TY - BOOK TI - Molecular and Biochemical Toxicology A3 - Smart, Robert C. A3 - Hodgson, Ernest DA - 2008/8/1/ PY - 2008/8/1/ DO - 10.1002/9780470285251 PB - John Wiley & Sons, Inc. SN - 9780470285251 9780470102114 UR - http://dx.doi.org/10.1002/9780470285251 ER - TY - CHAP TI - Carcinogenesis AU - Smart, Robert C. AU - Ewing, Sarah J. AU - Loomis, Kari D. T2 - Molecular and Biochemical Toxicology AB - This chapter contains sections titled: Introduction and Historical Perspective Human Cancer Categorization of Agents Associated with Carcinogenesis Somatic Mutation Theory Epigenetic Mechanism of Tumorigenesis Multistage Tumorigenesis Tumor Viruses Cellular Oncogenes Tumor Suppressor Genes Mutator Phenotype/DNA Stability Genes Interactions of Oncogenes and Tumor Suppressor Genes Genetically Modified Mouse Models Conclusions Suggested Reading PY - 2008/7/11/ DO - 10.1002/9780470285251.ch24 ET - 4th SP - 537–586 PB - John Wiley & Sons, Inc. UR - http://dx.doi.org/10.1002/9780470285251.ch24 ER - TY - CHAP TI - Overview of Molecular Techniques in Toxicology: Genes and Transgenes AU - Smart, Robert C. T2 - Molecular and Biochemical Toxicology A2 - Smart, R.C. A2 - Hodgson, E. AB - This chapter contains sections titled: Applicability of Molecular Techniques to Toxicology Overview of the Genetic Code and Flow of Genetic Information Molecular Cloning Southern and Northern Blot Analyses Polymerase Chain Reaction (PCR) Some Methods to Evaluate Gene Expression and Regulation Methods to Evaluate Gene Function Suggested Reading PY - 2008/7/11/ DO - 10.1002/9780470285251.ch2 ET - 4th SP - 5-24 PB - John Wiley & Sons, Inc. UR - http://dx.doi.org/10.1002/9780470285251.ch2 ER - TY - CHAP TI - Molecular and Biochemical Toxicology: Definition and Scope AU - Hodgson, Ernest AU - Smart, Robert C. T2 - Molecular and Biochemical Toxicology A2 - Smart, R.C. A2 - Hodgson, E. AB - This chapter contains sections titled: Introduction Toxicology Biochemical Toxicology Cellular Toxicology Molecular Toxicology Proteomics and Metabolomics Conclusions Suggested Reading PY - 2008/7/11/ DO - 10.1002/9780470285251.ch1 ET - 4th SP - 1-4 PB - John Wiley & Sons, Inc. UR - http://dx.doi.org/10.1002/9780470285251.ch1 ER - TY - CONF TI - Duration of infection and efficacy of doxycycline treatment in dogs experimentally co-infected with anaplasma platys and ehrlichia canis AU - Beall, M.J. AU - Gaunt, S.D. AU - Chandrashekar, R. AU - DeBisceglie, K AU - Thatcher, B AU - Diniz, PPVP AU - Breitschwerdt, EB T2 - 26th Annual American College of Veterinary Internal Medicine Forum C2 - 2008/// C3 - Journal of Veterinary Internal Medicine CY - San Antonio, TX DA - 2008/// PY - 2008/6/4/ VL - 22 SP - 703 M1 - 3 ER - TY - CONF TI - Atovaquone and azithromycin for the treatment of cytauxzoon felis AU - Birkenheuer, A.J. AU - Cohn, L.A. AU - Levy, M.G. AU - Breitschwerdt, E.B. AU - Marr, H.S. T2 - 26th Annual American College of Veterinary Internal Medicine Forum C2 - 2008/// C3 - Journal of Veterinary Internal Medicine CY - San Antonio, TX DA - 2008/// PY - 2008/6/4/ VL - 22 SP - 703–704 M1 - 3 ER - TY - CONF TI - Experimental E-canis infection in dogs: Antibody responses pre- and post-treatment AU - Stillman, B.A. AU - Caterina-DeBisceglie, K. AU - Bradley, J. AU - Saucier, J AU - Breitschwerdt, EB AU - Gaunt, SD AU - O'Connor, T AU - Chandrashekar, R T2 - 26th Annual American College of Veterinary Internal Medicine Forum C2 - 2008/// C3 - Journal of Veterinary Internal Medicine CY - San Antonio, TX DA - 2008/// PY - 2008/6/4/ VL - 22 SP - 781–782 M1 - 3 ER - TY - CONF TI - Dynamics of exposure to vector-borne organisms in dogs in North America: 2004-2006 AU - Diniz, P.P.V.P. AU - Morgado, M AU - Hegarty, BC AU - Cherry, N AU - Sullivan, M AU - Breitschwerdt, EB T2 - 26th Annual American College of Veterinary Internal Medicine Forum C2 - 2008/// C3 - Journal of Veterinary Internal Medicine CY - San Antonio, TX DA - 2008/// PY - 2008/6/4/ VL - 22 SP - 784 M1 - 3 ER - TY - JOUR TI - Surveillance for Zoonotic Vector-Borne Infections Using Sick Dogs from Southeastern Brazil AU - Diniz, Pedro Paulo Vissotto de Paiva AU - Schwartz, Denise Saretta AU - de Morais, Helio Silva Autran AU - Breitschwerdt, Edward Bealmear T2 - Vector-Borne and Zoonotic Diseases AB - For many vector-borne organisms, dogs can be used as sentinels to estimate the risk of human infection. The objective of this study was to use dogs as sentinels for multiple vector-borne organisms in order to evaluate the potential for human infection with these agents in southeastern Brazil. Blood from 198 sick dogs with clinicopathological abnormalities consistent with tick-borne infections were selected at the São Paulo State University Veterinary Teaching Hospital in Botucatu and tested for DNA and/or antibodies against specific vector-borne pathogens. At least one organism was detected in 88% of the dogs, and Ehrlichia canis DNA was amplified from 78% of the blood samples. Bartonella spp. seroreactivity was found in 3.6%. Leishmania chagasi antibodies were detected in 1% of the dogs. There was no serological or polymerase chain reaction evidence of infection with Anaplasma phagocytophilum, Borrelia burgdorferi, Ehrlichia chaffeensis, Ehrlichia ewingii, and Rickettsia rickettsii. The full E. canis 16S rRNA gene sequence of one of the Brazilian strains obtained in this study was identical to the causative agent of human ehrlichiosis in Venezuela. Ehrlichia canis may pose a human health hazard and may be undiagnosed in southeastern Brazil, whereas exposure to the other organisms examined in this study is presumably infrequent. DA - 2008/1/2/ PY - 2008/1/2/ DO - 10.1089/vbz.2007.0129 VL - 7 IS - 4 SP - 689–698 SN - 1530-3667 1557-7759 UR - http://dx.doi.org/10.1089/vbz.2007.0129 ER - TY - JOUR TI - Molecular Characterization of Bartonella vinsonii subsp. berkhoffii Genotype III AU - Cadenas, M. B. AU - Bradley, J. AU - Maggi, R. G. AU - Takara, M. AU - Hegarty, B. C. AU - Breitschwerdt, E. B. T2 - Journal of Clinical Microbiology AB - ABSTRACT The molecular characterization of a Bartonella vinsonii subsp. berkhoffii genotype III strain (NCSU strain 06-CO1) isolated from the blood of a military working dog diagnosed with endocarditis is reported in this study. Several genes were amplified and sequenced for comparative sequence similarity with other strains. DA - 2008/3/26/ PY - 2008/3/26/ DO - 10.1128/JCM.02456-07 VL - 46 IS - 5 SP - 1858-1860 J2 - Journal of Clinical Microbiology LA - en OP - SN - 0095-1137 UR - http://dx.doi.org/10.1128/JCM.02456-07 DB - Crossref ER - TY - JOUR TI - 237 Reovirus inhibits interferon signaling through a novel mechanism involving nuclear accumulation of IRF9 AU - Sherry, Barbara AU - Zurney, Jennifer AU - Kobayashi, Takeshi AU - Holm, Geoffrey H. AU - Dermody, Terence S. T2 - Cytokine DA - 2008/9// PY - 2008/9// DO - 10.1016/j.cyto.2008.07.302 VL - 43 IS - 3 SP - 296 J2 - Cytokine LA - en OP - SN - 1043-4666 UR - http://dx.doi.org/10.1016/j.cyto.2008.07.302 DB - Crossref ER - TY - JOUR TI - SILIP: A novel stable isotope labeling method for in planta quantitative proteomic analysis AU - Schaff, J.E. AU - Mbeunkui, F. AU - Blackburn, K. AU - Bird, D.McK. AU - Goshe, M.B. T2 - Plant Journal AB - Due to ease of manipulation, metabolic isotope coding of samples for proteomic analysis is typically performed in cell culture, thus preventing an accurate in vivo quantitative analysis, which is only achievable in intact organisms. To address this issue in plant biology, we developed SILIP (stable isotope labeling in planta) using tomato plants (Solanum lycopersicum cv. Rutgers) as a method that allows soil-grown plants to be efficiently labeled using a 14N/15N isotope coding strategy. After 2 months of growth on 14N- and 15N-enriched nitrogen sources, proteins were extracted from four distinct tomato tissues (roots, stems, leaves and flowers), digested, and analyzed by LC/MS/MS (data-dependent acquisition, DDA) and alternating low- and elevated-energy MS scans (data-independent acquisition, MS(E)). Using a derived relationship to generate a theoretical standard curve, the measured ratio of the M (monoisotopic) and M-1 isotopologues of 70 identified 15N-labeled peptides from 16 different proteins indicated that 15N incorporation was almost 99%, which is in excellent agreement with the 99.3% 15N-enriched nitrate used in the soil-based medium. Values for the various tissues ranged from 98.2 +/- 0.3% 15N incorporation in leaves to 98.8 2 +/- 0.2% in stems, demonstrating uniform labeling throughout the plant. In addition, SILIP is compatible with root-knot nematode (Meloidogyne spp.) development, and thus provides a new quantitative proteomics tool to study both plant and plant-microorganism systems. DA - 2008/// PY - 2008/// DO - 10.1111/j.1365-313X.2008.03639.x VL - 56 IS - 5 SP - 840-854 UR - http://www.scopus.com/inward/record.url?eid=2-s2.0-57649093858&partnerID=MN8TOARS KW - plant proteomics KW - quantitative analysis KW - mass spectrometry KW - liquid chromatography KW - isotope-coded KW - isotope ratio ER - TY - JOUR TI - Pioneering women in plant pathology AU - Ristaino, Jean Beagle DA - 2008/// PY - 2008/// ER - TY - CONF TI - Suppression of Fungal Pathogen Phytophthora capsici by Mycophagous Soil Fauna. AU - Qi, Rende AU - Tu, Cong AU - Shew, H David AU - Louws, Frank AU - Zhang, Yong AU - Ristaino, Jean AU - Hu, Shuijin C2 - 2008/// C3 - The 2008 Joint Annual Meeting DA - 2008/// ER - TY - JOUR TI - Phytophthora infestans identified in archival potato tubers from trials at Rothamsted, 1876-1879 AU - Ristaino, JB AU - Hu, CH AU - Fitt, Bruce DL T2 - Journal of Plant Pathology: an international journal of the Italian Phytopathological Society DA - 2008/// PY - 2008/// ER - TY - CONF TI - Major contributions of early women plant pathologists to our science: Strategies, struggles, and success AU - Ristaino, J T2 - AMER PHYTOPATHOLOGICAL SOC 3340 PILOT KNOB ROAD, ST PAUL, MN 55121 USA C2 - 2008/// C3 - PHYTOPATHOLOGY DA - 2008/// VL - 98 SP - S6-S6 M1 - 6 ER - TY - JOUR TI - Grace Marion Waterhouse AU - Brady, BL AU - Stamps, DJ AU - Ristaino, Jean Beagle T2 - Pioneering Women in Plant Pathology DA - 2008/// PY - 2008/// SP - 143 ER - TY - CONF TI - Genetic structure of populations of the tobacco blue mold pathogen, Peronospora tabacina in North America, Central America and the Caribbean and Europe AU - Blanco-Meneses, M AU - Carbone, I AU - Ivors, K AU - Ristaino, JB T2 - AMER PHYTOPATHOLOGICAL SOC 3340 PILOT KNOB ROAD, ST PAUL, MN 55121 USA C2 - 2008/// C3 - Phytopathology DA - 2008/// VL - 98 SP - S23-S23 M1 - 6 ER - TY - CONF TI - Gene flow of Phytophthora infestans between organic and conventional potato field in Southern Flevoland, The Netherlands AU - Hu, C AU - Govers, F AU - Ristaino, J C2 - 2008/// C3 - APS Centennial Meeting, Minneapolis, Minnesota, USA, 26-30 July 2008 DA - 2008/// VL - 98 SP - S69-S69 ER - TY - CONF TI - DNA sequence analysis of the late-blight pathogen gives clues to the world-wide migration AU - Ristaino, Jean Beagle C2 - 2008/// C3 - III International Late Blight Conference 834 DA - 2008/// SP - 27-40 ER - TY - JOUR TI - Preserving accuracy in GenBank T2 - Science DA - 2008/// PY - 2008/// VL - 319 IS - 5870 UR - http://www.scopus.com/inward/record.url?eid=2-s2.0-84881085358&partnerID=MN8TOARS ER - TY - CONF TI - Expressing a fraction of two determinants as a determinant AU - Kaltofen, Erich AU - Koiran, Pascal T2 - the twenty-first international symposium AB - Suppose the polynomials f and g in K[x1,...,xr] over the field K are determinants of non-singular m x m and n x n matrices, respectively, whose entries are in K ∪ x1,...,xr. Furthermore, suppose h = f/g is a polynomial in K[x1,..., xr]. We construct an s x s matrix C whose entries are in K ∪ x1,...,xr, such that h = det(C) and s = γ (m+n)6, where γ = O(1) if K is an infinite field or if for the finite field K = F{q} with q elements we have m = O(q), and where γ = (logq m)1+o(1) if q = o(m). Our construction utilizes the notion of skew circuits by Toda and WSK circuits by Malod and Portier. Our problem was motivated by resultant formulas derived from Chow forms. C2 - 2008/// C3 - Proceedings of the twenty-first international symposium on Symbolic and algebraic computation - ISSAC '08 DA - 2008/// DO - 10.1145/1390768.1390790 PB - ACM Press SN - 9781595939043 UR - http://dx.doi.org/10.1145/1390768.1390790 DB - Crossref ER - TY - CONF TI - Exact certification of global optimality of approximate factorizations via rationalizing sums-of-squares with floating point scalars AU - Kaltofen, Erich AU - Li, Bin AU - Yang, Zhengfeng AU - Zhi, Lihong T2 - the twenty-first international symposium AB - We generalize the technique by Peyrl and Parillo [Proc. SNC 2007] to computing lower bound certificates for several well-known factorization problems in hybrid symbolic-numeric computation. The idea is to transform a numerical sum-of-squares (SOS) representation of a positive polynomial into an exact rational identity. Our algorithms successfully certify accurate rational lower bounds near the irrational global optima for benchmark approximate polynomial greatest common divisors and multivariate polynomial irreducibility radii from the literature, and factor coefficient bounds in the setting of a model problem by Rump (up to n = 14, factor degree = 13. C2 - 2008/// C3 - Proceedings of the twenty-first international symposium on Symbolic and algebraic computation - ISSAC '08 DA - 2008/// DO - 10.1145/1390768.1390792 PB - ACM Press SN - 9781595939043 UR - http://dx.doi.org/10.1145/1390768.1390792 DB - Crossref ER - TY - JOUR TI - Statistical Methods for Mapping Multiple QTL AU - Zou, Wei AU - Zeng, Zhao-Bang T2 - International Journal of Plant Genomics AB - Since Lander and Botstein proposed the interval mapping method for QTL mapping data analysis in 1989, tremendous progress has been made in the last many years to advance new and powerful statistical methods for QTL analysis. Recent research progress has been focused on statistical methods and issues for mapping multiple QTL together. In this article, we review this progress. We focus the discussion on the statistical methods for mapping multiple QTL by maximum likelihood and Bayesian methods and also on determining appropriate thresholds for the analysis. DA - 2008/// PY - 2008/// DO - 10.1155/2008/286561 VL - 2008 SP - 1-8 J2 - International Journal of Plant Genomics LA - en OP - SN - 1687-5370 1687-5389 UR - http://dx.doi.org/10.1155/2008/286561 DB - Crossref ER - TY - JOUR TI - Epigenomic Consequences of Immortalized Plant Cell Suspension Culture AU - Tanurdzic, Milos AU - Vaughn, Matthew W AU - Jiang, Hongmei AU - Lee, Tae-Jin AU - Slotkin, R. Keith AU - Sosinski, Bryon AU - Thompson, William F AU - Doerge, R. W AU - Martienssen, Robert A T2 - PLoS Biology AB - Plant cells grown in culture exhibit genetic and epigenetic instability. Using a combination of chromatin immunoprecipitation and DNA methylation profiling on tiling microarrays, we have mapped the location and abundance of histone and DNA modifications in a continuously proliferating, dedifferentiated cell suspension culture of Arabidopsis. We have found that euchromatin becomes hypermethylated in culture and that a small percentage of the hypermethylated genes become associated with heterochromatic marks. In contrast, the heterochromatin undergoes dramatic and very precise DNA hypomethylation with transcriptional activation of specific transposable elements (TEs) in culture. High throughput sequencing of small interfering RNA (siRNA) revealed that TEs activated in culture have increased levels of 21-nucleotide (nt) siRNA, sometimes at the expense of the 24-nt siRNA class. In contrast, TEs that remain silent, which match the predominant 24-nt siRNA class, do not change significantly in their siRNA profiles. These results implicate RNA interference and chromatin modification in epigenetic restructuring of the genome following the activation of TEs in immortalized cell culture. DA - 2008/12/9/ PY - 2008/12/9/ DO - 10.1371/journal.pbio.0060302 VL - 6 IS - 12 SP - e302 J2 - PLoS Biol LA - en OP - SN - 1545-7885 UR - http://dx.doi.org/10.1371/journal.pbio.0060302 DB - Crossref ER - TY - PAT TI - Refined routes to chlorin building blocks AU - Lindsey, J. S. AU - Taniguchi, M. AU - Ra, D. AU - Mo, G. AU - Balasubramanian, T. C2 - 2008/// DA - 2008/// PY - 2008/// ER - TY - PAT TI - Procedure for preparing redox-active polymers on surfaces AU - Bocian, D. F. AU - Liu, Z AU - Lindsey, J. S. C2 - 2008/// DA - 2008/// PY - 2008/// ER - TY - PAT TI - Methods and compositions for protein production in tobacco plants with reduced nicotine AU - Conkling, M. A. AU - Song, W. AU - Mendu, N. C2 - 2008/// DA - 2008/// PY - 2008/// ER - TY - PAT TI - Boron complexation strategy for use in manipulating 1-acyldipyrromethanes AU - Lindsey, J. S. AU - Muthukumaran, K. AU - Ptaszek, M. AU - Huma, H. Z. S. C2 - 2008/// DA - 2008/// PY - 2008/// ER - TY - JOUR TI - Prediction of wood density breeding values of Pinus taeda elite parents from unbalanced data: A method for adjustment of site and age effects using common checklots AU - Isik, Fikret AU - Li, Bailian AU - Goldfarb, Barry AU - McKeand, Steve T2 - ANNALS OF FOREST SCIENCE AB - • Wood density of elite parents of loblolly pine (Pinus taeda L.) was investigated in 6 to 18 year-old progeny trials. The sampling was carried out separately in seven testing regions in the southeastern US. A checklot was the only connection between elite parents planted at different trials in a testing region. • We used a data normalization method suggested for unbalanced designs in cDNA microarray experiments to remove confounding site and age effects using the checklot as a reference sample. Wood density breeding values of parents were predicted by fitting a linear mixed model to the normalized data. • Using the reference samples to remove site and age effects appears to be an effective method for analysis of unbalanced progeny tests data. In general, wood density (kg/m3) decreased from coastal to inland plantings and from the southern to the northern planting. Considerable genetic variation for wood density was detected among these fast-growing elite parents in six of seven testing regions, with half-sib family mean heritabilities ranging from 0.71 to 0.97 within a testing region. With the exception of two regions, checklots were stable across trials in a region, based on regressing the checklot means on trial means. DA - 2008/6// PY - 2008/6// DO - 10.1051/forest:2008018 VL - 65 IS - 4 SP - SN - 1286-4560 KW - loblolly pine KW - reference sample KW - data normalization KW - genetic variation KW - heritability ER - TY - JOUR TI - Analysis of cellulose microfibril angle using a linear mixed model in Pinus taeda clones AU - Isik, Fikret AU - Gumpertz, Marcia AU - Li, Bailian AU - Goldfarb, Barry AU - Sun, Xuan T2 - CANADIAN JOURNAL OF FOREST RESEARCH AB - Variation in microfibril angle (MFA) (degrees) among loblolly pine ( Pinus taeda L.) full-sib families and clones was investigated using 43 clones from nine full-sib crosses tested at two locations. When the experiments were 12 years old, a total of 316 trees were drilled and 12 mm thick wood increment cores were collected. MFA for each growth ring in the wood core was measured using the SilviScan-2 tool. A quadratic mixed model was fitted to evaluate the MFA variation over different rings. Among the error covariance structures tested in the model, autoregressive order 1 was the best model for producing MFA estimates with the smallest errors. Estimated MFA was about 33° in the pith (ring 1) of the trees and decreased to 18° in the outer wood (ring 11). Full-sib crosses and clones within crosses explained about 12.5% of the total phenotypic variation. Repeatability of full-sib family means (H 2 f = 0.46) was moderate but repeatability of clone means was high (H 2 c = 0.79). Although it is possible to improve (decrease) MFA with recurrent selection in tree improvement programs to improve lumber quality, cost efficient and rapid methods for measuring MFA are needed. DA - 2008/6// PY - 2008/6// DO - 10.1139/X08-010 VL - 38 IS - 6 SP - 1676-1689 SN - 1208-6037 ER - TY - JOUR TI - An evaluation of selection for volume growth in loblolly pine AU - Sherrill, J. R. AU - Mullin, T. J. AU - Bullock, B. P. AU - McKeand, S. E. AU - Purnell, R. C. AU - Gumpertz, M. L. AU - Isik, F. T2 - SILVAE GENETICA AB - Abstract Total inside-bark volume is the most important selection criterion for productivity in tree breeding programs in the Southeastern U.S. Tree breeders typically estimate total inside-bark volume based on outside-bark diameter at breast height and total height without accounting for stem taper or bark thickness. To make a direct determination of total inside- and outside-bark volume, a loblolly pine (Pinus taeda L.) open-pollinated family trial replicated with cultural treatments of weed control and fertilization was measured. This direct measurement was compared to typical volume estimates. In this trial, approximately 40 individuals from each of 25 open-pollinated first- and second-generation families were destructively sampled in the 13 th growing season. Selection for volume using a combined-variable (diameter 2 * height) equation was found to be highly effective for making volume gain. There was a high correlation between estimated and directly-measured total inside-bark volumes (0.99). Bark thickness and stem taper had low importance for stem volume selection. There was a positive genetic correlation between bark thickness and diameter at breast height (0.66). This indicates that selection for larger diameters may produce individuals with thicker bark, which may eventually affect total inside-bark volume estimates. DA - 2008/// PY - 2008/// DO - 10.1515/sg-2008-0004 VL - 57 IS - 1 SP - 22-28 SN - 2509-8934 KW - bark thickness KW - genotype by treatment interaction KW - stem taper KW - volume gain KW - Pinus taeda L. KW - stem volume KW - tree improvement KW - stem form ER - TY - JOUR TI - Construction and characterization of mutant Dengue2 virus vaccine candidates displaying a host-range phenotype AU - Smith, K. M. AU - Nanda, K. AU - Slominski, C. J. AU - Hernandez, R. AU - Brown, D. T. AU - Thomas, M. E. T2 - Vaccine DA - 2008/// PY - 2008/// ER - TY - JOUR TI - Preliminary findings: analysis of carbon storage in Fraser fir plantations AU - Furiness, C. AU - Frampton, J. T2 - Limbs & Needles DA - 2008/// PY - 2008/// VL - 35 IS - 1 SP - 22 ER - TY - JOUR TI - Determination of Ground-State Hole-Transfer Rates Between Equivalent Sites in Oxidized Multiporphyrin Arrays Using Time-Resolved Optical Spectroscopy AU - Song, Hee-eun AU - Kirmaier, Christine AU - Taniguchi, Masahiko AU - Diers, James R. AU - Bocian, David F. AU - Lindsey, Jonathan S. AU - Holten, Dewey T2 - JOURNAL OF THE AMERICAN CHEMICAL SOCIETY AB - Excited-state charge separation in molecular architectures has been widely explored, yet ground-state hole (or electron) transfer, particularly involving equivalent pigments, has been far less studied, and direct quantitation of the rate of transfer often has proved difficult. Prior studies of ground-state hole transfer between equivalent zinc porphyrins using electron paramagnetic resonance techniques give a lower limit of approximately (50 ns)(-1) on the rates. Related transient optical studies of hole transfer between inequivalent sites [zinc porphyrin (Zn) and free base porphyrin (Fb)] give an upper limit of approximately (20 ps)(-1). Thus, a substantial window remains for the unknown rates of ground-state hole transfer between equivalent sites. Herein, the ground-state hole-transfer processes are probed in a series of oxidized porphyrin triads (ZnZnFb) with the focus being on determination of the rates between the nominally equivalent sites (Zn/Zn). The strategy builds upon recent time-resolved optical studies of the photodynamics of dyads wherein a zinc porphyrin is electrochemically oxidized and the attached free base porphyrin is photoexcited. The resulting energy- and hole-transfer processes in the oxidized ZnFb dyads are typically complete within 100 ps of excitation. Such processes are also present in the triads and serve as a starting point for determining the rates of ground-state hole transfer between equivalent sites in the triads. The rate constant of the Zn/Zn hole transfer is found to be (0.8 ns)(-1) for diphenylethyne-linked zinc porphyrins and increases only slightly to (0.6 ns)(-1) when a shorter phenylene linker is utilized. The rate decreases slightly to (1.1 ns)(-1) when steric constraints are introduced in the diarylethyne linker. In general, the rate constants for ground-state Zn/Zn hole transfer in oxidized arrays are a factor of 40 slower than those for Zn/Fb transfer. Collectively, the findings should aid the design of next-generation molecular architectures for applications in solar-energy conversion. DA - 2008/11/19/ PY - 2008/11/19/ DO - 10.1021/ja805673m VL - 130 IS - 46 SP - 15636-15648 SN - 0002-7863 ER - TY - JOUR TI - Tackling the characterization of canine chromosomal breakpoints with an integrated in-situ/in-silico approach: The canine PAR and PAB AU - Young, Andrea C. AU - Kirkness, Ewen F. AU - Breen, Matthew T2 - CHROMOSOME RESEARCH DA - 2008/12// PY - 2008/12// DO - 10.1007/s10577-008-1268-9 VL - 16 IS - 8 SP - 1193-1202 SN - 1573-6849 KW - Canis familiaris KW - chromosomal breakpoints KW - dog KW - PAR KW - PAB ER - TY - JOUR TI - Photophysical characterization of imidazolium-substituted Pd(II), In(III), and Zn(II) porphyrins as photosensitizers for photodynamic therapy AU - Kee, Hooi Ling AU - Bhaumik, Jayeeta AU - Diers, James R. AU - Mroz, Pawel AU - Hamblin, Michael R. AU - Bocian, David F. AU - Lindsey, Jonathan S. AU - Holten, Dewey T2 - JOURNAL OF PHOTOCHEMISTRY AND PHOTOBIOLOGY A-CHEMISTRY AB - The photophysical properties of four imidazolium-substituted metalloporphyrins have been assessed to gain insights into the relative efficacy of the compounds for photodynamic therapy (PDT). A set of zinc(II), palladium(II), and chloro-indium(III) porphyrins all bear a net positive charge owing to the diethylimidazolium unit; one zinc chelate bears a negative charge owing to a bis(sulfobutyl)imidazolium unit. The photophysical properties of the cationic and anionic zinc porphyrins are very similar to one another in organic solvents, phosphate-buffered saline, and in the presence of bovine serum albumin. The properties of the zinc and palladium porphyrins bearing charged peripheral groups are generally similar to those of neutral analogs in organic solvents. The palladium porphyrin shows an essentially quantitative yield (≥0.99) of the triplet excited state compared to the zinc porphyrins (∼0.9), and all are quantitatively quenched (at the diffusion limit) by molecular oxygen in air-saturated fluid solution. If the rate constant and yield of quenching of the triplet excited state by energy or electron transfer to molecular oxygen is the same in the cellular environment as in solution, then these processes combined with the triplet yield contribute only a factor of 1.3 to the higher PDT activity of analogous palladium versus zinc porphyrins, which is much smaller than what is observed. Therefore, other factors such as transient reduction of the excited porphyrin or delivery to the target site must predominantly underlie the difference in PDT efficacy of these sensitizers. DA - 2008/12/15/ PY - 2008/12/15/ DO - 10.1016/j.jphotochem.2008.08.006 VL - 200 IS - 2-3 SP - 346-355 SN - 1873-2666 KW - Photodynamic therapy KW - Porphyrin KW - Photosensitizer KW - Imidazolium KW - Photophysics ER - TY - JOUR TI - Epigenomic consequences of immortalized plant cell suspension culture AU - Tanurdzic, M. AU - Vaughn, M. W. AU - Jiang, H. AU - Lee, T. J. AU - Slotkin, R. K. AU - Sosinski, B. AU - Thompson, W. F. AU - Doerge, R. W. AU - Martienssen, R. A. T2 - PLoS Biology DA - 2008/// PY - 2008/// VL - 6 IS - 12 SP - 2880-2895 ER - TY - JOUR TI - A genome assembly-integrated dog 1 Mb BAC microarray: a cytogenetic resource for canine cancer studies and comparative genomic analysis AU - Thomas, R. AU - Duke, S. E. AU - Karlsson, E. K. AU - Evans, A. AU - Ellis, P. AU - Lindblad-Toh, K. AU - Langford, C. F. AU - Breen, M. T2 - CYTOGENETIC AND GENOME RESEARCH AB - Molecular cytogenetic studies have been instrumental in defining the nature of numerical and structural chromosome changes in human cancers, but their significance remains to be fully understood. The emergence of high quality genome assemblies for several model organisms provides exciting opportunities to develop novel genome-integrated molecular cytogenetic resources that now permit a comparative approach to evaluating the relevance of tumor-associated chromosome aberrations, both within and between species. We have used the dog genome sequence assembly to identify a framework panel of 2,097 bacterial artificial chromosome (BAC) clones, selected at intervals of approximately one megabase. Each clone has been evaluated by multicolor fluorescence in situ hybridization (FISH) to confirm its unique cytogenetic location in concordance with its reported position in the genome assembly, providing new information on the organization of the dog genome. This panel of BAC clones also represents a powerful cytogenetic resource with numerous potential applications. We have used the clone set to develop a genome-wide microarray for comparative genomic hybridization (aCGH) analysis, and demonstrate its application in detection of tumor-associated DNA copy number aberrations (CNAs) including single copy deletions and amplifications, regional aneuploidy and whole chromosome aneuploidy. We also show how individual clones selected from the BAC panel can be used as FISH probes in direct evaluation of tumor karyotypes, to verify and explore CNAs detected using aCGH analysis. This cytogenetically validated, genome integrated BAC clone panel has enormous potential for aiding gene discovery through a comparative approach to molecular oncology. DA - 2008/// PY - 2008/// DO - 10.1159/000163088 VL - 122 IS - 2 SP - 110-121 SN - 1424-859X ER - TY - JOUR TI - Resistance of Pinus taeda families under artificial inoculations with diverse fusiform rust pathogen populations and comparison with field trials (vol 38, pg 2687, 2008) AU - Isik, Fikret AU - Amerson, Henry V. AU - Whetten, Ross W. AU - Garcia, Saul A. AU - Li, Bailian AU - McKeand, Steven E. T2 - CANADIAN JOURNAL OF FOREST RESEARCH-REVUE CANADIENNE DE RECHERCHE FORESTIERE DA - 2008/12// PY - 2008/12// DO - 10.1139/x08-910 VL - 38 IS - 12 SP - 3151-3151 SN - 0045-5067 ER - TY - CONF TI - Seed orchard management strategies for deployment of intensively selected loblolly pine families in the southern US AU - McKeand, S. E. AU - Gerwig, D. M. AU - Cumbie, W. P. AU - Jett, J. B. C2 - 2008/// C3 - Seed orchards, Proceedings from a conference at Umea, Sweden DA - 2008/// SP - 177-182 SN - 978-91-85911-28-8 ER - TY - JOUR TI - Genetic parameter estimates for growth traits from diallel tests of loblolly pine throughout the southeastern United States AU - McKeand, S. E. AU - Li, B. AU - Grissom, J. E. AU - Isik, F. AU - Jayawickrama, K. J. S. T2 - SILVAE GENETICA AB - Abstract Variation in heritability and in genetic correlation estimates were evaluated for juvenile tree height and volume for six testing areas of loblolly pine (Pinus taeda L.) in the southeastern United States. Variance components and their functions (heritability and type B genetic correlations) were estimated from 265 six-parent disconnected diallel series, tested in almost 1000 trials (4 tests per diallel series). Original data were collected at age 6 years from about one million trees (265 diallel series x 30 crosses x 36 trees per cross/site x 4 sites) planted in field tests. Genetic tests were from the second cycle of breeding in the North Carolina State University - Industry Cooperative Tree Improvement Program. The overall unbiased individual-tree narrow-sense heritability for height was 0.19 and for volume was 0.16. The broad-sense heritabilities for height (0.24) and for volume (0.22) were higher than narrow-sense heritabilities due to the presence of non-additive genetic variance. There were moderate regional differences in these estimates, with tests in the Lower Gulf Coastal Plain tending to have the highest heritabilities for growth traits. There was very little association between site index and heritability, but heritabilities were higher on sites with the highest survival and highest test precision. Genotype x environment interactions were generally low both for half-sib and full-sib families, indicating that families can be operationally deployed to different sites with little concern about unpredictable performance. DA - 2008/// PY - 2008/// DO - 10.1515/sg-2008-0016 VL - 57 IS - 3 SP - 101-110 SN - 2509-8934 KW - genetic correlation KW - genetic gain KW - genotype x environment interaction KW - heritability KW - Pinus taeda L. ER - TY - JOUR TI - Infestation Rate of Hemlock Woolly Adelgid (Hemiptera: Adelgidae) Among Three North American Hemlock (Tsuga) Species Following Artificial Inoculation AU - Jetton, Robert M. AU - Hain, Fred P. AU - Dvorak, William S. AU - Frampton, John T2 - JOURNAL OF ENTOMOLOGICAL SCIENCE DA - 2008/10// PY - 2008/10// DO - 10.18474/0749-8004-43.4.438 VL - 43 IS - 4 SP - 438-442 SN - 0749-8004 KW - Adelges tsugae KW - Tsuga caroliniana KW - Tsuga canadensis KW - Tsuga heterophylla KW - host susceptibility ER - TY - JOUR TI - Transgenic Arabidopsis Plants Expressing the Type 1 Inositol 5-Phosphatase Exhibit Increased Drought Tolerance and Altered Abscisic Acid Signaling AU - Perera, Imara Y. AU - Hung, Chiu-Yueh AU - Moore, Candace D. AU - Stevenson-Paulik, Jill AU - Boss, Wendy F. T2 - PLANT CELL AB - The phosphoinositide pathway and inositol-1,4,5-trisphosphate (InsP(3)) are implicated in plant responses to stress. To determine the downstream consequences of altered InsP(3)-mediated signaling, we generated transgenic Arabidopsis thaliana plants expressing the mammalian type I inositol polyphosphate 5-phosphatase (InsP 5-ptase), which specifically hydrolyzes soluble inositol phosphates and terminates the signal. Rapid transient Ca(2+) responses to a cold or salt stimulus were reduced by approximately 30% in these transgenic plants. Drought stress studies revealed, surprisingly, that the InsP 5-ptase plants lost less water and exhibited increased drought tolerance. The onset of the drought stress was delayed in the transgenic plants, and abscisic acid (ABA) levels increased less than in the wild-type plants. Stomatal bioassays showed that transgenic guard cells were less responsive to the inhibition of opening by ABA but showed an increased sensitivity to ABA-induced closure. Transcript profiling revealed that the drought-inducible ABA-independent transcription factor DREB2A and a subset of DREB2A-regulated genes were basally upregulated in the InsP 5-ptase plants, suggesting that InsP(3) is a negative regulator of these DREB2A-regulated genes. These results indicate that the drought tolerance of the InsP 5-ptase plants is mediated in part via a DREB2A-dependent pathway and that constitutive dampening of the InsP(3) signal reveals unanticipated interconnections between signaling pathways. DA - 2008/10// PY - 2008/10// DO - 10.1105/tpc.108.061374 VL - 20 IS - 10 SP - 2876-2893 SN - 1040-4651 ER - TY - JOUR TI - BARTONELLA HENSELAE IN CAPTIVE AND HUNTER-HARVESTED BELUGA (DELPHINAPTERUS LEUCAS) AU - Maggi, Ricardo G. AU - Raverty, Stephen A. AU - Lester, Sally J. AU - Huff, David G. AU - Haulena, Martin AU - Ford, Susan L. AU - Nielsen, Ole AU - Robinson, John H. AU - Breitschwerdt, Edward B. T2 - JOURNAL OF WILDLIFE DISEASES AB - Previously, we reported the isolation of Bartonella henselae from the blood of harbor porpoises (Phocoena phocoena) and loggerhead sea turtles (Caretta caretta) from the North Carolina coast. Hematologic, pathologic, and microbiologic findings surrounding the death of a juvenile captive beluga in Vancouver initiated an outbreak investigation designed to define the molecular prevalence of Bartonella infection in belugas. Using polymerase chain reaction analyses targeting the intergenic spacer region (ITS), two B. henselae ITS strains were identified in 78% of captive and free-ranging hunter-harvested belugas. These findings may have public health implications and may influence aquarium management procedures for captive marine mammals. DA - 2008/10// PY - 2008/10// DO - 10.7589/0090-3558-44.4.871 VL - 44 IS - 4 SP - 871-877 SN - 1943-3700 KW - Bartonella KW - beluga KW - captive KW - Delphinapterus leucas KW - free-ranging ER - TY - JOUR TI - Mutations in a Delta(9)-Stearoyl-ACP-Desaturase Gene Are Associated with Enhanced Stearic Acid Levels in Soybean Seeds AU - Zhang, Ping AU - Burton, Joseph W. AU - Upchurch, Robert G. AU - Whittle, Edward AU - Shanklin, John AU - Dewey, Ralph E. T2 - CROP SCIENCE AB - Stearic acid (18:0) is typically a minor component of soybean [ Glycine max (L.) Merr.] oil, accounting for only 2 to 4% of the total fatty acid content. Increasing stearic acid levels of soybean oil would lead to enhanced oxidative stability, potentially reducing the need for hydrogenation, a process leading to the formation of undesirable trans fatty acids. Although mutagenesis strategies have been successful in developing soybean germplasm with elevated 18:0 levels in the seed oil, the specific gene mutations responsible for this phenotype were not known. We report a newly identified soybean gene, designated SACPD‐C , that encodes a unique isoform of Δ 9 –stearoyl‐ACP‐desaturase, the enzyme responsible for converting stearic acid to oleic acid (18:1). High levels of SACPD‐C transcript were only detected in developing seed tissue, suggesting that the encoded desaturase functions to enhance oleic acid biosynthetic capacity as the immature seed is actively engaged in triacylglycerol production and storage. The participation of SACPD‐C in storage triacylglycerol synthesis is further supported by the observation of mutations in this gene in two independent sources of elevated 18:0 soybean germplasm, A6 (30% 18:0) and FAM94‐41 (9% 18:0). A molecular marker diagnostic for the FAM94‐41 SACPD‐C gene mutation strictly associates with the elevated 18:0 phenotype in a segregating population, and could thus serve as a useful tool in the development of cultivars with oils possessing enhanced oxidative stability. DA - 2008/// PY - 2008/// DO - 10.2135/cropsci2008.02.0084 VL - 48 IS - 6 SP - 2305-2313 SN - 1435-0653 ER - TY - JOUR TI - Bartonella species detection in captive, stranded and free-ranging cetaceans AU - Harms, Craig A. AU - Maggi, Ricardo G. AU - Breitschwerdt, Edward B. AU - Clemons-Chevis, Connie L. AU - Solangi, Mobashir AU - Rotstein, David S. AU - Fair, Patricia A. AU - Hansen, Larry J. AU - Hohn, Aleta A. AU - Lovewell, Gretchen N. AU - McLellan, William A. AU - Pabst, D. Ann AU - Rowles, Teri K. AU - Schwacke, Lori H. AU - Townsend, Forrest I. AU - Wells, Randall S. T2 - Veterinary Research AB - We present prevalence of Bartonella spp. for multiple cohorts of wild and captive cetaceans. One hundred and six cetaceans including 86 bottlenose dolphins (71 free-ranging, 14 captive in a facility with a dolphin experiencing debility of unknown origin, 1 stranded), 11 striped dolphins, 4 harbor porpoises, 3 Risso's dolphins, 1 dwarf sperm whale and 1 pygmy sperm whale (all stranded) were sampled. Whole blood (n = 95 live animals) and tissues (n = 15 freshly dead animals) were screened by PCR (n = 106 animals), PCR of enrichment cultures (n = 50 animals), and subcultures (n = 50 animals). Bartonella spp. were detected from 17 cetaceans, including 12 by direct extraction PCR of blood or tissues, 6 by PCR of enrichment cultures, and 4 by subculture isolation. Bartonella spp. were more commonly detected from the captive (6/14, 43%) than from free-ranging (2/71, 2.8%) bottlenose dolphins, and were commonly detected from the stranded animals (9/21, 43%; 3/11 striped dolphins, 3/4 harbor porpoises, 2/3 Risso's dolphins, 1/1 pygmy sperm whale, 0/1 dwarf sperm whale, 0/1 bottlenose dolphin). Sequencing identified a Bartonella spp. most similar to B. henselae San Antonio 2 in eight cases (4 bottlenose dolphins, 2 striped dolphins, 2 harbor porpoises), B. henselae Houston 1 in three cases (2 Risso's dolphins, 1 harbor porpoise), and untyped in six cases (4 bottlenose dolphins, 1 striped dolphin, 1 pygmy sperm whale). Although disease causation has not been established, Bartonella species were detected more commonly from cetaceans that were overtly debilitated or were cohabiting in captivity with a debilitated animal than from free-ranging animals. The detection of Bartonella spp. from cetaceans may be of pathophysiological concern. DA - 2008/8/23/ PY - 2008/8/23/ DO - 10.1051/vetres:2008036 VL - 39 IS - 6 SP - 59 SN - 0928-4249 1297-9716 UR - http://dx.doi.org/10.1051/vetres:2008036 ER - TY - JOUR TI - The Red clover necrotic mosaic virus RNA-2 encoded movement protein is a second suppressor of RNA silencing AU - Powers, Jason G. AU - Sit, Tim L. AU - Heinsohn, Curtis AU - George, Carol G. AU - Kim, Kook-Hyung AU - Lommel, Steven A. T2 - VIROLOGY AB - The replication complex of Red clover necrotic mosaic virus (RCNMV) has been shown to possess silencing suppression activity. Here a newly developed viral-based assay for the identification of silencing suppression activity was used to provide evidence for a second, mechanistically distinct method of silencing suppression provided for by the RCNMV movement protein (MP). This new assay relies on Turnip crinkle virus with its capsid protein replaced with green fluorescent protein to act as a reporter (TCV-sGFP). In the presence of a protein with silencing suppression activity TCV-sGFP readily moves from cell-to-cell, but in the absence of such a protein TCV-sGFP is confined to small foci of infection. This TCV-sGFP assay was used to identify MP as a suppressor of RNA silencing, to delimit essential amino acids for this activity and uncouple silencing and movement functions. DA - 2008/11/25/ PY - 2008/11/25/ DO - 10.1016/j.virol.2008.09.004 VL - 381 IS - 2 SP - 277-286 SN - 0042-6822 KW - TCV-sGFP KW - Red clover necrotic mosaic virus KW - RCNMV KW - Movement protein KW - MP KW - VSR KW - RNA silencing ER - TY - JOUR TI - Quantitative Trait Loci Mapping and The Genetic Basis of Heterosis in Maize and Rice AU - Franco Garcia, Antonio Augusto AU - Wang, Shengchu AU - Melchinger, Albrecht E. AU - Zeng, Zhao-Bang T2 - GENETICS AB - Abstract Despite its importance to agriculture, the genetic basis of heterosis is still not well understood. The main competing hypotheses include dominance, overdominance, and epistasis. NC design III is an experimental design that has been used for estimating the average degree of dominance of quantitative trait loci (QTL) and also for studying heterosis. In this study, we first develop a multiple-interval mapping (MIM) model for design III that provides a platform to estimate the number, genomic positions, augmented additive and dominance effects, and epistatic interactions of QTL. The model can be used for parents with any generation of selfing. We apply the method to two data sets, one for maize and one for rice. Our results show that heterosis in maize is mainly due to dominant gene action, although overdominance of individual QTL could not completely be ruled out due to the mapping resolution and limitations of NC design III. For rice, the estimated QTL dominant effects could not explain the observed heterosis. There is evidence that additive × additive epistatic effects of QTL could be the main cause for the heterosis in rice. The difference in the genetic basis of heterosis seems to be related to open or self pollination of the two species. The MIM model for NC design III is implemented in Windows QTL Cartographer, a freely distributed software. DA - 2008/11// PY - 2008/11// DO - 10.1534/genetics.107.082867 VL - 180 IS - 3 SP - 1707-1724 SN - 1943-2631 ER - TY - JOUR TI - Design and synthesis of water-soluble bioconjugatable trans-AB-porphyrins AU - Muresan, Ana Z. AU - Lindsey, Jonathan S. T2 - TETRAHEDRON AB - Three free base porphyrins have been prepared that bear a polar and facially encumbering 2,4,6-tris-(carboxymethoxy)phenyl motif at one meso (5-) position. The only other substituent (15-position) comprises phenyl, formyl, or p-aminophenyl. The porphyrins exhibit solubility in water (or aqueous buffer solutions) at pH ≥7 and concentrations >1 mM at room temperature. The concise syntheses, water solubility, and bioconjugatable handle make these porphyrin constructs suitable for biological applications. DA - 2008/12/8/ PY - 2008/12/8/ DO - 10.1016/j.tet.2008.08.096 VL - 64 IS - 50 SP - 11440-11448 SN - 1464-5416 KW - Porphyrin KW - Water KW - Hydrocarbon KW - Water-soluble KW - Facial encumbrance KW - Bioconjugation ER - TY - JOUR TI - Coxiella-Like Infection in Psittacines and a Toucan AU - Shivaprasad, H. L. AU - Cadenas, M. B. AU - Diab, S. S. AU - Nordhausen, R. AU - Bradway, D. AU - Crespo, R. AU - Breitschwerdt, E. B. T2 - Avian Diseases AB - Seven psittacine birds and a toucan (Ramphastos toco) were diagnosed as infected with Coxiella-like bacteria, based on polymerase chain reaction and bacterial 16S rRNA gene sequence obtained from each bird's liver tissue. Most of the birds exhibited lethargy and weakness for several days prior to death. Gross lesions included mild to moderate emaciation and severely enlarged and mottled pale livers and spleens. Microscopically, there was multifocal necrosis of hepatocytes with infiltration of a mixed population of inflammatory cells, including lymphocytes, heterophils, plasma cells, and macrophages randomly scattered throughout in most birds. In several birds within the macrophages there were vacuoles containing basophilic small cocco-bacilli organisms measuring about 0.5-1 microm. The spleens had increased numbers of mononuclear phagocytic system cells, some of which had vacuoles that contained similar organisms, as observed in the liver. There was inflammation in the epicardium and endocardium, interstitium of the lungs, kidney, adrenal and thyroid glands, lamina propria of the intestine, and in occasional birds in the brain, bursa of Fabricius, and bone marrow associated with similar organisms in the macrophages. Transmission electron microscopy of the liver and lungs in most birds and in the thyroid glands of one bird revealed pleomorphic round to elongated bacteria measuring about 0.45 microm in diameter and more than 1.0 microm in length. Most of these organisms contained a peripheral zone of loosely arranged electron dense material that was located immediately beneath a trilaminar membrane. Occasional organisms contained nucleoids. This is the first documentation of disease presumptively associated with Coxiella-like bacteria in birds. DA - 2008/9// PY - 2008/9// DO - 10.1637/8192-120707-Reg VL - 52 IS - 3 SP - 426-432 J2 - Avian Diseases LA - en OP - SN - 0005-2086 1938-4351 UR - http://dx.doi.org/10.1637/8192-120707-Reg DB - Crossref KW - avian KW - psittacines KW - toucan KW - hepatitis KW - splenitis KW - Coxiella ER - TY - JOUR TI - A sequence-anchored genetic linkage map for the moss, Physcomitrella patens AU - Schaff, J. E. AU - Mbeunkui, F. AU - Blackburn, K. AU - Bird, D. M. AU - Goshe, M. B. T2 - Plant Journal DA - 2008/// PY - 2008/// VL - 56 IS - 5 SP - 840-854 ER - TY - JOUR TI - A Bipodal-Tethered Porphyrin for Attachment to Silicon Surfaces in Studies of Molecular Information Storage AU - Schmidt, Izabela AU - Jiao, Jieying AU - Bocian, David F. AU - Lindsey, Jonathan S. T2 - JOURNAL OF NANOSCIENCE AND NANOTECHNOLOGY AB - An approach for molecular information storage entails use of redox-active molecules attached to an electroactive surface. Understanding the structural features that give rise to robust molecular monolayers with high charge density is an essential objective. Toward this goal, a zinc-porphyrin bearing an all-carbon bipodal tether, 5-(1,6-heptadien-4-yl)-15-phenyl-10,20-di- p -tolylporphinatozinc(II), has been synthesized by the reaction of a dipyrromethane and a dipyrromethane-1,9-dicarbinol. The porphyrin was attached to Si(100) via a high temperature procedure. The resulting molecular monolayers exhibited surface coverages, adsorption geometries, electron-transfer rates, and stabilities under repeated electrochemical cycling that are similar to those obtained with monopodal carbon tethers. The observation that the bipodal anchor does not provide enhanced stability is surprising given that attachment is achieved via two versus one covalent linkage. DA - 2008/9// PY - 2008/9// DO - 10.1166/jnn.2008.IC85 VL - 8 IS - 9 SP - 4813-4817 SN - 1533-4899 KW - Porphyrin KW - Bipod KW - Silicon KW - Memory KW - DRAM ER - TY - JOUR TI - Using a bud volume index with the top-stop nipper to control leader growth of Fraser fir Christmas trees AU - Rutledge, M. E. AU - Frampton, J. AU - Hinesley, L. E. AU - Blank, G. T2 - HortTechnology DA - 2008/// PY - 2008/// VL - 18 IS - 4 SP - 583-587 ER - TY - JOUR TI - Site-specific evolutionary rates in proteins are better modeled as non-independent and strictly relative AU - Fernandes, Andrew D. AU - Atchley, William R. T2 - BIOINFORMATICS AB - In a nucleotide or amino acid sequence, not all sites evolve at the same rate, due to differing selective constraints at each site. Currently in computational molecular evolution, models incorporating rate heterogeneity always share two assumptions. First, the rate of evolution at each site is assumed to be independent of every other site. Second, the values of these rates are assumed to be drawn from a known prior distribution. Although often assumed to be small, the actual effect of these assumptions has not been previously quantified in the literature.Herein we describe an algorithm to simultaneously infer the set of n-1 relative rates that parameterize the likelihood of an n-site alignment. Unlike previous work (a) these relative rates are completely identifiable and distinct from the branch-length parameters, and (b) a far more general class of rate priors can be used, and their effects quantified. Although described in a Bayesian framework, we discuss a future maximum likelihood extension.Using both synthetic data and alignments from the Myc, Max and p53 protein families, we find that inferring relative rather than absolute rates has several advantages. First, both empirical likelihoods and Bayes factors show strong preference for the relative-rate model, with a mean Delta ln P=-0.458 per alignment site. Second, the computed likelihoods and Bayes factors were essentially independent of the relative-rate prior, indicating that good estimates of the posterior rate distribution are not required a priori. Third, a novel finding is that rates can be accurately inferred even when up to approximately 4 substitutions per site have occurred. Thus biologically relevant putative hypervariable sites can be identified as easily as conserved sites. Lastly, our model treats rates and tree branch-lengths as completely identifiable, allowing for the first time coherent simultaneous inference of branch-lengths and site-specific evolutionary rates.Source code for the utility described is available under a BSD-style license at http://www.fernandes.org/txp/article/9/site-specific-relative-evolutionary-rates. DA - 2008/10/1/ PY - 2008/10/1/ DO - 10.1093/bioinformatics/btn395 VL - 24 IS - 19 SP - 2177-2183 SN - 1460-2059 ER - TY - JOUR TI - Serological and Molecular Prevalence of Borrelia burgdorferi, Anaplasma phagocytophilum, and Ehrlichia Species in Dogs from Minnesota AU - Beall, Melissa J. AU - Chandrashekar, Ramaswamy AU - Eberts, Matthew D. AU - Cyr, Katie E. AU - Diniz, Pedro Paulo V.P. AU - Mainville, Celine AU - Hegarty, Barbara C. AU - Crawford, John M. AU - Breitschwerdt, Edward B. T2 - Vector-Borne and Zoonotic Diseases AB - A population of 731 naturally exposed pet dogs examined at a private practice in Baxter, Minnesota, an area endemic for Lyme disease and anaplasmosis, was tested by serological and molecular methods for evidence of exposure to or infection with selected vector-borne pathogens. Serum samples were tested by enzyme-linked immunosorbent assay (ELISA) for Anaplasma phagocytophilum, Borrelia burgdorferi, and Ehrlichia canis antibodies and for Dirofilaria immitis antigen. Blood samples from 273 dogs were also analyzed by polymerase chain reaction (PCR) for Anaplasma and Ehrlichia species DNA. Based on the owner history and the attending veterinarian's physical examination findings, dogs exhibiting illness compatible with anaplasmosis or borreliosis were considered clinical cases, and their results were compared to the healthy dog population. Antibodies to only A. phagocytophilum were detected in 217 (29%) dogs; to only B. burgdorferi, in 80 (11%) dogs; and seroreactivity to both organisms, in 188 (25%) dogs. Of 89 suspected cases of canine anaplasmosis or borreliosis, A. phagocytophilum or B. burgdorferi antibodies were detected in 22 dogs (25%) and 8 dogs (9%) respectively, whereas antibodies to both organisms were found in 38 dogs (43%). Ehrlichia canis antibodies and D. immitis antigen were each detected in 11 (1.5%) dogs. Anaplasma phagocytophilum DNA was amplified from 7 of 222 (3%) healthy dogs and 19 of 51 (37%) clinical cases. Seroreactivity to both A. phagocytophilum and B. burgdorferi was detected more frequently in suspected cases of anaplasmosis and/or borreliosis than seroreactivity to either organism alone. Based on PCR testing, A. phagocytophilum DNA was more prevalent in suspected cases of anaplasmosis or borreliosis than in healthy dogs from the same region. DA - 2008/8// PY - 2008/8// DO - 10.1089/vbz.2007.0236 VL - 8 IS - 4 SP - 455-464 J2 - Vector-Borne and Zoonotic Diseases LA - en OP - SN - 1530-3667 1557-7759 UR - http://dx.doi.org/10.1089/vbz.2007.0236 DB - Crossref KW - tick(s) ER - TY - JOUR TI - Sequence and genetic map of Meloidogyne hapla: A compact nematode genome for plant parasitism AU - Opperman, Charles H. AU - Bird, David M. AU - Williamson, Valerie M. AU - Rokhsar, Dan S. AU - Burke, Mark AU - Cohn, Jonathan AU - Cromer, John AU - Diener, Steve AU - Gajan, Jim AU - Graham, Steve AU - Houfek, T. D. AU - Liu, Qingli AU - Mitros, Therese AU - Schaff, Jennifer AU - Schaffer, Reenah AU - Scholl, Elizabeth AU - Sosinski, Bryon R. AU - Thomas, Varghese P. AU - Windham, Eric T2 - PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA AB - We have established Meloidogyne hapla as a tractable model plant-parasitic nematode amenable to forward and reverse genetics, and we present a complete genome sequence. At 54 Mbp, M. hapla represents not only the smallest nematode genome yet completed, but also the smallest metazoan, and defines a platform to elucidate mechanisms of parasitism by what is the largest uncontrolled group of plant pathogens worldwide. The M. hapla genome encodes significantly fewer genes than does the free-living nematode Caenorhabditis elegans (most notably through a reduction of odorant receptors and other gene families), yet it has acquired horizontally from other kingdoms numerous genes suspected to be involved in adaptations to parasitism. In some cases, amplification and tandem duplication have occurred with genes suspected of being acquired horizontally and involved in parasitism of plants. Although M. hapla and C. elegans diverged >500 million years ago, many developmental and biochemical pathways, including those for dauer formation and RNAi, are conserved. Although overall genome organization is not conserved, there are areas of microsynteny that may suggest a primary biological function in nematodes for those genes in these areas. This sequence and map represent a wealth of biological information on both the nature of nematode parasitism of plants and its evolution. DA - 2008/9/30/ PY - 2008/9/30/ DO - 10.1073/pnas.0805946105 VL - 105 IS - 39 SP - 14802-14807 SN - 1091-6490 UR - http://www.scopus.com/inward/record.url?eid=2-s2.0-54149092490&partnerID=MN8TOARS KW - compaction KW - dauer KW - development KW - horizontal gene transfer KW - gene ER - TY - JOUR TI - Phylogenetic relationships of Phytophthora andina, a new species from the highlands of Ecuador that is closely related to the Irish potato famine pathogen Phytophthora infestans AU - Gomez-Alpizar, Luis AU - Hu, Chia-Hui AU - Oliva, Ricardo AU - Forbes, Gregory AU - Ristaino, Jean Beagle T2 - MYCOLOGIA AB - Phylogenetic relationships of Phytophthora infestans sensu lato in the Andean highlands of South America were examined. Three clonal lineages (US-1, EC-1, EC-3) and one heterogeneous lineage (EC-2) were found in association with different host species in genus Solanum. The EC-2 lineage includes two mitochondrial (mtDNA) haplotypes, Ia and Ic. Isolates of P. infestans sensu lato EC-2 fit the morphological description of P. infestans but are different from any genotypes of P. infestans described to date. All isolates of P. infestans sensu lato from Ecuador were amplified by a P. infestans specific primer (PINF), and restriction fragment length patterns were identical in isolates amplified with ITS primers 4 and 5. The EC-1 clonal lineage of P. infestans sensu lato from S. andreanum, S. columbianum, S. paucijugum, S. phureja, S. regularifolium, S. tuberosum and S. tuquerense was confirmed to be P. infestans based on sequences of the cytochrome oxidase I (cox I) gene and intron 1 of ras gene. The EC-2 isolates with the Ic haplotype formed a distinct branch in the same clade with P. infestans and P. mirabilis, P. phaseoli and P. ipomoeae for both cox I and ras intron 1 phylogenies and were identified as the newly described species P. andina. Ras intron 1 sequence data suggests that P. andina might have arisen via hybridization between P. infestans and P. mirabilis. DA - 2008/// PY - 2008/// DO - 10.3852/07-074R1 VL - 100 IS - 4 SP - 590-602 SN - 1557-2536 UR - http://www.scopus.com/inward/record.url?eid=2-s2.0-54049104885&partnerID=MN8TOARS KW - Oomycetes KW - Phytophthora KW - potato KW - Stramenopiles ER - TY - JOUR TI - Endoplasmic reticulum quality control and the unfolded protein response: Insights from plants AU - Vitale, Alessandro AU - Boston, Rebecca S. T2 - TRAFFIC AB - Protein quality control (QC) within the endoplasmic reticulum and the related unfolded protein response (UPR) pathway of signal transduction are major regulators of the secretory pathway, which is involved in virtually any aspect of development and reproduction. The study of plant‐specific processes such as pathogen response, seed development and the synthesis of seed storage proteins and of particular toxins is providing novel insights, with potential implications for the general recognition events and mechanisms of action of QC and UPR. DA - 2008/10// PY - 2008/10// DO - 10.1111/j.1600-0854.2008.00780.x VL - 9 IS - 10 SP - 1581-1588 SN - 1398-9219 KW - endoplasmic reticulum KW - molecular chaperones KW - protein bodies KW - protein degradation KW - protein folding ER - TY - JOUR TI - Resistance of Pinus taeda families under artificial inoculations with diverse fusiform rust pathogen populations and comparison with field trials AU - Isik, Fikret AU - Amerson, Henry V. AU - Whetten, Ross W. AU - Garcia, Saul A. AU - Li, Bailian AU - McKeand, Steven E. T2 - CANADIAN JOURNAL OF FOREST RESEARCH-REVUE CANADIENNE DE RECHERCHE FORESTIERE AB - Controlled inoculations with 10 bulk inocula of Cronartium quercuum (Berk) Miyabe ex Shirai f.sp. fusiforme were carried out on open-pollinated progeny of 25 fast-growing Pinus taeda L. parents. The parents had a range of breeding values for resistance to fusiform rust in progeny field trials. There were highly significant differences among the half-sib families in response to inoculations, and these differences were very reproducible; the half-sib family-mean heritability of resistance to controlled inoculation was 0.97. All of the families that were susceptible in the field were susceptible in controlled inoculations, and most (12 of 17) of the field-resistant families were resistant in response to controlled inoculations. Significant pathogenic variability was observed among the different bulk inocula, although this accounted for only 1.9% of the total variation. Genetic differences among families within field-resistant or field-susceptible groups accounted for 13.7% of the total variation. The family by inocula interaction was highly significant, but a single field-resistant family contributed 44% of the total family by inocula interaction variance, and two other field-resistant families also showed significant interactions. DA - 2008/10// PY - 2008/10// DO - 10.1139/X08-111 VL - 38 IS - 10 SP - 2687-2696 SN - 0045-5067 ER - TY - JOUR TI - Regiospecifically alpha-C-13-labeled porphyrins for studies of ground-state hole transfer in multiporphyrin arrays AU - Muresan, Ana Z. AU - Thamyongkit, Patchanita AU - Diers, James R. AU - Holten, Dewey AU - Lindsey, Jonathan S. AU - Bocian, David F. T2 - JOURNAL OF ORGANIC CHEMISTRY AB - Insight into the electronic communication between the individual constituents of multicomponent molecular architectures is essential for the rational design of molecular electronic and/or photonic devices. To clock the ground-state hole/electron-transfer process in oxidized multiporphyrin architectures, a p-diphenylethyne-linked zinc porphyrin dyad was prepared wherein one porphyrin bears two 13C atoms and the other porphyrin is unlabeled. The 13C atoms are located at the 1- and 9-positions (α-carbons symmetrically disposed to the position of linker attachment), which are sites of electron/spin density in the a1u HOMO of the porphyrin. The 13C labels were introduced by reaction of KS13CN with allyl bromide to give the allyl isothiocyanate, which upon Trofimov pyrrole synthesis followed by methylation gave 2-(methylthio)pyrrole-2-13C. Reaction of the latter with paraformaldehyde followed by hydrodesulfurization gave dipyrromethane-1,9-13C, which upon condensation with a dipyrromethane-1,9-dicarbinol bearing three pentafluorophenyl groups gave the tris(pentafluorophenyl)porphyrin bearing 13C labels at the 1,9-positions and an unsubstituted meso (5-) position. Zinc insertion, bromination at the 5-position, and Suzuki coupling with an unlabeled porphyrin bearing a suitably functionalized diphenylethyne linker gave the regiospecifically labeled zinc porphyrin dyad. Examination of the monocation of the isotopically labeled dyad via electron paramagnetic resonance (EPR) spectroscopy (and comparison with the monocations of benchmark monomers, where hole transfer cannot occur) showed that the hole transfer between porphyrin constituents of the dyad is slow (<106 s−1) on the EPR time scale at room temperature. The slow rate stems from the a1u HOMO of the electron-deficient porphyrins, which has a node at the site of linker connection. In contrast, analogous dyads of electron-rich porphyrins (wherein the HOMO is a2u and has a lobe at the site of linker connection) studied previously exhibit rates of hole transfer that are fast (>5 × 107 s−1) on the EPR time scale at room temperature. DA - 2008/9/19/ PY - 2008/9/19/ DO - 10.1021/jo8012836 VL - 73 IS - 18 SP - 6947-6959 SN - 0022-3263 ER - TY - JOUR TI - Neurogenetic networks for startle-induced locomotion in Drosophila melanogaster AU - Yamamoto, Akihiko AU - Zwarts, Liesbeth AU - Callaerts, Patrick AU - Norga, Koenraad AU - Mackay, Trudy F. C. AU - Anholt, Robert R. H. T2 - PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA AB - Understanding how the genome empowers the nervous system to express behaviors remains a critical challenge in behavioral genetics. The startle response is an attractive behavioral model for studies on the relationship between genes, brain, and behavior, as the ability to respond rapidly to harmful changes in the environment is a universal survival trait. Drosophila melanogaster provides a powerful system in which genetic studies on individuals with controlled genetic backgrounds and reared under controlled environmental conditions can be combined with neuroanatomical studies to analyze behaviors. In a screen of 720 lines of D. melanogaster , carrying single P[GT1] transposon insertions, we found 267 lines that showed significant changes in startle-induced locomotor behavior. Excision of the transposon reversed this effect in five lines out of six tested. We infer that most of the 267 lines show mutant effects on startle-induced locomotion that are caused by the transposon insertions. We selected a subset of 15 insertions in the same genetic background in autosomal genes with strong mutant effects and crossed them to generate all 105 possible nonreciprocal double heterozygotes. These hybrids revealed an extensive network of epistatic interactions on the behavioral trait. In addition, we observed changes in neuroanatomy that were caused by these 15 mutations, individually and in their double heterozygotes. We find that behavioral and neuroanatomical phenotypes are determined by a common set of genes that are organized as partially overlapping genetic networks. DA - 2008/8/26/ PY - 2008/8/26/ DO - 10.1073/pnas.0804889105 VL - 105 IS - 34 SP - 12393-12398 SN - 0027-8424 KW - behavioral genetics KW - epistasis KW - sensorimotor integration KW - startle behavior KW - P-element insertional mutagenesis ER - TY - JOUR TI - Investigation of the scope of a new route to ABCD-bilanes and ABCD-porphyrins AU - Dogutan, Dilek Kiper AU - Lindsey, Jonathan S. T2 - JOURNAL OF ORGANIC CHEMISTRY AB - A new route to bilanes and porphyrins bearing four distinct meso substituents has been studied to elucidate the scope and gain entry to previously inaccessible compounds. The route entails (i) synthesis of a 1-bromo-19-acylbilane by acid-catalyzed condensation of a 1-acyldipyrromethane and a 9-bromodipyrromethane-1-carbinol and (ii) intramolecular cyclization of the 1-bromo-19-acylbilane in the presence of a metal salt (MgBr2, 3 mol equiv) and a non-nucleophilic base (DBU, 10 mol equiv) in a noncoordinating solvent (toluene) at 115 degrees C exposed to air to afford the corresponding magnesium(II) porphyrin. In this study, two sets of bilanes were initially prepared to explore substituent effects. In the first set, all bilanes vary only in the nature of the substituent at the 10-position. In the second set, all bilanes vary only in the nature of the substituent attached to the acyl unit (the 20-position). The substituents examined at the 10- and 20-positions include alkyl, aryl (electron-rich, electron-deficient, hindered), heteroaryl, ester, or no substituent (-H). The bilanes were obtained in 35-87% yield, and the target porphyrins in up to 60% yield. Further study of the scope focused on bilanes and porphyrins bearing three heterocyclic substituents (o-, m-, p-pyridyl) or four alkyl groups (ethyl, propyl, butyl, pentyl), in which case microwave irradiation was used for the porphyrin-forming step. Altogether, 17 bilanes and 19 porphyrins were prepared and characterized. In summary, the new route provides access to meso-substituted bilanes and porphyrins for which access is limited via other methods. DA - 2008/9/5/ PY - 2008/9/5/ DO - 10.1021/jo8010396 VL - 73 IS - 17 SP - 6728-6742 SN - 1520-6904 ER - TY - JOUR TI - Global analysis of Arabidopsis gene expression uncovers a complex array of changes impacting pathogen response and cell cycle during geminivirus infection AU - Ascencio-Ibanez, J. T. AU - Sozzani, R. AU - Lee, T. J. AU - Chu, T. M. AU - Wolfinger, R. D. AU - Cella, R. AU - Hanley-Bowdoin, L. T2 - Plant Physiology DA - 2008/// PY - 2008/// VL - 148 IS - 1 SP - 436-454 ER - TY - JOUR TI - Geminivirus-mediated gene silencing from Cotton leaf crumple virus is enhanced by low temperature in cotton AU - Tuttle, John R. AU - Idris, A. M. AU - Brown, Judith K. AU - Haigler, Candace H. AU - Robertson, Dominique T2 - PLANT PHYSIOLOGY AB - A silencing vector for cotton (Gossypium hirsutum) was developed from the geminivirus Cotton leaf crumple virus (CLCrV). The CLCrV coat protein gene was replaced by up to 500 bp of DNA homologous to one of two endogenous genes, the magnesium chelatase subunit I gene (ChlI) or the phytoene desaturase gene (PDS). Cotyledons of cotton cultivar 'Deltapine 5415' bombarded with the modified viral vectors manifested chlorosis due to silencing of either ChlI or PDS in approximately 70% of inoculated plants after 2 to 3 weeks. Use of the green fluorescence protein gene showed that replication of viral DNA was restricted to vascular tissue and that the viral vector could transmit to leaves, roots, and the ovule integument from which fibers originate. Temperature had profound effects on vector DNA accumulation and the spread of endogenous gene silencing. Consistent with reports that silencing against viruses increases at higher temperatures, plants grown at a 30 degrees C/26 degrees C day/night cycle had a greater than 10-fold reduction in viral DNA accumulation compared to plants grown at 22 degrees C/18 degrees C. However, endogenous gene silencing decreased at 30 degrees C/26 degrees C. There was an approximately 7 d delay in the onset of gene silencing at 22 degrees C/18 degrees C, but silencing was extensive and persisted throughout the life of the plant. The extent of silencing in new growth could be increased or decreased by changing temperature regimes at various times following the onset of silencing. Our experiments establish the use of the CLCrV silencing vector to study gene function in cotton and show that temperature can have a major impact on the extent of geminivirus-induced gene silencing. DA - 2008/9// PY - 2008/9// DO - 10.1104/pp.108.123869 VL - 148 IS - 1 SP - 41-50 SN - 1532-2548 ER - TY - JOUR TI - Functional genomics of probiotic Lactobacilli AU - Klaenhammer, Todd R. AU - Altermann, Eric AU - Pfeiler, Erika AU - Buck, Brock Logan AU - Goh, Yong-Jun AU - O'Flaherty, Sarah AU - Barrangou, Rodolphe AU - Duong, Tri T2 - JOURNAL OF CLINICAL GASTROENTEROLOGY AB - Lactic acid bacteria (LAB) have been used in fermentation processes for millennia. Recent applications such as the use of living cultures as probiotics have significantly increased industrial interest. Related bacterial strains can differ significantly in their genotype and phenotype, and features from one bacterial strain or species cannot necessarily be applied to a related one. These strain or family-specific differences often represent unique and applicable traits. Since 2002, the complete genomes of 13 probiotic LABs have been published. The presentation will discuss these genomes and highlight probiotic traits that are predicted, or functionally linked to genetic content. We have conducted a comparative genomic analysis of 4 completely sequenced Lactobacillus strains versus 25 lactic acid bacterial genomes present in the public database at the time of analysis. Using Differential Blast Analysis, each genome is compared with 3 other Lactobacillus and 25 other LAB genomes. Differential Blast Analysis highlighted strain-specific genes that were not represented in any other LAB used in this analysis and also identified group-specific genes shared within lactobacilli. Lactobacillus-specific genes include mucus-binding proteins involved in cell-adhesion and several transport systems for carbohydrates and amino acids. Comparative genomic analysis has identified gene targets in Lactobacillus acidophilus for functional analysis, including adhesion to mucin and intestinal epithelial cells, acid tolerance, bile tolerance, and quorum sensing. Whole genome transcriptional profiling of L. acidophilus, and isogenic mutants thereof, has revealed the impact of varying conditions (pH, bile, carbohydrates) and food matrices on the expression of genes important to probiotic-linked mechanisms. DA - 2008/9// PY - 2008/9// DO - 10.1097/MCG.0b013e31817da140 VL - 42 IS - 8 SP - S160-S162 SN - 0192-0790 KW - Lactobacillus acidophilus KW - Lactobacillus johnsonii KW - Lactobacillus plantarum KW - Lactobacillus gasseri KW - genomics KW - probiotic cultures KW - lactic acid bacteria ER - TY - JOUR TI - C/EBP beta represses p53 to promote cell survival downstream of DNA damage independent of oncogenic Ras and p19(Arf) AU - Ewing, S. J. AU - Zhu, S. AU - Zhu, F. AU - House, J. S. AU - Smart, R. C. T2 - CELL DEATH AND DIFFERENTIATION AB - CCAAT/enhancer-binding protein-β (C/EBPβ) is a mediator of cell survival and tumorigenesis. When C/EBPβ−/− mice are treated with carcinogens that produce oncogenic Ras mutations in keratinocytes, they respond with abnormally elevated keratinocyte apoptosis and a block in skin tumorigenesis. Although this aberrant carcinogen-induced apoptosis results from abnormal upregulation of p53, it is not known whether upregulated p53 results from oncogenic Ras and its ability to induce p19Arf and/or activate DNA-damage response pathways or from direct carcinogen-induced DNA damage. We report that p19Arf is dramatically elevated in C/EBPβ−/− epidermis and that C/EBPβ represses a p19Arf promoter reporter. To determine whether p19Arf is responsible for the proapoptotic phenotype in C/EBPβ−/− mice, C/EBPβ−/−;p19Arf−/− mice were generated. C/EBPβ−/−;p19Arf−/− mice responded to carcinogen treatment with increased p53 and apoptosis, indicating p19Arf is not essential. To ascertain whether oncogenic Ras activation induces aberrant p53 and apoptosis in C/EBPβ−/− epidermis, we generated K14-ER:Ras;C/EBPβ−/− mice. Oncogenic Ras activation induced by 4-hydroxytamoxifen did not produce increased p53 or apoptosis. Finally, when C/EBPβ−/− mice were treated with differing types of DNA-damaging agents, including alkylating chemotherapeutic agents, they displayed aberrant levels of p53 and apoptosis. These results indicate that C/EBPβ represses p53 to promote cell survival downstream of DNA damage and suggest that inhibition of C/EBPβ may be a target for cancer cotherapy to increase the efficacy of alkylating chemotherapeutic agents. DA - 2008/11// PY - 2008/11// DO - 10.1038/cdd.2008.105 VL - 15 IS - 11 SP - 1734-1744 SN - 1476-5403 KW - apoptosis KW - p53 KW - C/EBP beta KW - p19(Arf) KW - DNA damage KW - keratinocytes ER - TY - JOUR TI - Anti-Mullerian hormone and inhibin B in the definition of ovarian aging and the menopause transition AU - Sowers, MaryFran R. AU - Eyvazzadeh, Aimee D. AU - McConnell, Daniel AU - Yosef, Matheos AU - Jannausch, Mary L. AU - Zhang, Daowen AU - Harlow, Sioban AU - Randolph, John F., Jr. T2 - JOURNAL OF CLINICAL ENDOCRINOLOGY & METABOLISM AB - The objective of the study was to determine whether anti-Mullerian hormone (AMH) and inhibin B are viable endocrine biomarkers for framing the menopause transition from initiation to the final menstrual period (FMP).We assayed AMH, inhibin B, and FSH in 300 archival follicular phase specimens from 50 women with six consecutive annual visits commencing in 1993 when all women were in the pre- and perimenopausal menopause stages. Subsequently each woman had a documented FMP. The assay results were fitted as individual-woman profiles and then related to time to FMP and age at FMP as outcomes.Based on annual values from six time points prior to the FMP, (log)AMH longitudinal profiles declined and were highly associated with a time point 5 yr prior to FMP [including both observed and values below detection (P < 0.0001 and P = 0.0001, respectively)]. Baseline AMH profiles were also associated with age at FMP (P = 0.035). Models of declining (log)inhibin B profiles (including both observed and values below detection) were associated with time to FMP (P < 0.0001 and P = 0.0003, respectively). There was no significant association of (log)inhibin B profiles with age at FMP.AMH, an endocrine marker that reflects the transition of resting primordial follicles to growing follicles, declined to a time point 5 yr prior to the FMP; this may represent a critical biological juncture in the menopause transition. Low and nondetectable levels inhibin B levels also were observed 4-5 yr prior to the FMP but were less predictive of time to FMP or age at FMP. DA - 2008/9// PY - 2008/9// DO - 10.1210/jc.2008-0567 VL - 93 IS - 9 SP - 3478-3483 SN - 1945-7197 ER - TY - JOUR TI - Comparison of Electron-Transfer Rates for Metal- versus Ring-Centered Redox Processes of Porphyrins in Monolayers on Au(111) AU - Jiao, Jieying AU - Schmidt, Izabela AU - Taniguchi, Masahiko AU - Lindsey, Jonathan S. AU - Bocian, David F. T2 - LANGMUIR AB - The standard electron-transfer rate constants ( k ( 0 )) are measured for redox processes of Fe versus Zn porphyrins in monolayers on Au(111); the former undergoes a metal-centered redox process (conversion between Fe (III) and Fe (II) oxidation states) whereas the latter undergoes a ring-centered redox process (conversion between the neutral porphyrin and the pi-cation radical). Each porphyrin contains three meso-mesityl groups and a benzyl thiol for surface attachment. Under identical solvent (propylene carbonate)/electrolyte (1.0 M Bu 4NCl) conditions, the Zn (II) center has a coordinated Cl (-) ion when the porphyrin is in either the neutral or oxidized state. In the case of the Fe porphyrin, two species are observed a low-potential form ( E l (0) approximately -0.6 V) wherein the metal center has a coordinated Cl (-) ion when it is in either the Fe (II) or Fe (III) state and a high-potential form ( E h (0) approximately +0.2 V) wherein the metal center undergoes ligand exchange upon conversion from the Fe (III) to Fe (II) states. The k ( 0 ) values observed for all of the porphyrins depend on surface concentration, with higher concentrations resulting in slower rates, consistent with previous studies on porphyrin monolayers. The k ( 0 ) values for the ring-centered redox process (Zn chelate) are 10-40 times larger than those for the metal-centered process (Fe chelate); the k ( 0 ) values for the two forms of the Fe porphyrin differ by a factor of 2-4 (depending on surface concentration), the Cl (-) exchanging form generally exhibiting a faster rate. The faster rates for the ring- versus metal-centered redox process are attributed to the participating molecular orbitals and their proximity to the surface (given that the porphyrins are relatively upright on the surface): a pi molecular orbital that has significant electron density at the meso-carbon atoms (one of which is the site of attachment of the linker to the surface anchoring thiol) versus a d-orbital that is relatively well localized on the metal center. DA - 2008/10/21/ PY - 2008/10/21/ DO - 10.1021/la8019843 VL - 24 IS - 20 SP - 12047-12053 SN - 0743-7463 ER - TY - JOUR TI - Sequential transphosphorylation of the BRI1/BAK1 receptor kinase complex impacts early events in brassinosteroid signaling AU - Wang, Xiaofeng AU - Kota, Uma AU - He, Kai AU - Blackburn, Kevin AU - Li, Jia AU - Goshe, Michael B. AU - Huber, Steven C. AU - Clouse, Steven D. T2 - DEVELOPMENTAL CELL AB - Brassinosteroids (BRs) regulate plant development through a signal transduction pathway involving the BRI1 and BAK1 transmembrane receptor kinases. The detailed molecular mechanisms of phosphorylation, kinase activation, and oligomerization of the BRI1/BAK1 complex in response to BRs are uncertain. We demonstrate that BR-dependent activation of BRI1 precedes association with BAK1 in planta, and that BRI1 positively regulates BAK1 phosphorylation levels in vivo. BRI1 transphosphorylates BAK1 in vitro on specific kinase-domain residues critical for BAK1 function. BAK1 also transphosphorylates BRI1, thereby quantitatively increasing BRI1 kinase activity toward a specific substrate. We propose a sequential transphosphorylation model in which BRI1 controls signaling specificity by direct BR binding followed by substrate phosphorylation. The coreceptor BAK1 is then activated by BRI1-dependent transphosphorylation and subsequently enhances signaling output through reciprocal BRI1 transphosphorylation. This model suggests both conservation and distinct differences between the molecular mechanisms regulating phosphorylation-dependent kinase activation in plant and animal receptor kinases. DA - 2008/8/12/ PY - 2008/8/12/ DO - 10.1016/j.devcel.2008.06.011 VL - 15 IS - 2 SP - 220-235 SN - 1534-5807 ER - TY - JOUR TI - Fitness of isolates of Phytophthora capsici resistant to mefenoxam from squash and pepper fields in North Carolina AU - Cafe-Filho, Adalberto C. AU - Ristaino, Jean Beagle T2 - PLANT DISEASE AB - Despite the wide adoption of mefenoxam (Ridomil Gold EC) for vegetables in North Carolina, the incidence of Phytophthora blight on pepper (Capsicum annuum) and squash (Cucurbita pepo) is high. Seventy-five isolates of Phytophthora capsici were collected in five pepper and one squash field in order to assess mefenoxam sensitivity. The relative fitness of resistant and sensitive isolates was contrasted in vitro by their respective rates of colony growth and their ability to produce sporangia in unamended V8 juice agar medium. In in vivo experiments, the aggressiveness of isolates on pepper was evaluated. The frequency of resistant isolates in North Carolina populations was 63%, considerably higher than resistance levels in areas where mefenoxam is not widely adopted. Resistant isolates grew on amended media at rates >80 to 90% and >100% of the nonamended control at 100 μg ml -1 and 5 μg ml -1 , respectively. Sensitive isolates did not growth at 5 or 100 μg ml -1 . All isolates from three fields, including two pepper and a squash field, were resistant to mefenoxam. Populations from other fields were composed of either mixes of sensitive and resistant isolates or only sensitive isolates. Response to mefenoxam remained stable during the course of in vitro and in planta experiments. Occurrence of a mefenoxam-resistant population of P. capsici on squash is reported here for the first time in North Carolina. When measured by rate of colony growth, sporulation in vitro, or aggressiveness in planta, fitness of resistant isolates was not reduced. Mefenoxam-resistant isolates from squash were as aggressive on pepper as sensitive or resistant pepper isolates. These results suggest that mefenoxam-resistant populations of P. capsici are as virulent and fit as sensitive populations. DA - 2008/10// PY - 2008/10// DO - 10.1094/PDIS-92-10-1439 VL - 92 IS - 10 SP - 1439-1443 SN - 1943-7692 UR - http://www.scopus.com/inward/record.url?eid=2-s2.0-55549111617&partnerID=MN8TOARS KW - fungicide resistance KW - metalaxyl KW - oomycetes KW - phenylamides KW - stramenopiles ER - TY - JOUR TI - Epibulbar melanoma in a foal AU - McMullen, Richard J. AU - Clode, Alison B. AU - Pandiri, Arun Kumar R. AU - Malarkey, David E. AU - Michau, Tammy Miller AU - Gilger, Brian C. T2 - VETERINARY OPHTHALMOLOGY AB - A case of epibulbar melanoma in a 6-month-old, gelded, chestnut Hanoverian foal is reported. The location and clinical appearance upon initial presentation led to the tentative diagnosis of staphyloma or a congenital mass of unknown origin. An attempt was made to surgically excise the mass under general anesthesia, but due to its infiltrative nature and intraoperative appearance, most, but not all was removed without compromising the integrity of the globe. Histopathological evaluation revealed a multinodular to packeted, poorly demarcated, unencapsulated, infiltrative exophytic melanocytic neoplasm composed of bundles and nests of plump spindloid to polygonal heavily pigmented epithelioid neoplastic cells interspersed with pigment-laden macrophages within a fine fibrovascular stroma. Upon examination after enucleation, neoplastic cells were found to infiltrate into the lateral cornea, sclera and the choroid. This is a unique case of an epibulbar melanoma with choroidal invasion in a foal. Based on the sudden onset and rapid growth as well as the histological evidence of invasion, well-differentiated features, heavy pigmentation, and no apparent mitoses, this neoplasm was considered to be a low-grade malignant melanoma. At 14 months after excision there is no evidence of recurrence. DA - 2008/// PY - 2008/// DO - 10.1111/j.1463-5224.2008.00637.x VL - 11 SP - 44-50 SN - 1463-5216 KW - congenital KW - epibulbar KW - equine KW - melanoma KW - neoplasia ER - TY - JOUR TI - Acute HIV-1 infection: targeting the regulator AU - Dean, Gregg T2 - BLOOD AB - In this issue of Blood , Jiang and colleagues describe the use of humanized rag2-/-γC-/- mice to demonstrate the preferential infection and depletion of Tregs during acute HIV-1 infection. The role of regulatory T cells (Tregs), phenotypically defined as CD4+CD25+FoxP3+, has been the subject of DA - 2008/10/1/ PY - 2008/10/1/ DO - 10.1182/blood-2008-07-165704 VL - 112 IS - 7 SP - 2600-2600 SN - 0006-4971 ER - TY - JOUR TI - Serum Cardiac Troponin I Concentration in Dogs with Ehrlichiosis AU - Diniz, P.P.V.P. AU - de Morais, H.S.A. AU - Breitschwerdt, E.B. AU - Schwartz, D.S. T2 - Journal of Veterinary Internal Medicine AB - Background: Ehrlichiosis is a multisystemic disease with the potential to cause cardiomyocyte injury in naturally infected dogs. Hypothesis: Myocardial injury occurs in dogs infected with Ehrlichia canis . Animals: One‐hundred and ninety‐four dogs from Brazil with clinical and laboratory abnormalities indicative of ehrlichiosis. Sixteen healthy dogs served as controls. Methods: Electrocardiogram, echocardiogram, noninvasive blood pressure measurement, and serum cardiac troponin I (cTnI) concentrations were evaluated. Serologic assays and PCR determined the exposure and infection status for E. canis, Anaplasma spp., Babesia canis vogeli, Bartonella spp., Borrelia burgdorferi, Dirofilaria immitis, Ehrlichia chaffeensis, Ehrlichia ewingii, Leishmania chagasi , and spotted‐fever group Rickettsia . Dogs were assigned to groups according to PCR status: E. canis infected, infected with other vector‐borne organisms, sick dogs lacking PCR evidence for infection, and healthy controls. Results: E. canis ‐infected dogs had higher serum cTnI concentrations than controls (median: 0.04 ng/dL; range 0.04–9.12 ng/dL; control median: 0.04 ng/dL; range: 0.04–0.10 ng/dL; P = .012), and acute E. canis infection was associated with myocardial injury (odds ratio [OR]: 2.67, confidence interval [CI] 95%: 1.12–6.40, P = .027). Severity of anemia was correlated with increased risk of cardiomyocyte damage ( r = 0.84, P < .001). Dogs with clinical signs of systemic inflammatory response syndrome (SIRS) were at higher risk for myocardial injury than were other sick dogs (OR: 2.55, CI 95%: 1.31–4.95, P = .005). Conclusions and Clinical Importance: Acute infection with E. canis is a risk factor for myocardial injury in naturally infected Brazilian dogs. Severity of anemia and SIRS might contribute to the pathophysiology of myocardial damage. DA - 2008/9// PY - 2008/9// DO - 10.1111/j.1939-1676.2008.0145.x VL - 22 IS - 5 SP - 1136-1143 LA - en OP - SN - 0891-6640 1939-1676 UR - http://dx.doi.org/10.1111/j.1939-1676.2008.0145.x DB - Crossref KW - canine monocytic ehrlichiosis KW - myocarditis KW - risk factor KW - SIRS KW - troponin ER - TY - JOUR TI - Pleiotropic effects of Drosophila neuralized on complex behaviors and brain structure AU - Rollmann, Stephanie A. AU - Zwarts, Liesbeth AU - Edwards, Alexis C. AU - Yamamoto, Akihiko AU - Callaerts, Patrick AU - Norga, Koenraad AU - Mackay, Trudy F. C. AU - Anholt, Robert R. H. T2 - GENETICS AB - Abstract Understanding how genotypic variation influences variation in brain structures and behavioral phenotypes represents a central challenge in behavioral genetics. In Drosophila melanogaster, the neuralized (neur) gene plays a key role in development of the nervous system. Different P-element insertional mutations of neur allow the development of viable and fertile adults with profoundly altered behavioral phenotypes that depend on the exact location of the inserted P element. The neur mutants exhibit reduced responsiveness to noxious olfactory and mechanosensory stimulation and increased aggression when limited food is presented after a period of food deprivation. These behavioral phenotypes are correlated with distinct structural changes in integrative centers in the brain, the mushroom bodies, and the ellipsoid body of the central complex. Transcriptional profiling of neur mutants revealed considerable overlap among ensembles of coregulated genes in the different mutants, but also distinct allele-specific differences. The diverse phenotypic effects arising from nearby P-element insertions in neur provide a new appreciation of the concept of allelic effects on phenotype, in which the wild type and null mutant are at the extreme ends of a continuum of pleiotropic allelic effects. DA - 2008/7// PY - 2008/7// DO - 10.1534/genetics.108.088435 VL - 179 IS - 3 SP - 1327-1336 SN - 1943-2631 ER - TY - JOUR TI - Failure to identify an association between serologic or molecular evidence ofBartonellainfection and idiopathic rhinitis in dogs AU - Hawkins, Eleanor C. AU - Johnson, Lynelle R. AU - Guptill, Lynn AU - Marr, Henry S. AU - Breitschwerdt, Edward B. AU - Birkenheuer, Adam J. T2 - Journal of the American Veterinary Medical Association AB - Abstract Objective —To determine whether infection with or exposure to Bartonella spp was associated with idiopathic rhinitis in dogs. Design —Case-control study. Animals —44 dogs with idiopathic nasal discharge and 63 age- and weight-matched control dogs without nasal discharge and no clinical signs of bartonellosis. Procedures —Serum was tested for antibodies against Bartonella henselae and Bartonella vinsonii subsp berkhoffii with indirect fluorescent antibody assays. Blood was tested for Bartonella DNA with a PCR assay. Results —Results of the antibody and PCR assays were negative for all 44 dogs with idiopathic nasal discharge. One control dog had antibodies against B henselae ; a second control dog had positive PCR assay results. We did not detect a significant association between assay results and group designation. Conclusions and Clinical Relevance —The present study failed to confirm an association between idiopathic rhinitis and exposure to or infection with Bartonella spp in dogs. Findings do not rule out the possibility that Bartonella infection may cause nasal discharge in some dogs, but the failure to find any evidence of exposure to or infection with Bartonella spp in dogs with idiopathic nasal discharge suggested that Bartonella infection was not a common cause of the disease. DA - 2008/8/15/ PY - 2008/8/15/ DO - 10.2460/javma.233.4.597 VL - 233 IS - 4 SP - 597-599 J2 - Journal of the American Veterinary Medical Association LA - en OP - SN - 0003-1488 UR - http://dx.doi.org/10.2460/javma.233.4.597 DB - Crossref ER - TY - JOUR TI - Enhancement of extra chromosomal recombination in somatic cells by affecting the ratio of homologous recombination (HR) to non-homologous end joining (NHEJ) AU - Zaunbrecher, Gretchen M. AU - Mir, Bashir AU - Dunne, Patrick W. AU - Breen, Matthew AU - Piedrahita, Jorge A. T2 - ANIMAL BIOTECHNOLOGY AB - Abstract Advancements in somatic cell gene targeting have been slow due to the finite lifespan of somatic cells and the overall inefficiency of homologous recombination. The rate of homologous recombination is determined by mechanisms of DNA repair, and by the balance between homologous recombination (HR) and non-homologous end joining (NHEJ). A plasmid-to-plasmid, extra chromosomal recombination system was used to study the effects of the manipulation of molecules involved in NHEJ (Mre11, Ku70/80, and p53) on HR/NHEJ ratios. In addition, the effect of telomerase expression, cell synchrony, and DNA nuclear delivery was examined. While a mutant Mre11 and an anti-Ku aptamer did not significantly affect the rate of NHEJ or HR, transient expression of a p53 mutant increased overall HR/NHEJ by 2.5 fold. However, expression of the mutant p53 resulted in increased aneuploidy of the cultured cells. Additionally, we found no relationship between telomerase expression and changes in HR/NHEJ. In contrast, cell synchrony by thymidine incorporation did not induce chromosomal abnormalities, and increased the ratio of HR/NHEJ 5-fold by reducing the overall rate of NHEJ. Overall our results show that attempts at reducing NHEJ by use of Mre11 or anti-Ku aptamers were unsuccessful. Cell synchrony via thymidine incorporation, however, does increase the ratio of HR/NHEJ and this indicates that this approach may be of use to facilitate targeting in somatic cells by reducing the numbers of colonies that need to be analyzed before a HR is identified. Keywords: Extra chromosomalHomologous recombinationSomatic cells The authors are grateful to Dr. Peter Lansdorp for transformation of cell lines with GFP and hTERT-GFP, and to Bhanu Chowdhary and Yanling Wang for their technical assistance. Funding for this work came from an NIH grant HL51587 to JAP. Notes + Indicates number of hours after addition of thymidine to the culture media. − Indicates hours after removal of thymidine from the + 24 hour cultured cells. DA - 2008/// PY - 2008/// DO - 10.1080/10495390701670099 VL - 19 IS - 1 SP - 6-21 SN - 1532-2378 KW - extra chromosomal KW - homologous recombination KW - somatic cells ER - TY - JOUR TI - Analysis of the genome sequence of Lactobacillus gasseri ATCC 33323 reveals the molecular basis of an autochthonous intestinal organism AU - Azcarate-Peril, M. Andrea AU - Altermann, Eric AU - Goh, Yong Jun AU - Tallon, Richard AU - Sanozky-Dawes, Rosemary B. AU - Pfeiler, Erika A. AU - O'Flaherty, Sarah AU - Buck, B. Logan AU - Dobson, Alleson AU - Duong, Tri AU - Miller, Michael J. AU - Barrangou, Rodolphe AU - Klaenhammer, Todd R. T2 - APPLIED AND ENVIRONMENTAL MICROBIOLOGY AB - ABSTRACT This study presents the complete genome sequence of Lactobacillus gasseri ATCC 33323, a neotype strain of human origin and a native species found commonly in the gastrointestinal tracts of neonates and adults. The plasmid-free genome was 1,894,360 bp in size and predicted to encode 1,810 genes. The GC content was 35.3%, similar to the GC content of its closest relatives, L. johnsonii NCC 533 (34%) and L. acidophilus NCFM (34%). Two identical copies of the prophage LgaI (40,086 bp), of the Sfi11-like Siphoviridae phage family, were integrated tandomly in the chromosome. A number of unique features were identified in the genome of L. gasseri that were likely acquired by horizontal gene transfer and may contribute to the survival of this bacterium in its ecological niche. L. gasseri encodes two restriction and modification systems, which may limit bacteriophage infection. L. gasseri also encodes an operon for production of heteropolysaccharides of high complexity. A unique alternative sigma factor was present similar to that of B. caccae ATCC 43185, a bacterial species isolated from human feces. In addition, L. gasseri encoded the highest number of putative mucus-binding proteins (14) among lactobacilli sequenced to date. Selected phenotypic characteristics that were compared between ATCC 33323 and other human L. gasseri strains included carbohydrate fermentation patterns, growth and survival in bile, oxalate degradation, and adhesion to intestinal epithelial cells, in vitro. The results from this study indicated high intraspecies variability from a genome encoding traits important for survival and retention in the gastrointestinal tract. DA - 2008/8// PY - 2008/8// DO - 10.1128/AEM.00054-08 VL - 74 IS - 15 SP - 4610-4625 SN - 1098-5336 ER - TY - JOUR TI - A Listeria monocytogenes mutant defective in bacteriophage attachment is attenuated in orally inoculated mice and impaired in enterocyte intracellular growth AU - Spears, Patricia A. AU - Suyemoto, M. Mitsu AU - Palermo, Angela M. AU - Horton, John R. AU - Hamrick, Terri S. AU - Havell, Edward A. AU - Orndorff, Paul E. T2 - INFECTION AND IMMUNITY AB - ABSTRACT A Listeria monocytogenes bacteriophage was used to identify a phage-resistant Tn 917 insertion mutant of the mouse-virulent listerial strain F6214-1. The mutant was attenuated when it was inoculated orally into female A/J mice and failed to replicate efficiently in cultured mouse enterocytes. Phage binding studies indicated that the mutant had a cell surface alteration that precluded phage attachment. All phenotypes associated with the mutation could be complemented in trans by a single open reading frame (ORF) that corresponded to the ORF interrupted by the Tn 917 insertion. The complementation effected was, in all cases, at a level indistinguishable from that of the parent. The Tn 917 insertion interrupted a gene that is predicted to encode a group 2 glycosyl transferase (provisionally designated glcV ). A similar glcV gene is present in Listeria welshimeri and Listeria innocua and in some serotypes of L. monocytogenes . We speculate that the loss of the glcV product results in a defective phage receptor and that this alteration coincidentally influences a feature of the normal host-pathogen interaction required for virulence. Interestingly, the glcV lesion, while preventing phage attachment, did not alter the mutant's ability to bind to cultured mouse enterocyte monolayers. Rather, the mutation appeared to alter a subsequent step in intracellular replication measured by a reduction in plaque-forming efficiency and plaque size. In vivo, the mutant was undetectable in the liver and spleen 48 h after oral inoculation. The mutation is significant in part because it is one of the few that produce attenuation when the mutant is delivered orally. DA - 2008/9// PY - 2008/9// DO - 10.1128/IAI.00283-08 VL - 76 IS - 9 SP - 4046-4054 SN - 0019-9567 ER - TY - JOUR TI - Examination of chlorin-bacteriochlorin energy-transfer dyads as prototypes for near-infrared molecular imaging probes AU - Kee, Hooi Ling AU - Nothdurft, Ralph AU - Muthiah, Chinnasamy AU - Diers, James R. AU - Fan, Dazhong AU - Ptaszek, Marcin AU - Bocian, David F. AU - Lindsey, Jonathan S. AU - Culver, Joseph P. AU - Holten, Dewey T2 - PHOTOCHEMISTRY AND PHOTOBIOLOGY AB - Abstract New classes of synthetic chlorin and bacteriochlorin macrocycles are characterized by narrow spectral widths, tunable absorption and fluorescence features across the red and near‐infrared (NIR) regions, tunable excited‐state lifetimes (<1 to >10 ns) and chemical stability. Such properties make dyad constructs based on synthetic chlorin and bacteriochlorin units intriguing candidates for the development of NIR molecular imaging probes. In this study, two such dyads ( FbC‐FbB and ZnC‐FbB ) were investigated. The dyads contain either a free base (Fb) or zinc (Zn) chlorin (C) as the energy donor and a free base bacteriochlorin (B) as the energy acceptor. In both constructs, energy transfer from the chlorin to bacteriochlorin occurs with a rate constant of ∼(5 ps) −1 and a yield of >99%. Thus, each dyad effectively behaves as a single chromophore with an exceptionally large Stokes shift (85 nm for FbC‐FbB and 110 nm for ZnC‐FbB ) between the red‐region absorption of the chlorin and the NIR fluorescence of the bacteriochlorin (λ f = 760 nm, Φ f = 0.19, τ ∼ 5.5 ns in toluene). The long‐wavelength transitions (absorption, emission) of each constituent of each dyad exhibit narrow (≤20 nm) spectral widths. The narrow spectral widths enabled excellent selectivity in excitation and detection of one chlorin–bacteriochlorin energy‐transfer dyad in the presence of the other upon diffuse optical tomography of solution‐phase phantoms. DA - 2008/// PY - 2008/// DO - 10.1111/j.1751-1097.2008.00409.x VL - 84 IS - 5 SP - 1061-1072 SN - 1751-1097 ER - TY - JOUR TI - Characterization of root-knot nematode resistance in Medicago truncatula AU - Dhandaydham, M. AU - Charles, L. AU - Zhu, H. AU - Starr, J. L. AU - Huguet, T. AU - Cook, D. R. AU - Prosperi, J. M. AU - Opperman, C. T2 - Journal of Nematology DA - 2008/// PY - 2008/// VL - 40 IS - 1 SP - 46-54 ER - TY - JOUR TI - Bartonella sp. Bacteremia in Patients with Neurological and Neurocognitive Dysfunction AU - Breitschwerdt, E. B. AU - Maggi, R. G. AU - Nicholson, W. L. AU - Cherry, N. A. AU - Woods, C. W. T2 - Journal of Clinical Microbiology AB - We detected infection with a Bartonella species (B. henselae or B. vinsonii subsp. berkhoffii) in blood samples from six immunocompetent patients who presented with a chronic neurological or neurocognitive syndrome including seizures, ataxia, memory loss, and/or tremors. Each of these patients had substantial animal contact or recent arthropod exposure as a potential risk factor for Bartonella infection. Additional studies should be performed to clarify the potential role of Bartonella spp. as a cause of chronic neurological and neurocognitive dysfunction. DA - 2008/7/16/ PY - 2008/7/16/ DO - 10.1128/JCM.00832-08 VL - 46 IS - 9 SP - 2856-2861 J2 - Journal of Clinical Microbiology LA - en OP - SN - 0095-1137 UR - http://dx.doi.org/10.1128/JCM.00832-08 DB - Crossref ER - TY - JOUR TI - The molecular intersection of brassinosteroid-regulated growth and flowering in Arabidopsis AU - Clouse, Steven D. T2 - PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA AB - Many waterbodies across the United States do not meet water quality standards. To help determine where and to what extent improvements should be sought, policymakers must consider the costs of regulations with their monetized values. We ...Scientific knowledge related to quantifying the monetized benefits for landscape-wide water quality improvements does not meet current regulatory and benefit–cost analysis needs in the United States. In this study we addressed this knowledge gap by ... DA - 2008/5/27/ PY - 2008/5/27/ DO - 10.1073/pnas.0803552105 VL - 105 IS - 21 SP - 7345-7346 SN - 0027-8424 ER - TY - JOUR TI - Sindbis virus conformational changes induced by a neutralizing anti-E1 monoclonal antibody AU - Hernandez, Raquel AU - Paredes, Angel AU - Brown, Dennis T. T2 - JOURNAL OF VIROLOGY AB - ABSTRACT A rare Sindbis virus anti-E1 neutralizing monoclonal antibody, Sin-33, was investigated to determine the mechanism of in vitro neutralization. A cryoelectron microscopic reconstruction of Sindbis virus (SVHR) neutralized with FAb from Sin-33 (FAb-33) revealed conformational changes on the surface of the virion at a resolution of 24 Å. FAb-33 was found to bind E1 in less than 1:1 molar ratios, as shown by the absence of FAb density in the reconstruction and stoichiometric measurements using radiolabeled FAb-33, which determined that about 60 molecules of FAb-33 bound to the 240 possible sites in a single virus particle. FAb-33-neutralized virus particles became sensitive to digestion by endoproteinase Glu-C, providing further evidence of antibody-induced structural changes within the virus particle. The treatment of FAb-33-neutralized or Sin-33-neutralized SVHR with low pH did not induce the conformational rearrangements required for virus membrane-cell membrane fusion. Exposure to low pH, however, increased the amount of Sin-33 or FAb-33 that bound to the virus particles, indicating the exposure of additional epitopes. The neutralization of SVHR infection by FAb-33 or Sin-33 did not prevent the association of virus with host cells. These data are in agreement with the results of previous studies that demonstrated that specific antibodies can inactivate the infectious state of a metastable virus in vitro by the induction of conformational changes to produce an inactive structure. A model is proposed which postulates that the induction of conformational changes in the infectious state of a metastable enveloped virus may be a general mechanism of antibody inactivation of virus infectivity. DA - 2008/6// PY - 2008/6// DO - 10.1128/JVI.02673-07 VL - 82 IS - 12 SP - 5750-5760 SN - 0022-538X ER - TY - JOUR TI - Sequence variation of microRNAs and their binding sites in Arabidopsis AU - Ehrenreich, Ian M. AU - Purugganan, Michael D. T2 - PLANT PHYSIOLOGY AB - Major differences exist between plants and animals both in the extent of microRNA (miRNA)-based gene regulation and the sequence complementarity requirements for miRNA-messenger RNA pairing. Whether these differences affect how these sites evolve at the molecular level is unknown. To determine the extent of sequence variation at miRNAs and their targets in a plant species, we resequenced 16 miRNA families (66 miRNAs in total) and all 52 of the characterized binding sites for these miRNAs in the plant model Arabidopsis (Arabidopsis thaliana), accounting for around 50% of the known miRNAs and binding sites in this species. As has been shown previously in humans, we find that both miRNAs and their target binding sites have very low nucleotide variation and divergence compared to their flanking sequences in Arabidopsis, indicating strong purifying selection on these sites in this species. Sequence data flanking the mature miRNAs, however, exhibit normal levels of polymorphism for the accessions in this study and, in some cases, nonneutral evolution or subtle effects on predicted pre-miRNA secondary structure, suggesting that there is raw material for the differential function of miRNA alleles. Overall, our results show that despite differences in the architecture of miRNA-based regulation, miRNAs and their targets are similarly constrained in both plants and animals. DA - 2008/4// PY - 2008/4// DO - 10.1104/pp.108.116582 VL - 146 IS - 4 SP - 1974-1982 SN - 1532-2548 ER - TY - JOUR TI - Genes related to apoptosis predict necrosis of the liver as a phenotype observed in rats exposed to a compendium of hepatotoxicants AU - Huang, L. AU - Heinloth, A. N. AU - Zeng, Z. B. AU - Paules, R. S. AU - Bushel, P. R. T2 - BMC Genomics DA - 2008/// PY - 2008/// VL - 9 ER - TY - JOUR TI - Biochemical and functional evidence of p53 homology is inconsistent with molecular phylogenetics for distant sequences AU - Fernandes, Andrew D. AU - Atchley, William R. T2 - JOURNAL OF MOLECULAR EVOLUTION DA - 2008/7// PY - 2008/7// DO - 10.1007/s00239-008-9124-2 VL - 67 IS - 1 SP - 51-67 SN - 0022-2844 KW - p53 KW - p63 KW - p73 KW - beta-Sandwich KW - transcription factor KW - p731 ER - TY - JOUR TI - Bartonella-Associated Meningoradiculoneuritis and Dermatitis or Panniculitis in 3 Dogs AU - Cross, J.R. AU - Rossmeisl, J.H. AU - Maggi, R.G. AU - Breitschwerdt, E.B. AU - Duncan, R.B. T2 - Journal of Veterinary Internal Medicine AB - An 11-year-old castrated male mixed breed (dog 1) was evaluated for an 11-day history of progressive paraparesis. The dog traveled extensively throughout the United States and was frequently exposed to ticks. Physical examination abnormalities included a 1-cm subdermal nodule on the caudolateral aspect of the thorax. Neurologic deficits were consistent with a T3–L3 myelopathy, including ambulatory pelvic limb ataxia and paraparesis with intact pelvic limb reflexes, and spinal hyperpathia in the midlumbar area. A CBC, serum biochemical profile, and urinalysis were within reference ranges. Cytologic examination of the subdermal mass revealed pyogranulomatous inflammation. Thoracic radiographs revealed a mild bronchiolar pattern. The dog was anesthetized for radiography and computed tomography (CT) of the thoracolumbar spine. No significant abnormalities were noted on radiographs. The CT revealed an extradural soft-tissue mass (Fig 1A) that was hyperattenuating relative to the spinal cord, and enhanced uniformly following contrast administration (iothalamate sodium [0.45 mL/kg], IV). The mass extended irregularly from the caudal L3 to the cranial L5 vertebral bodies, resulting in significant spinal cord compression in the L4 midbody region and focal bone resorption of the L4 vertebral body (Fig 1B). Differential diagnoses for the CT abnormalities included soft-tissue sarcoma (nerve sheath neoplasia, fibrosarcoma), extradural meningioma, lymphoma, or inflammatory granuloma. (A) Transverse, postcontrast computed tomography (CT) image from dog 1 at the level of the L4 vertebral body. Note the uniformly contrast enhancing mass (arrow) within the vertebral canal resulting in spinal cord compression. Window = 300, level = 40. (B) Transverse, postcontrast CT image from dog 1, obtained at the same level (A). The arrows delineate a focal area of vertebral bone resorption associated with the mass (m) lesion. Window = 1500, level = 300. A right hemilaminectomy defect extending from L3 to L5 was created, which revealed a tan, friable extradural mass extending from L3 to L5. After removal of the extradural mass in a piecemeal fashion, multifocal intradural masses were visualized between L4 and L5, which prompted performance of a regional durectomy and subsequent removal of the intradural masses using blunt dissection. Rhizotomy of the L4 dorsal and ventral roots was performed because of gross thickening of visualized intra- and extradural portions of the nerve roots and nerve. The extradural mass, intradural masses, dura, nerve, and nerve roots were submitted for histopathologic examination. The hemilaminectomy defect was covered with gelfoam and the surgical wound was closed routinely. Histopathologic examination of extra- and intradural portions of the masses revealed abundant fibrous tissue with multiple infiltrative foci consisting of primarily neutrophils with fewer macrophages and giant cells admixed with fibrin and necrotic debris (Fig 2A). There were lymphocytes and plasma cells in the periphery of the lesion. The L4 spinal nerve and nerve roots were infiltrated with islands of lymphocytes, plasma cells, and few neutrophils (Fig 2B). No infectious organisms were detected in excised tissue sections subjected to Gram, Warthin-Starry silver, acid-fast, or periodic acid Schiff (PAS) stains. The final histopathologic diagnosis was pyogranulomatous, lymphoplasmocytic meningoradiculoneuritis. (A) Photomicrograph of intradural portions of the mass lesion from dog 1 demonstrating a pyogranulomatous and lymphoplasmocytic meningitis. Hematoxylin and eosin stain. (B) Photomicrograph of L4 spinal nerve of dog 1. Islands of lymphoplasmocytic infiltrates are visible within the section. Hematoxylin and eosin stain. Serologic assays for blastomycosis, coccidiomycosis, neosporosis, and toxoplasmosis antibodies were negative, as was a cryptococcal capsular antigen test. By indirect fluorescent antibody (IFA) testing, the dog was seroreactive to Bartonella vinsonii subsp. berkhoffi antigens (titer 1 : 64).b Postoperative care included morphine (0.25 mg/kg SC q6h), deracoxib (2.7 mg/kg PO q24h), and lactated Ringers solution (2.75 mL/kg/h IV). Paraparesis improved the day after surgery, and the dog was discharged 5 days later. The owner was instructed to administer azithromycin (13.9 mg/kg PO q24h for 5 days followed by q48h for 23 days). The neurologic examination was normal on recheck day 8 after surgery. B. vinsonii subsp. berkhoffi DNA was detected in an EDTA-anticoagulated blood sample collected at that time using real-time polymerase chain reaction (PCR).b Real-time PCR for Bartonella species performed on formalin-fixed, paraffin-embedded tissues were negative.b,c The histologic findings, serology, and PCR results were interpreted as consistent with active B. vinsonii subsp. berkhoffi infection. Four months later, no clinical abnormalities were present on reexamination. A repeated B. vinsonii subsp. berkhoffi antibody titer was <1 : 16, and PCR on EDTA-anticoagulated blood was negative.b A CT examination of the surgical site was performed under anesthesia and revealed a scant amount of contrast enhancing soft tissue material in the dorsal aspect of the L4 vertebral body in the region of previously noted bone resorption, with no evidence of spinal cord compression (Fig 3). Therapy with azithromycin was repeated (7 mg/kg PO q12h for 6 weeks), based on the CT results. The dog remains clinically normal to date (29 months after surgery). Transverse, postcontrast computed tomography (CT) image from dog 1 at the level of the L4 vertebral body obtained at the 4-month recheck examination, demonstrating relief of spinal cord compression previously associated with the mass lesion. Contrast enhancement of the gelfoam used to cover the hemilaminectomy defect can be seen. Window = 300, level = 40. An 8-year-old, spayed female Shetland sheepdog (dog 2) was referred with a 5-week history of progressive paraparesis despite prednisone therapy (1.3 mg/kg/day PO). The dog had been observed to have multiple infestations with fleas and ticks. Results of a CBC, serum biochemical profile, urinalysis, and thoracolumbar radiographs performed before referral revealed no significant abnormalities. Physical examination abnormalities included multiple, variably sized (0.3–1 cm in diameter) subcutaneous nodules located in the thoracic, flank, and inguinal areas, and palpable hepatomegaly. Neurologic abnormalities were consistent with a T3–L3 myelopathy and included ambulatory pelvic limb ataxia and paraparesis, exaggerated pelvic limb spinal reflexes, a normal cutaneous trunci reflex, and spinal hyperpathia at the L2 level. Fine needle aspirates of nodules from the flank and inguinal areas were cytologically consistent with pyogranulomatous inflammation. Additional cytologic specimens stained with Gram and acid-fast stains did not result in the visualization of organisms. Abdominal ultrasonography revealed a diffusely hyperechoic hepatic parenchyma. No significant abnormalities were noted on thoracic radiographs. The dog was anesthetized for thoracolumbar magnetic resonance imaging, which revealed an intradural-extramedullary mass lesion extending from the L3 vertebra cranially to exit the left L2–L3 intervertebral foramen that was contiguous with a focal nodular thickening of the left L3 spinal nerve. Relative to skeletal muscle, the mass was isointense on T1 images and hyperintense on T2 images, and demonstrated marked homogenous enhancement on postcontrast (gadolinium-DTPA [0.1 mmol/kg], IV) T1 images. Intradural portions of the mass were associated with spinal cord compression in the L2–L3 region. Lumbar cerebrospinal fluid (CSF) was consistent with a mild mixed cellular pleocytosis, characterized by nondegenerate neutrophils and lymphocytes. Nerve sheath neoplasia and meningioma were considered the primary differential diagnoses. The dog was aseptically prepared for an L2–L3 hemilaminectomy and excisional biopsy of one of the lateral thoracic subcutaneous nodules. After creation of a hemilaminectomy defect at L2–L3, a reddish-gray subdural mass centered over the body of L3 was visualized. The mass coursed to the caudal limits of hemilaminectomy, so the defect was extended to involve L3–L4. A durectomy extending from L2 to L4 was performed, which revealed a reddish, friable, intradural-extramedullary mass, which was subsequently removed in a piecemeal fashion. Both intradural and extradural portions of the L3 dorsal and ventral nerve roots and nerve appeared thickened and nodular, and an L3 rhizotomy was performed. The excised mass was submitted for histopathologic analysis and aerobic and fungal cultures. The hemilaminectomy defect was covered with autologous fat, and the surgical site was closed routinely. Biopsies of the intradural mass, excised spinal nerve and nerve roots, and subcutaneous nodule were consistent with pyogranulomatous and lymphoplasmocytic meningitis, radiculoneuritis, and panniculitis, respectively. No etiologic agents were detected following Gram, acid-fast, and PAS staining procedures. Postoperatively, morphine (0.4 mg/kg SC q4–6h) and lactated Ringer's solution (3 mL/kg/h IV) were administered for 36 hours. Serology was negative for blastomycosis and neosporosis. CSF assays for Toxoplasma gondii antibodies and crytococcal capsular antigen were negative. The dog was seropositive to B. vinsonii subsp. berkhoffi (titer 1 : 256).b Real-time PCR for Bartonella species DNA, performed on EDTA-anticoagulated plasma and freshly excised portions of the intradural mass, were both positive for B. vinsonii subsp. berkhoffi DNA.c Aerobic and fungal cultures of the excised mass, CSF, and the skin nodule did not yield bacterial or fungal growth. The dog was discharged to the owners 72 hours after surgery with instructions to administer azithromycin (11.7 mg/kg PO q24h for 6 weeks), and taper the prednisone (0.6 mg/kg PO q24h). Thirteen days after surgery, the subcutaneous nodules had largely resolved, with only 2 small nodules being noted in the inguinal region. The dog was ambulating with mild pelvic limb ataxia, and the owners were instructed to continue the azithromycin for 3 more weeks, and reduce the prednisone therapy to 0.6 mg/kg PO q48h for 1 week. Six months later, the dog was normal on recheck examination. The serum B. vinsonii antibody titer was < 1:16, and Bartonella PCR using EDTA-anticoagulated blood was negative.b Another recheck performed 17 months postoperatively identified no physical abnormalities. A 9-year-old, castrated male Miniature Schnauzer was evaluated for an 18-day history of progressive paraparesis. The dog lived primarily indoors and did not travel extensively. Physical examination abnormalities included multiple cutaneous nodules over the right quadriceps and right popliteal lymphadenopathy. Neurologic deficits were consistent with a T3–L3 myelopathy (paraplegia, clonic pelvic limb reflexes, intact nociception, and cutaneous trunci reflex absent caudal to L1). A CBC, biochemical profile, urinalysis, and fine needle aspirates of the right popliteal lymph node and cutaneous nodules were performed. The CBC and urinalysis were unremarkable. Serum biochemical abnormalities included increased alanine aminotransferase (166 U/L; reference range, 17–66 U/L) and ALP activities (234 U/L; reference range, 14–105 U/L), and hyperglobulinemia (4.4 g/dL; reference range, 2.2–4.0 g/dL). The lymph node and cutaneous nodule aspirates were consistent with reactive hyperplasia and pyogranulomatous inflammation, respectively. The dog was anesthetized for a thoracolumbar CT, which revealed a ventral extradural, homogenously contrast enhancing mass lesion causing spinal cord compression in the L1–L2 region. Lumbar CSF analysis was consistent with a lymphocytic pleocytosis. Following a left L1–L2 hemilaminectomy, a tan, friable, extradural mass was visualized and removed. Portions of the mass were adherent to the dura, which was locally excised, and the surgical site was closed routinely. Postoperative treatment consisted of morphine (0.4 mg/kg SC q4–6h) and urinary bladder expression q8h. The excised mass was histologically consistent with pyogranulomatous meningitis, with no etiologic agents visualized. An EDTA-anticoagulated blood sample was positive for B. vinsonii subsp. berkhoffi DNA using real-time PCR.c Five days postoperatively, the dog was discharged with weak voluntary motor activity. The owners were instructed to begin treatment with clindamycin (13 mg/kg PO q12h) and enrofloxacin (5.9 mg/kg PO q12h). On recheck 10 days later, the subcutaneous nodules had diminished in size, and the dog was able to ambulate with sling assistance and urinate voluntarily. The clindamycin and enrofloxacin were continued for 6 weeks. Eight weeks later, the referring veterinarian reported that the cutaneous nodules and lymphadenopathy had resolved, and the dog was ambulatory with mild ataxia. To the authors' knowledge, these are the first dogs with B. vinsonii subsp. berkhoffi associated with inflammation involving the spinal meninges, nerve roots, and nerves. Although direct causation has not been established, Bartonella spp. have been associated with various canine nervous system disorders, including myelopathy,1 neutrophilic or granulomatous meningoencephalitis,2 anterior uveitis and choroiditis,3 and meningitis.4Bartonella may affect any topographical portion of the nervous system, resulting in meningoencephalitis, transverse myelitis, vertebral osteomyelitis associated with extradural abscessation, and focal pyogranulomatous meningoradiculoneuritis.1,2,4–6 Proposed mechanisms for Bartonella-associated neurologic disease include direct bacterial invasion and indirect toxin-, immune-, or vasculitis-mediated injury.7 In in vitro studies using feline microglial cells, B. henselae was able to invade microglia, but not astrocytes, and intracellular infection did not induce ultrastructural microglial changes.8 The cellular localization of Bartonella spp. within specific nervous tissues has not been studied, and the extent to which organisms can persist within cells of the CNS is unknown. Experimental infection of cats with B. henselae and B. clarridgeiae suggests that CNS tissues can harbor Bartonella during periods of abacteremia.9 All dogs in this series had concurrent pyogranulomatous dermatitis or panniculitis. Bartonella spp. have been associated with granulomatous inflammation in extraneural canine tissues, including lymphadenitis, rhinitis, panniculitis, and polysystemic granulomatous disease.4,10–12Bartonella species DNA was found in the blood of a dog with α1-proteinase inhibitor deficiency, panniculitis, polyarthritis, and meningitis.4 The role of Bartonella as a cause or cofactor in the development of panniculitis in dogs requires additional investigation. Diagnosing bartonellosis can be difficult. Unless immunocompromised, the Warthrin–Starry silver stain is insensitive for the diagnosis of Bartonella infection in humans.13 Isolation of Bartonella species by conventional blood culture also has a low sensitivity in dogs and humans.14,15 In a study of canine endocarditis, 5/18 dogs demonstrated seroreactivity to Bartonella spp. antigens and had postmortem evidence of Bartonella spp. DNA in diseased aortic valves, although only 1/5 dogs was positive by conventional blood culture.15 Because of the chronic, insidious nature of bartonellosis, prior antibiotic treatment complicates the diagnosis of Bartonella infection by making culture and PCR less sensitive.16,17 Although serology can be useful to establish prior exposure to or active infection with Bartonella species, nearly half of infected dogs can be seronegative by IFA testing.4,14,16 Combinations of pre-enrichment liquid culture in an insect cell culture–based growth medium with PCR detection of Bartonella spp. resulted in enhanced detection or isolation of several Bartonella spp. as compared with conventional culture techniques.14 The ideal antibiotic for the treatment of Bartonella infections in dogs is unknown. Bartonella isolates have demonstrated in vitro susceptibility to β-lactams, aminoglycosides, macrolides, doxycycline, and rifampin.18 Tetracyclines, fluoroquinolones, β-lactams, and combinations of these drugs have been used to treat dogs and cats with Bartonella spp. infections.2,11,17 However, in vivo treatment becomes more difficult as Bartonella species have been found within erythrocytes, macrophages, endothelial cells, microglial cells, pericytes, and neutrophils in cell cultures and tissues obtained from multiple animal species.8 Using antibiotics that accumulate within leukocytes, such as azithromycin and fluoroquinolones, may prove beneficial because these cells can migrate to infectious foci such as granulomatous lesions.19 Evidence suggests that specific strains of B. henselae can be resistant to macrolides.20 One study indicated that enrofloxacin was more efficient at eliminating Bartonella-associated bacteremia than doxycycline, but neither drug eliminated bacteremia in all experimentally infected cats.17 It has been speculated that the variation in antimicrobial therapeutic outcome may not be solely related to the efficacy of each antibiotic, but to the immune-mediated changes caused by Bartonella species.21 The outcomes of the dogs in this series provide support for fluoroquinolone or azithromycin usage for Bartonella infections. In this report, additional decompressive surgery also allowed for rapid improvement in neurologic deficits. Surgical decompression, in combination with appropriate antimicrobial therapy, is indicated in humans and dogs with specific focal infectious spinal cord diseases.5,22 Although not evaluated in this study, medical management alone might result in progressive neurological deterioration or residual dysfunction. Bartonella spp. infections should be considered as a differential diagnosis in dogs with thoracolumbar myelopathy, especially if accompanied by a nodular dermatosis. Serology, pre-enrichment blood culture, and PCR from tissues should be used to support the diagnosis. Treatment through surgical decompression with azithromycin or doxycycline-enrofloxacin combinations may be appropriate for Bartonella-associated pyogranulomatous inflammation in dogs. Prospective studies are needed to determine the incidence of focal neurologic lesions in animals and humans that are positive for Bartonella spp. based on serology, culture, or PCR. aCross JR, Rossmeisl JH. Bartonella-associated pyogranulomatous meningoradiculoneuritis and nodular dermatitis in 3 dogs. J Vet Int Med 2007;21(3):591 (abstract #68) bIntracellular Pathogens Research Laboratory, Center for Comparative Medicine and Translational Research, College of Veterinary Medicine, North Carolina State University, Raleigh, NC cVeterinary Diagnostic Laboratory, Colorado State University, College of Veterinary Medicine and Biomedical Sciences, Fort Collins, CO DA - 2008/5// PY - 2008/5// DO - 10.1111/j.1939-1676.2008.0087.x VL - 22 IS - 3 SP - 674-678 LA - en OP - SN - 0891-6640 1939-1676 UR - http://dx.doi.org/10.1111/j.1939-1676.2008.0087.x DB - Crossref ER - TY - JOUR TI - Top-down identification and quantification of stable isotope labeled proteins from Aspergillus flavus using online nano-flow reversed-phase liquid chromatography coupled to a LTQ-FTICR mass spectrometer AU - Collier, Timothy S. AU - Hawkridge, Adam M. AU - Georgianna, D. Ryan AU - Payne, Gary A. AU - Muddiman, David C. T2 - ANALYTICAL CHEMISTRY AB - Online liquid chromatography-mass spectrometric (LC-MS) analysis of intact proteins (i.e., top-down proteomics) is a growing area of research in the mass spectrometry community. A major advantage of top-down MS characterization of proteins is that the information of the intact protein is retained over the vastly more common bottom-up approach that uses protease-generated peptides to search genomic databases for protein identification. Concurrent to the emergence of top-down MS characterization of proteins has been the development and implementation of the stable isotope labeling of amino acids in cell culture (SILAC) method for relative quantification of proteins by LC-MS. Herein we describe the qualitative and quantitative top-down characterization of proteins derived from SILAC-labeled Aspergillus flavus using nanoflow reversed-phase liquid chromatography directly coupled to a linear ion trap Fourier transform ion cyclotron resonance mass spectrometer (nLC-LTQ-FTICR-MS). A. flavus is a toxic filamentous fungus that significantly impacts the agricultural economy and human health. SILAC labeling improved the confidence of protein identification, and we observed 1318 unique protein masses corresponding to 659 SILAC pairs, of which 22 were confidently identified. However, we have observed some limiting issues with regard to protein quantification using top-down MS/MS analyses of SILAC-labeled proteins. The role of SILAC labeling in the presence of competing endogenously produced amino acid residues and its impact on quantification of intact species are discussed in detail. DA - 2008/7/1/ PY - 2008/7/1/ DO - 10.1021/ac800254z VL - 80 IS - 13 SP - 4994-5001 SN - 1520-6882 ER - TY - JOUR TI - The drosophila gap gene giant has an anterior segment identity function mediated through disconnected and teashirt AU - Sanders, Lisa R. AU - Pate, Mukund AU - Mahaffey, James W. T2 - GENETICS AB - Abstract The C2H2 zinc-finger-containing transcription factors encoded by the disconnected (disco) and teashirt (tsh) genes contribute to the regionalization of the Drosophila embryo by establishing fields in which specific Homeotic complex (Hom-C) proteins can function. In Drosophila embryos, disco and the paralogous disco-related (disco-r) are expressed throughout most of the epidermis of the head segments, but only in small patches in the trunk segments. Conversely, tsh is expressed extensively in the trunk segments, with little or no accumulation in the head segments. Little is known about the regulation of these genes; for example, what limits their expression to these domains? Here, we report the regulatory effects of gap genes on the spatial expression of disco, disco-r, and tsh during Drosophila embryogenesis. The data shed new light on how mutations in giant (gt) affect patterning within the anterior gt domain, demonstrating homeotic function in this domain. However, the homeosis does not occur through altered expression of the Hom-C genes but through changes in the regulation of disco and tsh. DA - 2008/5// PY - 2008/5// DO - 10.1534/genetics.107.084988 VL - 179 IS - 1 SP - 441-453 SN - 1943-2631 ER - TY - JOUR TI - Temperature-dependent regulation of proteins in Aspergillus flavus: Whole organism stable isotope labeling by amino acids AU - Georgianna, D. Ryan AU - Hawkridge, Adam M. AU - Muddiman, David C. AU - Payne, Gary A. T2 - JOURNAL OF PROTEOME RESEARCH AB - Stable isotope labeling by amino acids in cell culture (SILAC) has been used in many different organisms including yeast, mammalian cells, and Arabidopsis cell culture. We present an adaptation of this method to quickly quantify protein changes in response to environmental stimuli regulating biosynthesis of the carcinogen aflatoxin in the fungus Aspergillus flavus. Changes in relative protein concentrations in response to temperature were quantified and compared to changes in aflatoxin biosynthesis and the transcription of the aflatoxin biosynthetic genes. In a comparison between conducive (28 °C) and nonconducive (37 °C) temperatures for aflatoxin biosynthesis, 31 proteins were found to be more abundant at 37 °C and 18 more abundant at 28 °C. The change in expression of the aflatoxin pathway enzymes closely followed the strong repression of both aflatoxin biosynthesis and transcription of the aflatoxin pathway genes observed at 37 °C. Transcripts corresponding to the 379 proteins quantified by SILAC were analyzed using microarrays, but their expression did not always correlate well with transcript levels of encoding genes. This is the first reported labeling of a multicellular free-living prototroph using the SILAC procedure to compare 13C6-arginine-labeled samples to 12C6-arginine-labeled samples for quantitative proteomics. The data presented shows the utility of this procedure in quantifying changes in protein expression in response to environmental stimuli. DA - 2008/7// PY - 2008/7// DO - 10.1021/pr8001047 VL - 7 IS - 7 SP - 2973-2979 SN - 1535-3907 KW - SILAC KW - mass spectrometry KW - proteomics KW - Aspergillus flavus KW - aflatoxin ER - TY - JOUR TI - Metal-molecule interactions upon deposition of copper overlayers on reactively functionalized porphyrin monolayers on Si(100) AU - Anariba, Franklin AU - Schmidt, Lzabela AU - Muresan, Ana Z. AU - Lindsey, Jonathan S. AU - Bocian, David F. T2 - LANGMUIR AB - The interaction of evaporated Cu deposited on a series of porphyrins in monolayers covalently attached to Si(100) substrates was investigated using cyclic voltammetry and FTIR spectroscopy. Each porphyrin contains a triallyl tripod attached to the porphyrin via a p-phenylene unit. The tripod anchors the porphyrin to the Si(100) substrate via hydrosilylation of the allyl groups. Two of the porphyrins are Zn chelates that possess meso p-cyanophenyl substituentsone, ZnP-CND, contains a single group opposite (distal) to the tripodal surface anchor, whereas the other, ZnP-CNL, contains two groups orthogonal (lateral) to the surface anchor. A third Zn porphyrin, ZnP, containing nonreactive p-tolyl groups at all three nonanchoring meso positions, was examined for comparison. The fourth porphyrin, FbP-HD, is a metal-free species (free base) that contains nonreactive phenyl (distal) and p-tolyl groups (lateral) at the three nonanchoring meso positions. The fifth porphyrin, CuP-HD, is the Cu chelate of FbP-HD, and serves as a reference complex for evaluating the effects of Cu metal deposition onto FbP-HD. The studies indicate that all of the porphyrin monolayers are robust under the conditions of Cu deposition, experiencing no noticeable degradation. In addition, the Cu metal does not penetrate through the monolayer to form electrically conductive filaments. For the ZnP-CND monolayers, the deposited Cu quantitatively reacts/complexes with the distal cyano group. In contrast, for the ZnP-CNL monolayers no reaction/complexation of the lateral cyano groups is observed. For the FbP-HD monolayers, Cu deposition results in quantitative insertion of Cu into the free base porphyrin. Collectively, the studies demonstrate that porphyrin monolayers are amenable to direct deposition of Cu overlayers and that functionalization of the porphyrins can be used to mediate the attributes of the metal-molecule junction. DA - 2008/7/1/ PY - 2008/7/1/ DO - 10.1021/la800472c VL - 24 IS - 13 SP - 6698-6704 SN - 0743-7463 ER - TY - JOUR TI - Hepatic endopolyploidy as a cellular consequence of age-specific selection for rate of development in mice AU - Funk-Keenan, Jhondra AU - Haire, Frances AU - Woolard, Sara AU - Atchley, William R. T2 - JOURNAL OF EXPERIMENTAL ZOOLOGY PART B-MOLECULAR AND DEVELOPMENTAL EVOLUTION AB - Abstract Endopolyploidy is the generation of polyploid cells by DNA replication without subsequent cell division and is correlated with hypertrophic growth or growth via cell size. Thus, selection that alters growth may also change onset and frequency of endopolyploidy as a correlated response. We search for endopolyploidy in the liver in response to age‐specific restricted index selection for the rate of development. Polyploidy changes over ontogeny are described in five mouse lines: two selected for divergence in early growth (0–10 days of age), two selected for divergence in late growth (28–56 days of age), and one randombred control. Polyploid cell frequency within each line increased as ontogeny continued, as expected from previous research. However, selection for altered growth clearly plays a role in the frequency and onset of polyploid cells. Lines selected for divergence in early growth have polyploidy differences after weaning that are not seen in adult mice. However, lines selected for divergence in late growth are divergent in frequency of polyploid cells, starting near sexual maturity and continuing into adulthood. J. Exp. Zool. (Mol. Dev. Evol.) 310B:385–397, 2008 . © 2008 Wiley‐Liss, Inc. DA - 2008/7/15/ PY - 2008/7/15/ DO - 10.1002/jez.b.21211 VL - 310B IS - 5 SP - 385-397 SN - 1552-5007 ER - TY - JOUR TI - Feline bartonellosis and cat scratch disease AU - Breitschwerdt, Edward B. T2 - Veterinary Immunology and Immunopathology AB - Bartonella species are important emerging zoonotic pathogens. Transmission of these organisms in nature may be much more complex than is currently appreciated. Cats can be infected with five Bartonella species, including, Bartonella henselae, Bartonella clarridgeae, Bartonella bovis, Bartonella koehlerae and Bartonella quintana. In addition to cats, numerous domestic and wild animals, including bovine, canine, human, and rodent species can serve as chronically infected reservoir hosts for various intra-erythrocytic Bartonella species. In addition, an increasing number of arthropod vectors, including biting flies, fleas, keds, lice, sandflys and potentially ticks have been implicated in the transmission of various Bartonella species to animals or human beings. In the reservoir host, Bartonella species cause chronic intra-erythrocytic and vascular endothelial infections, with a relapsing bacteremia documented in experimentally infected cats. Although the immunopathology induced by Bartonella infection requires additional study, the organisms can localize to the heart valve (endocarditis), cause granulomatous inflammation in lymph nodes, liver or spleen, induce central nervous system dysfunction with or without cerebrospinal fluid changes, and may contribute to inflammatory polyarthritis. Hematological abnormalities are infrequent, but thrombocytopenia, lymphocytosis, neutropenia, and eosinophilia have been reported in B. henselae-infected cats. Serology, PCR and culture can be used to support a diagnosis of feline bartonellosis, however, due to the high rate of sub-clinical infections among various cat populations, documenting causation in an individual cat is difficult, if not impossible. Response to treatment can be used in conjunction with serology or organism isolation to support a clinical diagnosis of feline bartonellosis. As fleas are involved in the transmission among cats, the use of acaracide products to eliminate fleas from the environment is of critical importance to decrease the risk of B. henselae transmission among cats and to humans. DA - 2008/5// PY - 2008/5// DO - 10.1016/j.vetimm.2008.01.025 VL - 123 IS - 1-2 SP - 167-171 J2 - Veterinary Immunology and Immunopathology LA - en OP - SN - 0165-2427 UR - http://dx.doi.org/10.1016/j.vetimm.2008.01.025 DB - Crossref KW - cat KW - bartonella KW - cat scratch disease KW - epidemiology ER - TY - JOUR TI - Characterization of diazotrophs containing Mo-independent nitrogenases, isolated from diverse natural environments AU - Betancourt, Doris A. AU - Loveless, Telisa M. AU - Brown, James W. AU - Bishop, Paul E. T2 - APPLIED AND ENVIRONMENTAL MICROBIOLOGY AB - Molybdenum-independent nitrogenases were first described in the nitrogen-fixing bacterium Azotobacter vinelandii and have since been described in other diazotrophic bacteria. Previously, we reported the isolation of seven diazotrophs with Mo-independent nitrogenases from aquatic environments. In the present study, we extend these results to include diazotrophs isolated from wood chip mulch, soil, "paraffin dirt," and sediments from mangrove swamps. Mo-deficient, N-free media under both aerobic and anaerobic conditions were used for the isolations. A total of 26 isolates were genetically and physiologically characterized. Their phylogenetic placement was determined using 16S rRNA gene sequence analysis. Most of the isolates are members of the gamma subdivision of the class Proteobacteria and appear to be specifically related to fluorescent pseudomonads and azotobacteria. Two other isolates, AN1 and LPF4, are closely related to Enterobacter spp. and Paenibacillus spp., respectively. PCR and/or Southern hybridization were used to detect the presence of nitrogenase genes in the isolates. PCR amplification of vnfG and anfG was used to detect the genetic potential for the expression of the vanadium-containing nitrogenase and the iron-only nitrogenase in the isolates. This study demonstrates that diazotrophs with Mo-independent nitrogenases can be readily isolated from diverse natural environments. DA - 2008/6// PY - 2008/6// DO - 10.1128/AEM.02694-07 VL - 74 IS - 11 SP - 3471-3480 SN - 1098-5336 ER - TY - JOUR TI - Variation in genetic architecture of olfactory behaviour among wild-derived populations of Drosophila melanogaster AU - Lavagnino, N. J. AU - Anholt, R. R. H. AU - Fanara, J. J. T2 - JOURNAL OF EVOLUTIONARY BIOLOGY AB - Odour-guided behaviour is a quantitative trait determined by many genes that are sensitive to gene-environment interactions. Different natural populations are likely to experience different selection pressures on the genetic underpinnings of chemosensory behaviour. However, few studies have reported comparisons of the quantitative genetic basis of olfactory behaviour in geographically distinct populations. We generated isofemale lines of Drosophila melanogaster from six populations in Argentina and measured larval and adult responses to benzaldehyde. There was significant variation within populations for both larval and adult olfactory behaviour and a significant genotype x sex interaction (GSI) for adult olfactory behaviour. However, there is substantial variation in the contribution of GSI to the total phenotypic variance among populations. Estimates of evolvability are orders of magnitude higher for larvae than for adults. Our results suggest that the potential for evolutionary adaptation to the chemosensory environment is greater at the larval feeding stage than at the adult reproductive stage. DA - 2008/7// PY - 2008/7// DO - 10.1111/j.1420-9101.2008.01546.x VL - 21 IS - 4 SP - 988-996 SN - 1010-061X KW - behaviour KW - chemical ecology KW - genetic variation KW - phenotypic plasticity KW - quantitative genetics ER - TY - JOUR TI - Swallowtail bacteriochlorins. Lipophilic absorbers for the near-infrared AU - Borbas, K. Eszter AU - Ruzie, Christian AU - Lindsey, Jonathan S. T2 - ORGANIC LETTERS AB - Bacteriochlorins absorb strongly in the near-infrared spectral region and hence are ideally suited for diverse photomedical applications, yet naturally occurring bacteriochlorins have limited stability and synthetic malleability. A de novo route has been exploited to prepare synthetic bacteriochlorins that bear a geminal dimethyl group in each pyrroline ring for stability and a symmetrically branched 1,5-dimethoxypentyl group attached to each pyrrole ring for solubility in lipophilic media. DA - 2008/5/15/ PY - 2008/5/15/ DO - 10.1021/ol800436u VL - 10 IS - 10 SP - 1931-1934 SN - 1523-7060 ER - TY - JOUR TI - RNA interference (RNAi)-induced suppression of nicotine demethylase activity reduces levels of a key carcinogen in cured tobacco leaves AU - Lewis, Ramsey S. AU - Jack, Anne M. AU - Morris, Jerry W. AU - Robert, Vincent J. M. AU - Gavilano, Lily B. AU - Siminszky, Balazs AU - Bush, Lowell P. AU - Hayes, Alec J. AU - Dewey, Ralph E. T2 - PLANT BIOTECHNOLOGY JOURNAL AB - Summary Technologies for reducing the levels of tobacco product constituents that may contribute to unwanted health effects are desired. Target compounds include tobacco‐specific nitrosamines (TSNAs), a class of compounds generated through the nitrosation of pyridine alkaloids during the curing and processing of tobacco. Studies have reported the TSNA N ′‐nitrosonornicotine (NNN) to be carcinogenic in laboratory animals. NNN is formed via the nitrosation of nornicotine, a secondary alkaloid produced through enzymatic N ‐demethylation of nicotine. Strategies to lower nornicotine levels in tobacco ( Nicotiana tabacum L.) could lead to a corresponding decrease in NNN accumulation in cured leaves. The major nicotine demethylase gene of tobacco has recently been isolated. In this study, a large‐scale field trial was conducted to evaluate transgenic lines of burley tobacco carrying an RNA interference (RNAi) construct designed to inhibit the expression of this gene. Selected transgenic lines exhibited a six‐fold decrease in nornicotine content relative to untransformed controls. Analysis of cured leaves revealed a commensurate decrease in NNN and total TSNAs. The inhibition of nicotine demethylase activity is an effective means of decreasing significantly the level of a key defined animal carcinogen present in tobacco products. DA - 2008/5// PY - 2008/5// DO - 10.1111/j.1467-7652.2008.00324.x VL - 6 IS - 4 SP - 346-354 SN - 1467-7644 KW - nicotine demethylase KW - N'-nitrosonornicotine KW - RNA interference KW - tobacco carcinogens KW - tobacco-specific nitrosamines (TSNAs) ER - TY - JOUR TI - Phenotypic plasticity and genotype by environment interaction for olfactory behavior in Drosophila melanogaster AU - Sambandan, Deepa AU - Carbone, Mary Anna AU - Anholt, Robert R. H. AU - Mackay, Trudy E. C. T2 - GENETICS AB - Abstract Genotype by environment interactions (GEI) play a major part in shaping the genetic architecture of quantitative traits and are confounding factors in genetic studies, for example, in attempts to associate genetic variation with disease susceptibility. It is generally not known what proportion of phenotypic variation is due to GEI and how many and which genes contribute to GEI. Behaviors are complex traits that mediate interactions with the environment and, thus, are ideally suited for studies of GEI. Olfactory behavior in Drosophila melanogaster presents an opportunity to systematically dissect GEI, since large numbers of genetically identical individuals can be reared under defined environmental conditions and the olfactory system of Drosophila and its behavioral response to odorants have been well characterized. We assessed variation in olfactory behavior in a population of 41 wild-derived inbred lines and asked to what extent different larval-rearing environments would influence adult olfactory behavior and whether GEI is a minor or major contributing source of phenotypic variation. We found that ∼50% of phenotypic variation in adult olfactory behavior is attributable to GEI. In contrast, transcriptional analysis revealed that only 20 genes show GEI at the level of gene expression [false discovery rate (FDR) &lt; 0.05], some of which are associated with physiological responses to environmental chemicals. Quantitative complementation tests with piggyBac-tagged mutants for 2 of these genes (CG9664 and Transferrin 1) demonstrate that genes that show transcriptional GEI are candidate genes for olfactory behavior and that GEI at the level of gene expression is correlated with GEI at the level of phenotype. DA - 2008/6// PY - 2008/6// DO - 10.1534/genetics.108.086769 VL - 179 IS - 2 SP - 1079-1088 SN - 0016-6731 ER - TY - JOUR TI - Nicotiana benthamiana: Its history and future as a model for plant-pathogen interactions AU - Goodin, Michael M. AU - Zaitlin, David AU - Naidu, Rayapati A. AU - Lommel, Steven A. T2 - MOLECULAR PLANT-MICROBE INTERACTIONS AB - Nicotiana benthamiana is the most widely used experimental host in plant virology, due mainly to the large number of diverse plant viruses that can successfully infect it. Additionally, N. benthamiana is susceptible to a wide variety of other plant-pathogenic agents (such as bacteria, oomycetes, fungi, and so on), making this species a cornerstone of host–pathogen research, particularly in the context of innate immunity and defense signaling. Moreover, because it can be genetically transformed and regenerated with good efficiency and is amenable to facile methods for virus-induced gene silencing or transient protein expression, N. benthamiana is rapidly gaining popularity in plant biology, particularly in studies requiring protein localization, interaction, or plant-based systems for protein expression and purification. Paradoxically, despite being an indispensable research model, little is known about the origins, genetic variation, or ecology of the N. benthamiana accessions currently used by the research community. In addition to addressing these latter topics, the purpose of this review is to provide information regarding sources for tools and reagents that can be used to support research in N. benthamiana. Finally, we propose that N. benthamiana is well situated to become a premier plant cell biology model, particularly for the virology community, who as a group were the first to recognize the potential of this unique Australian native. DA - 2008/8// PY - 2008/8// DO - 10.1094/mpmi-21-8-1015 VL - 21 IS - 8 SP - 1015-1026 SN - 1943-7706 KW - AFLP KW - agroinfiltration KW - Arabidopsis KW - VIGS ER - TY - JOUR TI - Molecular characterization of Bartonella vinsonii subsp berkhoffii genotype III AU - Cadenas, M. B. AU - Bradley, J. AU - Maggi, Ricardo AU - Takara, M. AU - Hegarty, B. C. AU - Breitschwerdt, Edward T2 - Journal of Clinical Microbiology DA - 2008/// PY - 2008/// DO - 10.1128/JCM.62456-07 VL - 46 IS - 5 SP - 1858–1860 ER - TY - JOUR TI - Mannitol biosynthesis is required for plant pathogenicity by Alternaria alternata AU - Veleaz, Heriberto AU - Glassbrook, Norman J. AU - Daub, Margaret E. T2 - FEMS MICROBIOLOGY LETTERS AB - Mannitol has been hypothesized to play a role in antioxidant defense. In previous work, we confirmed the presence of the two mannitol biosynthetic enzymes, mannitol dehydrogenase (MtDH) and mannitol 1-phosphate 5-dehydrogenase (MPDH), in the fungus Alternaria alternata and created disruption mutants for both enzymes. These mutants were used to investigate the role of mannitol in pathogenicity of A. alternata on its host, tobacco. Conidia of all mutants were viable and germinated normally. GC-MS analysis demonstrated elevated levels of trehalose in the mutants, suggesting that trehalose may substitute for mannitol as a storage compound for germination. Tobacco inoculation showed no reduction in lesion severity caused by the MtDH mutant as compared with wild type; however, the MPDH mutant and a mutant in both enzymes caused significantly less disease. Microscopy analysis indicated that the double mutant was unaffected in the ability to germinate and produce appressoria on tobacco leaves and elicited a defense response from the host, indicating that it was able to penetrate and infect the host. We conclude that mannitol biosynthesis is required for pathogenesis of A. alternata on tobacco, but is not required for spore germination either in vitro or in planta or for initial infection. DA - 2008/8// PY - 2008/8// DO - 10.1111/j.1574-6968.2008.01224.x VL - 285 IS - 1 SP - 122-129 SN - 0378-1097 KW - mannitol metabolism KW - reactive oxygen species KW - antioxidant KW - plant disease ER - TY - JOUR TI - Genetic variation in early growth and bud production among natural populations of fraser fir AU - Emerson, J. L. AU - Frampton, J. AU - McKeand, S. E. T2 - HortScience DA - 2008/// PY - 2008/// VL - 43 IS - 3 SP - 661-666 ER - TY - JOUR TI - Factors Associated with the Occurrence of Epistaxis in Natural Canine Leishmaniasis (Leishmania infantum) AU - Petanides, T.A. AU - Koutinas, A.F. AU - Mylonakis, M.E. AU - Day, M.J. AU - Saridomichelakis, M.N. AU - Leontides, L.S. AU - Mischke, R. AU - Diniz, P. AU - Breitschwerdt, E.B. AU - Kritsepi, M. AU - Garipidou, V.A. AU - Koutinas, C.K. AU - Lekkas, S. T2 - Journal of Veterinary Internal Medicine AB - Background: Canine leishmaniasis (CanL) is a common cause of epistaxis in dogs residing in endemic areas. The pathogenesis of CanL‐associated epistaxis has not been fully explored because of the limited number of cases reported so far. Hypothesis: Epistaxis in CanL could be attributed to more than 1 pathomechanism such as hemostatic dysfunction, biochemical abnormalities, chronic rhinitis, and coinfections occurring in various combinations. Animals: Fifty‐one dogs with natural CanL. Methods: The allocation of 51 dogs in this cross‐sectional study was based on the presence (n = 24) or absence (n = 27) of epistaxis. The potential associations among epistaxis and concurrent infections ( Ehrlichia canis , Bartonella spp., and Aspergillus spp.), biochemical and hemostatic abnormalities, and nasal histopathology were investigated. Results: Hypergammaglobulinemia ( P = .044), increased serum viscosity ( P = .038), decreased platelet aggregation response to collagen ( P = .042), and nasal mucosa ulceration ( P = .039) were more common in the dogs with epistaxis than in those without epistaxis. The other significant differences between the 2 groups involved total serum protein ( P = .029) and γ‐globulin ( P = .013) concentrations, which were higher, and the percentage platelet aggregation to collagen, which was lower ( P = .012) in the epistaxis dogs. Clinical Importance: CanL‐associated epistaxis appears to be the result of multiple and variable pathogenetic factors such as thrombocytopathy, hyperglobulinemia‐induced serum hyperviscosity, and nasal mucosa ulceration. DA - 2008/7// PY - 2008/7// DO - 10.1111/j.1939-1676.2008.0129.x VL - 22 IS - 4 SP - 866-872 LA - en OP - SN - 0891-6640 1939-1676 UR - http://dx.doi.org/10.1111/j.1939-1676.2008.0129.x DB - Crossref KW - dysproteinemia KW - rhinitis KW - thrombocytopathy ER - TY - JOUR TI - Evaluation of conventional and real-time PCR assays for detection and differentiation of Spotted Fever Group Rickettsia in dog blood AU - Kidd, L. AU - Maggi, R. AU - Diniz, P.P.V.P. AU - Hegarty, B. AU - Tucker, M. AU - Breitschwerdt, E. T2 - Veterinary Microbiology AB - Spotted Fever Group Rickettsia is important cause of emerging and re-emerging infectious disease in people and dogs. Importantly, dogs can serve as sentinels for disease in people. Sensitive and specific diagnostic tests that differentiate among species of infecting Rickettsia are needed. The objective of this study was to develop a sensitive and specific PCR that differentiates SFG Rickettsia infecting dog blood. Conventional and real-time PCR assays were developed using primers that targeted a small region of the ompA gene. Their sensitivity, determined by testing a cloned target sequence in the presence of host DNA, was 15–30 and 5 copies of DNA, respectively. Testing of Rickettsia cultures and analysis of Rickettsia gene sequences deposited in GenBank verified DNA could be amplified and used to differentiate species. DNA from the blood of infected dogs was also tested. Importantly, Rickettsia DNA was detected before seroconversion in some dogs. The species of infecting Rickettsia was also identified. We conclude these assays may assist in the timely diagnosis of infection with SFG Rickettsia. They may also facilitate the discovery of novel SFG Rickettsia infecting dogs, and in the investigation of dogs as sentinels for emerging rickettsioses. DA - 2008/6// PY - 2008/6// DO - 10.1016/j.vetmic.2007.11.035 VL - 129 IS - 3-4 SP - 294-303 J2 - Veterinary Microbiology LA - en OP - SN - 0378-1135 UR - http://dx.doi.org/10.1016/j.vetmic.2007.11.035 DB - Crossref KW - dog KW - PCR KW - Rickettsia KW - spotted fever ER - TY - JOUR TI - Comprehensive characterization of hybrid junctions comprised of a porphyrin monolayer sandwiched between a coinage metal overlayer and a Si(100) substrate AU - Anariba, Franklin AU - Tiznado, Hugo AU - Diers, James R. AU - Schmidt, Izabela AU - Muresan, Ana Z. AU - Lindsey, Jonathan S. AU - Zaera, Francisco AU - Bocian, David F. T2 - JOURNAL OF PHYSICAL CHEMISTRY C AB - The promise of molecular electronics has stimulated a variety of approaches for making hybrid junctions wherein molecules are sandwiched between two metal contacts or a metal and a semiconductor contact. However, the fate of molecules subsequent to deposition of a top metal contact has generally not been well characterized. Toward this goal, the interaction of evaporated Cu, Ag, and Au films deposited in varying thicknesses (3, 5, and 8 nm) on a series of monolayer-coverage porphyrins covalently attached to Si(100) substrates was investigated. Each porphyrin contains a triallyl tripod attached to the porphyrin via a p-phenylene unit, which anchors the porphyrin to the Si(100) substrate via hydrosilylation of the allyl groups. All of the porphyrins are Zn chelates bearing meso p-tolyl substituents orthogonal (lateral) to the tripodal surface anchor, but contain different substituents in the meso position opposite (distal) to the surface anchor, p-tolyl, α,α,α-trifluoro-p-tolyl, pentafluorophenyl, or p-cyanophenyl, to test the potential for the distal meso substituents to impart different characteristics to the metal/molecule/Si junction. The methods of interrogation used include ellipsometry, atomic force microscopy, FTIR spectroscopy, Raman spectroscopy, X-ray photoelectron spectroscopy, cyclic voltammetry, and current−voltage measurements. The studies indicate that all of the porphyrin monolayers are robust under the conditions of metal deposition, experiencing no noticeable degradation, and that none of the distal functional groups reacts with the deposited metal, except for the nitrile group of the p-cyanophenyl-substituted porphyrin, which reacts/complexes with Cu. Neither Cu nor Ag penetrates through the monolayer to an appreciable extent to form electrically conductive filaments, whereas Au does penetrate through the porphyrin monolayer and contacts the Si substrate. Collectively, the studies provide a comprehensive assessment of the characteristics of the metal (Cu, Ag, Au)/porphyrin monolayer/Si junctions. DA - 2008/6/26/ PY - 2008/6/26/ DO - 10.1021/jp802428y VL - 112 IS - 25 SP - 9474-9485 SN - 1932-7447 ER - TY - JOUR TI - Comparative analysis of transcript abundance in Pinus sylvestris after challenge with a saprotrophic, pathogenic or mutualistic fungus AU - Adomas, Aleksandra AU - Heller, Gregory AU - Olson, Ake AU - Osborne, Jason AU - Karlsson, Magnus AU - Nahalkova, Jarmila AU - Van Zyl, Len AU - Sederoff, Ron AU - Stenlid, Jan AU - Finlay, Roger AU - Asiegbu, Frederick O. T2 - TREE PHYSIOLOGY AB - To investigate functional differences in the recognition and response mechanisms of conifer roots to fungi with different trophic strategies, Pinus sylvestris L. was challenged with a saprotrophic fungus Trichoderma aureoviride Rifai. The results were compared with separate studies investigating pine interactions with a pathogen, Heterobasidion annosum (Fr.) Bref. sensu stricto and an ectomycorrhizal symbiont, Laccaria bicolor Maire (Orton). Global changes in the expression of 2109 conifer genes were assayed 1, 5 and 15 days after inoculation. Gene expression data from a cDNA microarray were analyzed by the 2-interconnected mixed linear model statistical approach. The total number of genes differentially expressed compared with the uninfected control was similar after challenge with the pathogen and the ectomycorrhizal symbiont, but the number of differentially expressed genes increased over time for H. annosum, and decreased for L. bicolor. Inoculation of pine roots with T. aureoviride resulted overall in a much lower number of genes with changed transcript levels compared with inoculation with H. annosum or L. bicolor. Functional classification of the differentially expressed genes revealed that the ectomycorrhizal fungus triggered transient induction of defence-related genes. The response and induction of defence against the pathogen was delayed and the magnitude increased over time. Thus, there were specific transcriptional responses depending on whether the conifer roots were challenged with mutualistic, saprotrophic or pathogenic fungi. This suggests that pine trees are able to recognize diverse fungal species and specifically distinguish whether they are pathogenic, neutral or beneficial microbial agents. DA - 2008/6// PY - 2008/6// DO - 10.1093/treephys/28.6.885 VL - 28 IS - 6 SP - 885-897 SN - 1758-4469 KW - defence KW - ectomycorrhiza KW - Heterobasidion annostan KW - Laccaria bicolor KW - microarray KW - recognition KW - Trichoderma aureoviride ER - TY - JOUR TI - Canine cytogenetics - from band to basepair AU - Breen, M. T2 - CYTOGENETIC AND GENOME RESEARCH AB - Humans and dogs have coexisted for thousands of years, during which time we have developed a unique bond, centered on companionship. Along the way, we have developed purebred dog breeds in a manner that has resulted unfortunately in many of them being affected by serious genetic disorders, including cancers. With serendipity and irony the unique genetic architecture of the 21st century genome of Man’s best friend may ultimately provide many of the keys to unlock some of nature’s most intriguing biological puzzles. Canine cytogenetics has advanced significantly over the past 10 years, spurred on largely by the surge of interest in the dog as a biomedical model for genetic disease and the availability of advanced genomics resources. As such the role of canine cytogenetics has moved rapidly from one that served initially to define the gross genomic organization of the canine genome and provide a reliable means to determine the chromosomal location of individual genes, to one that enabled the assembled sequence of the canine genome to be anchored to the karyotype. Canine cytogenetics now presents the biomedical research community with a means to assist in our search for a greater understanding of how genome architectures altered during speciation and in our search for genes associated with cancers that affect both dogs and humans. The cytogenetics ‘toolbox’ for the dog is now loaded. This review aims to provide a summary of some of the recent advancements in canine cytogenetics. DA - 2008/// PY - 2008/// DO - 10.1159/000118740 VL - 120 IS - 1-2 SP - 50-60 SN - 1424-859X ER - TY - JOUR TI - Accessing the near-infrared spectral region with stable, synthetic, wavelength-tunable bacteriochlorins AU - Taniguchi, Masahiko AU - Cramer, David L. AU - Bhise, Anil D. AU - Kee, Hooi Ling AU - Bocian, David F. AU - Holten, Dewey AU - Lindsey, Jonathan S. T2 - NEW JOURNAL OF CHEMISTRY AB - The near-infrared (NIR) spectral region has been comparatively under-utilized for diverse materials and medical applications owing to the lack of chromophores that afford stability, solubility, synthetic malleability, and tunable photophysical features. Bacteriochlorins are attractive candidates in this regard; however, preparation via modification of naturally occurring bacteriochlorophylls or reduction of porphyrins or chlorins has proved cumbersome. To overcome such limitations, a dibromobacteriochlorin (BC-Br3Br13) was prepared de novo by the acid-catalyzed condensation of an 8-bromodihydrodipyrrin-acetal. BC-Br3Br13 bears (1) a geminal dimethyl group in each reduced ring to block adventitious dehydrogenation, and (2) bromo groups at the 3- and 13-positions for further chemical modifications. BC-Br3Br13 was subjected to four types of Pd-mediated coupling reaction (Suzuki, Stille, Sonogashira, dehalogenation) to give bacteriochlorins bearing substituents at the 3- and 13-positions (phenyl, vinyl, acetyl, phenylethynyl), and a benchmark bacteriochlorin lacking such substituents. The 3,13-divinylbacteriochlorin was transformed to the 3,13-diformylbacteriochlorin. Depending on the substituents at the 3- and 13-positions, the position of the long-wavelength absorption maximum (Qy(0,0) band) lies between 713 and 771 nm, the fluorescence emission maximum lies between 717 and 777 nm, and the fluorescence quantum yield ranges from 0.15 to 0.070. The ability to introduce a wide variety of functional groups via Pd-mediated coupling reactions and the tunable absorption and emission spectral properties suggest that synthetic bacteriochlorins are viable candidates for a wide variety of photochemical applications. DA - 2008/// PY - 2008/// DO - 10.1039/b717803d VL - 32 IS - 6 SP - 947-958 SN - 1369-9261 ER - TY - JOUR TI - A versatile assay for the identification of RNA silencing suppressors based on complementation of viral movement AU - Powers, Jason G. AU - Sit, Tim L. AU - Qu, Feng AU - Morris, T. Jack AU - Kim, Kook-Hyung AU - Lommel, Steven A. T2 - MOLECULAR PLANT-MICROBE INTERACTIONS AB - The cell-to-cell movement of Turnip crinkle virus (TCV) in Nicotiana benthamiana requires the presence of its coat protein (CP), a known suppressor of RNA silencing. RNA transcripts of a TCV construct containing a reporter gene (green fluorescent protein) (TCV-sGFP) in place of the CP open reading frame generated foci of three to five cells. TCV CP delivered in trans by Agrobacterium tumefaciens infiltration potentiated movement of TCV-sGFP and increased foci diameter, on average, by a factor of four. Deletion of the TCV movement proteins in TCV-sGFP (construct TCVDelta92-sGFP) abolished the movement complementation ability of TCV CP. Other known suppressors of RNA silencing from a wide spectrum of viruses also complemented the movement of TCV-sGFP when delivered in trans by Agrobacterium tumefaciens. These include suppressors from nonplant viruses with no known plant movement function, demonstrating that this assay is based solely on RNA silencing suppression. While the TCV-sGFP construct is primarily used as an infectious RNA transcript, it was also subcloned for direct expression from Agrobacterium tumefaciens for simple quantification of suppressor activity based on fluorescence levels in whole leaves. Thus, this system provides the flexibility to assay for suppressor activity in either the cytoplasm or nucleus, depending on the construct employed. DA - 2008/7// PY - 2008/7// DO - 10.1094/MPMI-21-7-0879 VL - 21 IS - 7 SP - 879-890 SN - 1943-7706 KW - IVIS lumina KW - p8 KW - p9 KW - pPZP212 KW - VSR ER - TY - JOUR TI - A new branch of endoplasmic reticulum stress signaling and the osmotic signal converge on plant-specific asparagine-rich proteins to promote cell death AU - Costa, Maximiller D. L. AU - Reis, Pedro A. B. AU - Valente, Maria Anete S. AU - Irsigler, Andre S. T. AU - Carvalho, Claudine M. AU - Loureiro, Marcelo E. AU - Aragao, Francisco J. L. AU - Boston, Rebecca S. AU - Fietto, Luciano G. AU - Fontes, Elizabeth P. B. T2 - JOURNAL OF BIOLOGICAL CHEMISTRY AB - NRPs (N-rich proteins) were identified as targets of a novel adaptive pathway that integrates endoplasmic reticulum (ER) and osmotic stress signals based on coordinate regulation and synergistic up-regulation by tunicamycin and polyethylene glycol treatments. This integrated pathway diverges from the molecular chaperone-inducing branch of the unfolded protein response (UPR) in several ways. While UPR-specific targets were inversely regulated by ER and osmotic stresses, NRPs required both signals for full activation. Furthermore, BiP (binding protein) overexpression in soybean prevented activation of the UPR by ER stress inducers, but did not affect activation of NRPs. We also found that this integrated pathway transduces a PCD signal generated by ER and osmotic stresses that result in the appearance of markers associated with leaf senescence. Overexpression of NRPs in soybean protoplasts induced caspase-3-like activity and promoted extensive DNA fragmentation. Furthermore, transient expression of NRPs in planta caused leaf yellowing, chlorophyll loss, malondialdehyde production, ethylene evolution, and induction of the senescence marker gene CP1. This phenotype was alleviated by the cytokinin zeatin, a potent senescence inhibitor. Collectively, these results indicate that ER stress induces leaf senescence through activation of plant-specific NRPs via a novel branch of the ER stress response. NRPs (N-rich proteins) were identified as targets of a novel adaptive pathway that integrates endoplasmic reticulum (ER) and osmotic stress signals based on coordinate regulation and synergistic up-regulation by tunicamycin and polyethylene glycol treatments. This integrated pathway diverges from the molecular chaperone-inducing branch of the unfolded protein response (UPR) in several ways. While UPR-specific targets were inversely regulated by ER and osmotic stresses, NRPs required both signals for full activation. Furthermore, BiP (binding protein) overexpression in soybean prevented activation of the UPR by ER stress inducers, but did not affect activation of NRPs. We also found that this integrated pathway transduces a PCD signal generated by ER and osmotic stresses that result in the appearance of markers associated with leaf senescence. Overexpression of NRPs in soybean protoplasts induced caspase-3-like activity and promoted extensive DNA fragmentation. Furthermore, transient expression of NRPs in planta caused leaf yellowing, chlorophyll loss, malondialdehyde production, ethylene evolution, and induction of the senescence marker gene CP1. This phenotype was alleviated by the cytokinin zeatin, a potent senescence inhibitor. Collectively, these results indicate that ER stress induces leaf senescence through activation of plant-specific NRPs via a novel branch of the ER stress response. The unfolded protein response (UPR) 5The abbreviations used are: UPR, unfolded protein response; AZC, l-azetidine-2-carboxylic acid; BAP, 6-benzylaminopurine; BiP, binding protein; CNX, calnexin; CRT, calreticulin; DCD, development and cell death; ER, endoplasmic reticulum; MDA, malondialdehyde; NIG, NSP-interacting GTPase; NRPs, N-rich proteins; PCD, programmed cell death; PDI, protein-disulfide isomerase; PEG, polyethylene glycol; SMP, seed maturation protein; MES, 4-morpholineethanesulfonic acid; GFP, green fluorescent protein; TUNEL, terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling; WT, wild type; RACE, rapid amplification of cDNA ends. 5The abbreviations used are: UPR, unfolded protein response; AZC, l-azetidine-2-carboxylic acid; BAP, 6-benzylaminopurine; BiP, binding protein; CNX, calnexin; CRT, calreticulin; DCD, development and cell death; ER, endoplasmic reticulum; MDA, malondialdehyde; NIG, NSP-interacting GTPase; NRPs, N-rich proteins; PCD, programmed cell death; PDI, protein-disulfide isomerase; PEG, polyethylene glycol; SMP, seed maturation protein; MES, 4-morpholineethanesulfonic acid; GFP, green fluorescent protein; TUNEL, terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling; WT, wild type; RACE, rapid amplification of cDNA ends. is activated by stress conditions that disrupt the endoplasmic reticulum (ER) homeostasis and the proper folding and maturation of secretory proteins. This complex signaling cascade is conserved in eukaryotes, although the mechanism of signal transduction differs across species. The central transducer of the UPR is IRE1, a transmembrane kinase that appears to be the only means of signal transduction in yeast, but is part of a tripartite signaling response in animals, which also includes the ER-transmembrane transducers PERK and ATF6 (1Maa Y. Hendershot L.M. J. Chem. Neuroanatomy. 2004; 28: 51-65Crossref PubMed Scopus (313) Google Scholar). In plants, there is compelling evidence that the UPR operates through IRE1 (2Buzeli R.A.A. Cascardo J.C.M. Rodrigues L.A.Z. Andrade M.O. Loureiro M.E. Otoni W.C. Fontes E.P.B. Plant Mol. Biol. 2002; 50: 757-771Crossref PubMed Scopus (37) Google Scholar, 3Denecke J. Carlsson L.E. Vidal S. Hoglund A.-S. Ek B. van Zeiji M.J. Sinjorgo K.M.C. Palva E.T. Plant Cell. 1995; 7: 391-406Crossref PubMed Scopus (184) Google Scholar, 4Irsigler A.S. Costa M.D. Zhang P. Reis P.A.B. Dewey R.E. Boston R.S. Fontes E.P.B. BMC Genomics. 2007; 8: 431Crossref PubMed Scopus (84) Google Scholar, 5Kamauchi S. Nakatani H. Nakano C. Urade R. Febs. J. 2005; 272: 3461-3476Crossref PubMed Scopus (163) Google Scholar, 6Kirst M.E. Meyer D.J. Gibbon B.C. Jung R. Boston R.S. Plant Physiol. 2005; 138: 218-231Crossref PubMed Scopus (52) Google Scholar, 7Martinez I.M. Chrispeels M.J. Plant Cell. 2003; 15: 561-576Crossref PubMed Scopus (331) Google Scholar), but characterization of this cytoprotective signaling pathway is still incipient. The only known components of plant UPR are the putative proximal sensors that include two Arabidopsis Ire1p homologues and two ATF6-related proteins, designated AtbZIP28 and AtbZIP60 (8Koizumi N. Martinez I.M. Kimata Y. Kohno K. Sano H. Chrispeels M.J. Plant Physiol. 2001; 127: 949-962Crossref PubMed Scopus (186) Google Scholar, 9Iwata Y. Koizumi N. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 5280-5285Crossref PubMed Scopus (266) Google Scholar, 10Liu J.-X. Srivastava R. Che P. Howell S.H. Plant Cell. 2007; 19: 4111-4119Crossref PubMed Scopus (323) Google Scholar). Although the N-terminal domain of plant IRE1 homologues can replace functionally the yeast IRE1, downstream components of the IRE1 signaling are yet to be identified (8Koizumi N. Martinez I.M. Kimata Y. Kohno K. Sano H. Chrispeels M.J. Plant Physiol. 2001; 127: 949-962Crossref PubMed Scopus (186) Google Scholar). AtbZIP60 and AtbZIP28 have been described as ER stress-induced leucine zipper (bZIP) transcription factor genes that are anchored to the ER membrane under normal conditions and may serve as ER stress sensors and transducers (9Iwata Y. Koizumi N. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 5280-5285Crossref PubMed Scopus (266) Google Scholar, 10Liu J.-X. Srivastava R. Che P. Howell S.H. Plant Cell. 2007; 19: 4111-4119Crossref PubMed Scopus (323) Google Scholar). Upon sensing the ER stress, AtbZIP28 is proteolytically released from the membrane and translocated to the nucleus by a mechanism that is predicted to be similar to that acting on mammalian ATF6 transducer (10Liu J.-X. Srivastava R. Che P. Howell S.H. Plant Cell. 2007; 19: 4111-4119Crossref PubMed Scopus (323) Google Scholar). Expression of a truncated form of either AtbZIP28 or AtbZIP60, harboring the bZIP domain, up-regulates UPR target genes under normal conditions (9Iwata Y. Koizumi N. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 5280-5285Crossref PubMed Scopus (266) Google Scholar, 10Liu J.-X. Srivastava R. Che P. Howell S.H. Plant Cell. 2007; 19: 4111-4119Crossref PubMed Scopus (323) Google Scholar). The UPR up-regulated chaperone BiP (binding protein) is central to this cytoprotective response as the indirect sensor of alterations in the ER environment. In addition to this role as molecular chaperone, in mammalian cells, BiP has been shown to regulate the UPR by controlling the activation status of the three transducers, IRE1, PERK, and ATF6 (11Malhotra J.D. Kaufman R.J. Semin. Cell Dev. Biol. 2007; 18: 716-731Crossref PubMed Scopus (759) Google Scholar). Under normal conditions, the luminal domains of these sensors are occupied with BiP, which is recruited by unfolded proteins upon ER stress. The dissociation of BiP promotes oligomerization and phosphorylation of IRE1 and PERK, as well as relocalization of ATF6 to the Golgi where its transcriptional domain is proteolytically released from the membrane into the nucleus. Although the UPR has not been extensively characterized in plants, conserved aspects between mammalian and plant UPR include its negative regulation by BiP, possibly through interaction with the putative proximal sensors, and the BiP cytoprotective role (12Leborgne-Castel N. Jelitto-Van Dooren E. Crofts A.J. Denecke J. Plant Cell. 1999; 11: 459-469Crossref PubMed Scopus (152) Google Scholar). In fact, we have previously demonstrated that BiP confers tolerance to water deficit in transgenic plants (13Alvim F.C. Carolino S.M.B. Cascardo J.C.M. Nunes C.C. Martinez C.A. Otoni W.C. Fontes E.P.B. Plant Physiol. 2001; 126: 1042-1054Crossref PubMed Scopus (185) Google Scholar). Constant challenges by adverse conditions and environmental stresses pose major constraints for development, growth, and productivity of plants. Our knowledge about stress-specific adaptive responses and cross-talk between signaling cascades has advanced considerably with genome-wide analyses and expression-profiling studies in different plant species under different stress conditions (14Denekamp M. Smeekens S.C. Plant Physiol. 2003; 132: 1415-1423Crossref PubMed Scopus (158) Google Scholar, 15Kreps J.A. Wu Y.J. Chang H.S. Zhu T. Wang X. Harper J.F. Plant Physiol. 2002; 130: 2129-2141Crossref PubMed Scopus (1190) Google Scholar, 16Seki M. Narusaka M. Ishida J. Nanjo T. Fujita M. Oono Y. Kamiya A. Nakajima M. Enju A. Sakurai T. Satou M. Akiyama K. Taji T. Yamaguchi-Shinozaki K. Carninci P. Kawai J. Hayashizaki Y. Shinozaki K. Plant J. 2002; 31: 279-292Crossref PubMed Scopus (1613) Google Scholar). A common theme that has emerged is that plants transduce environmental signals through integrated networks between different stress-induced adaptive responses. Despite the potential of ER stress response to accommodate adaptive pathways, its integration with environmental-induced responses is poorly understood in plants. Recently, we performed global expression profiling on soybean leaves exposed to polyethylene glycol (PEG) treatment or to UPR inducers to identify integrated networks between osmotic and ER stress-induced adaptive responses (4Irsigler A.S. Costa M.D. Zhang P. Reis P.A.B. Dewey R.E. Boston R.S. Fontes E.P.B. BMC Genomics. 2007; 8: 431Crossref PubMed Scopus (84) Google Scholar). In addition to uncovering specific responses to ER stress and osmotically regulated changes, a small proportion (5.5%) of the up-regulated genes represented a shared response that seemed to integrate the two signaling pathways. These co-regulated genes had similar induction kinetics and a synergistic response to the combination of osmotic- and ER stress-inducing treatments. Thus, they were considered to be downstream targets of the integrating pathway. Two ESTs (N-richI and N-richII), which showed the strongest synergistic induction, were homologous to genes encoding asparagine-rich proteins that have a plant-specific development and cell death (DCD) domain. Here we investigated the possibility that the integrated pathway might transduce a programmed cell death (PCD) signal generated by ER and osmotic stress. We demonstrated that induction of target genes, such as N-richI and N-richII ESTs, by ER stress-inducing agents occurs via a pathway distinct from the UPR. We also found that the N-rich proteins are critical mediators of osmotic- and ER stress-induced cell death in plants. Plant Growth, Soybean Suspension Cells, and Stress Treatments—Soybean (Glycine max) seeds (cultivar Conquista) were germinated in soil and grown under greenhouse conditions (avg: 21 °C, max: 31 °C, min: 15 °C) under natural conditions of light, 70% relative humidity, and approximately equal day and night length. Two weeks after germination, the seedlings were transferred to a 2-ml 10 μg/ml tunicamycin (Sigma) solution (DMSO, as control). After 12 and 24 h of treatment, the leaves were harvested, immediately frozen in liquid N2, and stored at -80 °C until use. Alternatively, the aerial portions of 3-week-old plants were excised below the cotyledons and directly placed into 15 ml of 10% (w/v) polyethylene glycol (PEG; MW 8000, Sigma), 10 μg/ml tunicamycin (Sigma), or 50 mm l-azetidine-2-carboxylic acid (AZC, Sigma) solutions. The first trifoliate leaves were harvested after 4, 10, or 16 h of PEG and tunicamycin treatments, then immediately frozen in liquid N2, and stored at -80 °C until use. Each stress treatment and RNA extraction were replicated in three independent experiments. Cotyledons cells from the soybean variety Conquista were cultured as described previously (17Cascardo J.C.M. Almeida R.S. Buzeli R.A.A. Carolino S.M.B. Otoni W.C. Fontes E.P.B. J. Biol. Chem. 2000; 275: 14494-14500Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar). Cells were subcultured every 10 days by diluting the culture 1:10 in fresh MS medium. All treatments were performed on 5-day-old subcultures. Tunicamycin was added to cultures by dilution of a 5 mg/ml stock in DMSO into normal growth medium to 10 μg/ml and incubated for the intervals indicated in the figure legends. For PEG-induced dehydration, the cells were washed and then cultured with normal growth medium containing 10% (w/v) PEG-8000 for the indicated intervals. Suspension cells were directly treated with 0.5 μg/ml cycloheximide, 10 μm BAP (6-benzylaminopurine), and 10 μm zeatin for the intervals indicated in the figure legends. After treatments, the cells were filtrated under vacuum, washed with 0.25 m NaCl, and immediately frozen in liquid N2. Rapid Amplification of 3′-cDNA Ends (3′-RACE) and NRP-B cDNA Cloning—3′-RACE was performed using the GeneRacer kit (Invitrogen). Total RNA isolated from PEG-treated soybean leaves was used for reverse transcription, and an N-richI EST-specific primer (FwNRich1, TACAGGCATCCAATTTGGCGAACC) and oligo dT primer from the kit were used in the polymerase chain reaction. The amplified fragment was cloned in pCR4 to generate pCR4-NRPB, which harbors the full-length NRP-B cDNA. Plasmid Construction—For transient expression in protoplasts, NRP-B and NRP-A (gi:57898928) cDNAs were amplified and inserted into the BglII and EcoRI sites of pMON921 (18Fontes E.P.B. Eagle P.A. Sipe P.S. Luckow V.A. Hanley-Bowdoin L. J. Biol. Chem. 1994; 269: 8459-8465Abstract Full Text PDF PubMed Google Scholar). The resulting clones pUFV967 and pUFV849 contain NRP-B or NRP-A cDNAs, respectively, under the control of the cauliflower mosaic virus 35S promoter and the 3′-end of the pea E9 rbcS gene. For agroinfiltration in tobacco leaves, NRP-A and NRP-B cDNAs were amplified by PCR and introduced by recombination into the entry vector pCR8/GW/TOPO (Invitrogen) to generate pUFV908 and pUFV874, respectively. NRP-A and NRP-B cDNAs were then transferred by recombination to the plant transformation binary vector 35S-YFP-cassetteA-Nos-pCAMBIA1300 yielding pUFV937 (35S:NRP-B) and pUFV938 (35S:NRP-A), which contain NRP-B or NRP-A cDNA, respectively, under the control of the 35S promoter and 3′-ends of nos. For subcellular localization, NRP-A and NRP-B cDNAs were amplified by PCR with appropriate extensions and introduced by recombination into the entry vector pDONR201 (Invitrogen) and then transferred to the binary vector pK7FWG2 to generate pK7F-NRP-B and pK7F-NRP-A, which contain the GFP gene fused in-frame after the last codon of the respective cDNAs. The U16322 plasmid harboring a full-length NIG (NSP-interacting GTPase, Ref. 19Carvalho, C. M., Fontenelle, M. R., Florentino, L. H., Santos, A. A., Zerbini, F. M., and Fontes, E. P. B. (2008) Plant J., in pressGoogle Scholar). cDNA was obtained from the Arabidopsis Biological Resource Center (ABRC) and used as a control. For this purpose, the full-length NIG cDNA was amplified by PCR from U16322 cDNA, cloned into pDONR201 and then transferred to the binary vector pK7WG2 to obtain pK7-NIG, which contains NIG cDNA under the control of the CaMV 35S promoter. Real-time RT-PCR Analysis—For quantitative RT-PCR, total RNA was extracted from frozen leaves or cells with TRIzol (Invitrogen) according to the instructions from the manufacturer. The RNA was treated with 2 units of RNase-free DNase (Promega) and further purified through RNeasy Mini kit (Qiagen) columns. First-strand cDNA was synthesized from 4 μg of total RNA using oligo-dT(18) and Transcriptase Reversa M-MLV (Invitrogen), according to the manufacturer's instructions. Real-time RT-PCR reactions were performed on an ABI7500 instrument (Applied Biosystems), using SYBR® Green PCR Master Mix (Applied Biosystems). The amplification reactions were performed as follows: 2 min at 50 °C, 10 min at 95 °C, and 40 cycles of 94 °C for 15 s and 60 °C for 1 min. To confirm quality and primer specificity, we verified the size of amplification products after electrophoresis through a 1.5% agarose gel, and analyzed the Tm (melting temperature) of amplification products in a dissociation curve, performed by the ABI7500 instrument. The primers used are listed in supplemental Table S1. For quantitation of gene expression in soybean protoplasts and seedlings, we used RNA helicase (4Irsigler A.S. Costa M.D. Zhang P. Reis P.A.B. Dewey R.E. Boston R.S. Fontes E.P.B. BMC Genomics. 2007; 8: 431Crossref PubMed Scopus (84) Google Scholar) as the endogenous control gene for data normalization in real-time RT-PCR analysis. For quantitation of gene expression in tobacco leaves, we used actin as a control gene. Fold variation, which is based on the comparison of the target gene expression (normalized to the endogenous control) between experimental and control samples, was quantified using the comparative Ct method: 2−^ - (ΔCtTreatment - ΔCtControl). The absolute gene expression was quantified using the 2-ΔCT method, and values were normalized to the endogenous control. Soybean Transformation—A plant expression cassette containing the BiPD gene was constructed by insertion of the 2.0-kb XbaI cDNA insert of pUFV24 (17Cascardo J.C.M. Almeida R.S. Buzeli R.A.A. Carolino S.M.B. Otoni W.C. Fontes E.P.B. J. Biol. Chem. 2000; 275: 14494-14500Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar) into the pBS35SdAMVNOS2 vector. The resulting clone pBS35SdAMVNOS2-BiP contains the BiP cDNA under control of a duplicated cauliflower mosaic virus 35S promoter with an enhancer region from the alfalfa mosaic virus and the polyadenylation signal of the nos gene. The Arabidopsis thaliana ahas gene (that confers tolerance to the herbicide imazapyr) was removed from the vector pAC321 (20Aragão F.J.L. Sarokin L. Vianna G.R. Rech E.L. Theor. Appl. Genet. 2000; 101: 1-6Crossref Scopus (111) Google Scholar) with XbaI and inserted into the vector pFACM1 to generate pFACMahas. The BiPD expression cassette was released with SalI and NotI from pBS35SdAMVNOS2-BiP and cloned into the vector pFACMahasm to yield pahasBip. The vector pahasBip was used to transform soybean (cv. Conquista) as previously described (20Aragão F.J.L. Sarokin L. Vianna G.R. Rech E.L. Theor. Appl. Genet. 2000; 101: 1-6Crossref Scopus (111) Google Scholar). Primary transformants were selected by PCR using the primers bipsoy235 (5′-GAGAGACTAATTGGAGAGGCTG-3′) and bipsoy645c (5′-ATAGGCAATGGCAGCAGCAGTG-3′), which amplify a 410-bp sequence from the BiPD gene coding region and cover an intron region from the genomic BiPD sequence. Segregation analyses of independently transformed soybean lines were performed by PCR, and accumulation of BiP was monitored in each subsequent generation by immunoblotting. RNA Gel Blotting—Total RNA was extracted from frozen leaves of control and tunicamycin-treated wild type and soybean transgenic seedlings, which were treated with tunicamycin or control DMSO for 24 h, with TRIzol (Invitrogen) according to the instructions from the manufacturer. Equal amounts of total RNA were denatured by formamide/formaldehyde and resolved on 1% agarose gels containing formaldehyde. The RNA was transferred to a nylon membrane by capillary transfer and immobilized by UV cross-linking (Stratalinker, Stratagene). The membrane was hybridized at high stringency conditions using the soyBiPD cDNA (17Cascardo J.C.M. Almeida R.S. Buzeli R.A.A. Carolino S.M.B. Otoni W.C. Fontes E.P.B. J. Biol. Chem. 2000; 275: 14494-14500Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar) as probe. The hybridization probe was radiolabeled with [α-32P]dCTP by random primed labeling (Amersham Biosciences). Autoradiography was performed at -80 °C using a Lightning-Plus intensifying screen. The amount of RNA and the integrity of ribosomal RNA were monitored by rehybridizing the membranes with an 18 S rDNA probe. Immunoblot Analysis—Total protein was extracted from control and tunicamycin-treated leaves of wild-type and soybean transgenic seedlings as previously described (17Cascardo J.C.M. Almeida R.S. Buzeli R.A.A. Carolino S.M.B. Otoni W.C. Fontes E.P.B. J. Biol. Chem. 2000; 275: 14494-14500Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar). SDS-PAGE was carried out, and the proteins were transferred from 10% SDS-polyacrylamide gels to nitrocellulose membranes by electroblotting. Immunoblot analyses were performed using polyclonal anti-BiP-carboxyl antibodies (2Buzeli R.A.A. Cascardo J.C.M. Rodrigues L.A.Z. Andrade M.O. Loureiro M.E. Otoni W.C. Fontes E.P.B. Plant Mol. Biol. 2002; 50: 757-771Crossref PubMed Scopus (37) Google Scholar), an anti-calreticulin serum (21Pagny S. Cabanes-Macheteau M. Gillikin J. Leborgne-Castel N. Lerouge P. Boston R.S. Faye L. Gomord V. Plant Cell. 2000; 12: 739-755Crossref PubMed Scopus (96) Google Scholar) or an anti-GmSBP2 serum (22Delú-Filho N. Pirovani C.P. Pedra J.H.F. Matrangolo F.S.V. Macêdo J.N.A. Otoni W.C. Fontes E.P.B. Plant Physiol. Biochem. 2000; 38 (353): 353Crossref Scopus (20) Google Scholar) at a 1:1000 dilution and a goat anti-rabbit IgG alkaline phosphatase conjugate (Sigma) at a 1:5000 dilution. Alkaline phosphatase activity was assayed using 5-bromo-4-chloro-3-indolyl phosphate (Sigma) and p-nitro blue tetrazolium (Sigma). Transient Expression in Soybean Protoplasts—Protoplasts were prepared from soybean suspension cells as essentially described by Fontes et al. (18Fontes E.P.B. Eagle P.A. Sipe P.S. Luckow V.A. Hanley-Bowdoin L. J. Biol. Chem. 1994; 269: 8459-8465Abstract Full Text PDF PubMed Google Scholar). The protoplasts were isolated 5 days after subculture by digestion for 3 h, under agitation at 40 rpm, with 0.5% cellulase, 0,5% macerozyme R-10, 0.1% pectolyase Y23, 0.6 m mannitol, 20 mm MES, pH 5.5. The extent of digestion was monitored by examining the cells microscopically at each 30-min interval. After filtration through nylon mesh of 65 μm, protoplasts were recovered by centrifugation, resuspended in 2 ml of 0.6 m mannitol, 20 mm MES, pH 5.5, separated by centrifugation in a sucrose gradient (20% (w/v) sucrose, 0.6 m mannitol, 20 mm MES, pH 5.5), and diluted into 2 ml of electroporation buffer (25 mm Hepes-KOH, pH 7.2, 10 mm KCl, 15 mm MgCl2, 0.6 m mannitol). Transient expression assays were performed by electroporation (250 V, 250 μF) of 10 μg of expression cassette DNA, and 30 μg of sheared salmon sperm DNA into 2 × 105 - 5 × 106 protoplasts in a final volume of 0.8 ml. Protoplasts were diluted into 8 ml of MS medium supplemented with 0.2 mg/ml 2,4-dichlorophenoxyacetic acid and 0.6 m mannitol, pH 5.5. After 36 h of incubation in the dark, the protoplasts were washed with 0.6 m mannitol, 20 mm MES, pH 5.5, and frozen in liquid N2 until use. Caspase-3-like Activity and in Situ Labeling of DNA Fragmentation (TUNEL)—Total protein was extracted from soybean cells 36-h post-electroporation. Caspase-3-like activity was determined using the ApoAlert® Caspase-3 Colorimetric Assay kit (Clontech) according to the manufacturer's instructions. The substrate was DEVD-pNA, and the inhibitor of caspase-3 activity was the synthetic tetrapeptide DEVD-fmk supplied by the kit. Free 3′-OH in the DNA was labeled by the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay using the ApoAlert DNA Fragmentation Assay kit (Clontech) as instructed by the manufacturer. Samples were observed with a Zeiss LSM 410 inverted confocal laser scanning microscope fitted with the configuration: excitation at 488 nm and emission at 515 nm. As a positive control, samples were treated with DNase1. Transient Overexpression in Nicotiana tabacum by Agrobacterium Infiltration—N. tabacum plants were grown in a greenhouse with natural day length illumination. Three weeks after germination, the plants were transferred to a growth chamber at 21 °C with a 16-hour light and 8-hour dark cycle. Agrobacterium infiltration was performed in the leaves of N. tabacum with Agrobacterium strain GV3101 carrying pUFV937 (35S: NRP-B) or pUFV938 (35S:NRP-A), NIG (35S:NIG), rpl10 (35S: rpL10) as previously described (23Batoko H. Zheng H.Q. Hawes C. Moore I. Plant Cell. 2000; 12: 2201-2218Crossref PubMed Scopus (462) Google Scholar). Subcellular Localization of Proteins—For subcellular localization of proteins, N. tabacun leaves were agroinoculated with pK7F-NRP-B and pK7F-NRP-A. About 72-h post-agroinfiltration, 1-cm2 leaf explants were excised incubated with 0.6 m mannitol for 10 min, and GFP fluorescence patterns were examined in epidermal cells with ×40 oil immersion objective and a Zeiss LSM510 META inverted laser scanning microscope equipped with an argon laser as excitation source. For imaging GFP, the 455-nm excitation line and the 500–530-nm band pass filter were used. The pinhole was usually set to give a 1–1.5-μm optical slice. Post-acquisition image processing was done using the LSM 5 Browser software (Carl-Zeiss) and Adobe Photoshop (Adobe Systems). Microsomal fractions were prepared from agroinoculated leaves, as described (24Pirovani C.P. Macêdo J.N.A. Contim L.A.S. Matrangolo F.S.V. Loureiro M.E. Fontes E.P.B. Eur. J. Biochem. 2002; 269: 3998-4008Crossref PubMed Scopus (16) Google Scholar), and immunoblotted with an anti-GFP serum. Determination of Chlorophyll Content and Lipid Peroxidation—Total chlorophyll content was determined spectrophotometrically at 663 and 646 nm after quantitative extraction from individual leaves with 80% (v/v) acetone in the presence of ∼1 mg of Na2CO3 (25Lichtenthaler H.K. Methods Enzymol. 1987; 148: 350-382Crossref Scopus (8704) Google Scholar). The extent of lipid peroxidation in leaves was estimated by measuring the amount of MDA (malondialdehyde, a decomposition product of the oxidation of polyunsaturated fatty acids). MDA content was determined by the reaction of thiobarbituric acid (TBA) as described by Cakmak and Horst (26Cakmak I. Horst W.J. Physiol. Plantarum. 1991; 83: 463-468Crossref Scopus (1301) Google Scholar). Ethylene Determination—For ethylene measurements as a function of NRP-A and NRP-B expression, whole leaves of 3-week-old tobacco plants were agroinoculated with pK7-NRP-A, pK7-NRP-B, or control binary vector. Agroinfiltrated tobacco plants were placed in 25-ml flasks containing a Petri dish with 3 ml of 1 m HgClO4·4H2O and sealed with rubber serum caps. After 5 days, the solution was collected into a sealed assay tube, and ethylene was released with the injection of 1 ml of 1 m KCl. 1 ml of gas was collected with a syringe, and ethylene was measured on a model 5840A gas chromatography system (Hewlett-Packard Canada Ltda) equipped with a flame ionization detector. A Porapak N (80–100 mesh, 100 cm × 6 mm) column was used at 60 °C. The injection port and detector temperatures were 100 and 150 °C, respectively. The flow of nitrogen, air, and hydrogen was at 30, 130, and 20 ml min-1, respectively. N-richI and N-richII ESTs Are Both Encoded by the NRP-B Gene—The N-richI and N-richII ESTs correspond to different regions of a contiguous soybean genomic sequence (scaffold_78:2951370.2949024). To determine if both ESTs corresponded to the same mRNAs, we obtained the full sequences of N-richI and N-richII cDNAs through RACE. A comparison of the cDNA sequences revealed that these ESTs represented the same gene which we designated NRP-B (N-rich protein B; supplemental Fig. S1A). The deduced NRP-B protein has an estimated Mr of 37092 and pI 7.05 and is most closely related to the previously identified NRP from soybean (27Ludwig A.A. Tenhaken R. Eur. J. Plant Pathol. 2001; 107: 323-336Crossref Scopus (24) Google Scholar), here designated NRP-A. The two NRPs share a highly conserved C-terminal DCD domain in addition to a high content of asparagine residues at their more divergent N termini. This structural organization places NRP-A and NRP-B in the subgroup I of plant-specific DCD-containing proteins (28Tenhaken R. Doerks T. Bork P. BMC Bioinformatics. 2005; 6: 169Crossref PubMed Scopus (33) Google Scholar). Analysis of several subgroup I DCD domain-containing proteins by ClustalW showed that NRP-A and NRP-B form a subgroup with the Arabidopsis protein of unknown function encoded by At5g42050 (gi 231 18422167, supplemental Fig. S1B). An examination of available microarray data indicated that the At5g42050 locus is induced by cycloheximide, osmotic stress, salt stress, and during senescence (GeneInvestigator). Integration of ER Stress and Osmotic Signals Leads to Maximal Express DA - 2008/7/18/ PY - 2008/7/18/ DO - 10.1074/jbc.M802654200 VL - 283 IS - 29 SP - 20209-20219 SN - 1083-351X ER - TY - JOUR TI - Tailoring a bacteriochlorin building block with cationic, amphipathic, or lipophilic substituents AU - Ruzie, Christian AU - Krayer, Michael AU - Balasubramanian, Thiagarajan AU - Lindsey, Jonathan S. T2 - JOURNAL OF ORGANIC CHEMISTRY AB - Bacteriochlorins are attractive candidates for photodynamic therapy (PDT) of diverse medical indications owing to their strong absorption in the near-infrared (NIR) region, but their use has been stymied by lack of access to stable, synthetically malleable molecules. To overcome these limitations, a synthetic free base 3,13-dibromobacteriochlorin (BC-Br3Br13) has been exploited as a building block in the synthesis of diverse bacteriochlorins via Pd-mediated coupling reactions (Sonogashira, Suzuki, and reductive carbonylation). Each bacteriochlorin is stable to adventitious dehydrogenation by virtue of the presence of a geminal dimethyl group in each pyrroline ring. The target bacteriochlorins bear cationic, lipophilic, or amphipathic substituents at the 3- and 13- (β-pyrrolic) positions. A dicarboxybacteriochlorin was converted to amide derivatives via the intermediate diacid chloride. A diformylbacteriochlorin was subjected to reductive amination to give aminomethyl derivatives. A set of 3,5-disubstituted aryl groups bearing lipophilic or amphipathic groups was introduced via Suzuki coupling. Altogether 22 free base bacteriochlorins have been prepared. Eight aminoalkylbacteriochlorins were quaternized with methyl iodide at two or four amine sites per molecule, which resulted in water solubility. Each bacteriochlorin exhibits a Qy absorption band in the range of 720−772 nm. The ability to introduce a wide variety of peripheral functional groups makes these bacteriochlorins attractive candidates for diverse applications in photomedicine including PDT in the NIR region. DA - 2008/8/1/ PY - 2008/8/1/ DO - 10.1021/jo800736c VL - 73 IS - 15 SP - 5806-5820 SN - 1520-6904 ER - TY - PAT TI - Synthesis of porphyrins designed for attachment to electroactive surfaces via one or more carbon tethers AU - Lindsey, J. S. AU - Loewe, R. S. C2 - 2008/// DA - 2008/// PY - 2008/// ER - TY - JOUR TI - Rational or statistical routes from 1-acyldipyrromethanes to meso-substituted porphyrins. Distinct patterns, multiple pyridyl substituents, and amphipathic architectures AU - Dogutan, Dilek Kiper AU - Ptaszek, Marcin AU - Lindsey, Jonathan S. T2 - JOURNAL OF ORGANIC CHEMISTRY AB - New methodology is described for the synthesis of porphyrins bearing four (A4, cis-A2B2, cis-ABC2, trans-A2B2) or fewer (A, cis-AB, cis-A2, trans-A2) meso substituents. The method entails condensation of two 1-acyldipyrromethanes in the presence of a metal salt (MgBr2, 3 mol equiv) and a noncoordinating base (DBU, 10 mol equiv) in a noncoordinating solvent (toluene) with heating (conventional or microwave irradiation) and exposure to air. The rational synthesis of trans-A2B2- or trans-A2-porphyrins was achieved via condensation of two identical 1-acyldipyrromethanes. The statistical synthesis of various meso-substituted porphyrins was achieved via condensation of two nonidentical 1-acyldipyrromethanes. Both routes possess attractive features including (1) no scrambling, (2) good yield (up to 60%) at high concentration (100 mM) for the macrocycle-forming step, (3) reasonable scope (aryl, heteroaryl, alkyl, or no substituent), (4) short reaction time (∼2 h) via microwave irradiation, (5) magnesium porphyrins as the products, which easily undergo demetalation, and (6) facile chromatographic purification. A key advantage of the statistical route is to obtain a cis-substituted porphyrin without the corresponding trans isomer. For example, reaction of an A/B-substituted 1-acyldipyrromethane and the fully unsubstituted 1-formyldipyrromethane gave the magnesium chelates of three porphyrins: the trans-A2B2-porphyrin, the “hybrid” cis-AB-porphyrin, and porphine (no trans-AB-porphyrin can form), which were readily demetalated and separated as the free base species. Altogether 26 1-acyldipyrromethanes and 26 target porphyrins have been prepared, including many with two different pyridyl substituents. One set of amphipathic porphyrins includes cis-A2B2- or cis-A2BC-porphyrins wherein A = pentyl and B/C = pyridyl (o-, m-, p-). Taken together, the rational and statistical routes enable facile conversion of readily available 1-acyldipyrromethanes to diverse porphyrins bearing 1−4 meso substituents for which access is limited via other methods. DA - 2008/8/15/ PY - 2008/8/15/ DO - 10.1021/jo800588n VL - 73 IS - 16 SP - 6187-6201 SN - 0022-3263 ER - TY - JOUR TI - High-throughput gene and SNP discovery in Eucalyptus grandis, an uncharacterized genome AU - Novaes, Evandro AU - Drost, Derek R. AU - Farmerie, William G. AU - Pappas, Georgios J., Jr. AU - Grattapaglia, Dario AU - Sederoff, Ronald R. AU - Kirst, Matias T2 - BMC GENOMICS AB - Abstract Background Benefits from high-throughput sequencing using 454 pyrosequencing technology may be most apparent for species with high societal or economic value but few genomic resources. Rapid means of gene sequence and SNP discovery using this novel sequencing technology provide a set of baseline tools for genome-level research. However, it is questionable how effective the sequencing of large numbers of short reads for species with essentially no prior gene sequence information will support contig assemblies and sequence annotation. Results With the purpose of generating the first broad survey of gene sequences in Eucalyptus grandis , the most widely planted hardwood tree species, we used 454 technology to sequence and assemble 148 Mbp of expressed sequences (EST). EST sequences were generated from a normalized cDNA pool comprised of multiple tissues and genotypes, promoting discovery of homologues to almost half of Arabidopsis genes, and a comprehensive survey of allelic variation in the transcriptome. By aligning the sequencing reads from multiple genotypes we detected 23,742 SNPs, 83% of which were validated in a sample. Genome-wide nucleotide diversity was estimated for 2,392 contigs using a modified theta (θ) parameter, adapted for measuring genetic diversity from polymorphisms detected by randomly sequencing a multi-genotype cDNA pool. Diversity estimates in non-synonymous nucleotides were on average 4x smaller than in synonymous, suggesting purifying selection. Non-synonymous to synonymous substitutions (Ka/Ks) among 2,001 contigs averaged 0.30 and was skewed to the right, further supporting that most genes are under purifying selection. Comparison of these estimates among contigs identified major functional classes of genes under purifying and diversifying selection in agreement with previous researches. Conclusion In providing an abundance of foundational transcript sequences where limited prior genomic information existed, this work created part of the foundation for the annotation of the E. grandis genome that is being sequenced by the US Department of Energy. In addition we demonstrated that SNPs sampled in large-scale with 454 pyrosequencing can be used to detect evolutionary signatures among genes, providing one of the first genome-wide assessments of nucleotide diversity and Ka/Ks for a non-model plant species. DA - 2008/6/30/ PY - 2008/6/30/ DO - 10.1186/1471-2164-9-312 VL - 9 SP - SN - 1471-2164 ER - TY - JOUR TI - Genetic variation and population structure in Fraser fir (Abies fraseri): a microsatellite assessment of young trees AU - Potter, Kevin M. AU - Frampton, John AU - Josserand, Sedley A. AU - Nelson, C. Dana T2 - CANADIAN JOURNAL OF FOREST RESEARCH-REVUE CANADIENNE DE RECHERCHE FORESTIERE AB - The island-like populations of Fraser fir ( Abies fraseri (Pursh) Poir.) have been isolated since the end of the late-Wisconsinian glaciation on the highest peaks of the Southern Appalachian Mountains and therefore offer an opportunity to investigate the genetic dynamics of a long-fragmented forest tree species. An analysis of eight microsatellite markers isolated from Fraser fir found that the species was out of Hardy–Weinberg equilibrium, with a significant deficiency of heterozygosity and a high degree of inbreeding (F IS = 0.223) relative to other conifers, perhaps associated in part with the young life stage of the trees included in the analysis. The analysis detected a significant but small amount of genetic differentiation among Fraser fir populations (F ST = 0.004) and revealed that the geographical and latitudinal distances between populations, but not population area, were significantly correlated with their pairwise genetic differences. Both gene flow and postglacial migration history may have influenced the genetic architecture of the species. The results will be useful in the genetic conservation of Fraser fir, a species experiencing severe mortality following infestation by an exotic insect. DA - 2008/8// PY - 2008/8// DO - 10.1139/X08-064 VL - 38 IS - 8 SP - 2128-2137 SN - 0045-5067 ER - TY - JOUR TI - Top-stop nipper reduces leader growth in fraser fir Christmas trees AU - Rutledge, M. E. AU - Frampton, J. AU - Hinesley, L. E. AU - Blank, G. T2 - HortTechnology DA - 2008/// PY - 2008/// VL - 18 IS - 2 SP - 256-260 ER - TY - JOUR TI - Targeting cell death in tumors by activating Caspases AU - MacKenzie, S. H. AU - Clark, A. C. T2 - Current Cancer Drug Targets DA - 2008/// PY - 2008/// VL - 8 IS - 2 SP - 98-109 ER - TY - JOUR TI - Soluble precipitable porphyrins for use in targeted molecular brachytherapy AU - Yao, Zhen AU - Borbas, K. Eszter AU - Lindsey, Jonathan S. T2 - NEW JOURNAL OF CHEMISTRY AB - In a new therapy that aims to concentrate and immobilize therapeutic radionuclides in nanoscale assemblies within solid tumors, a soluble precipitable reagent (SPR) is administered as the radionuclide carrier and is converted to non-diffusible precipitate by an enzyme located in tumor tissues. To meet the objective of such an SPR, we have prepared and examined a class of porphyrin–alkyldiphosphates that are soluble in aqueous solution and that are rendered insoluble upon removal of the two phosphate groups. The porphyrins examined herein are of the trans-AB architecture wherein the substituents are a bis(dihydroxyphosphoryloxy)alkyl group and a phenyl (or p-bromophenyl) group. Provisions for later incorporation of radionuclides have been established by preparation of the analogous copper chelate or the meso-iodo free-base porphyrin. Altogether, four porphyrins bearing a bis(dihydroxyphosphoryloxy)alkyl group were examined and found to exhibit satisfactory solubility in water (>1 mM). Dephosphorylation reactions have been carried out in vitro using the enzyme shrimp alkaline phosphatase. In each case, enzyme-induced precipitation was observed. The soluble-to-insoluble conversion has been examined by visual inspection, absorption spectroscopy, electrospray ionization mass spectrometry, and nephelometry using non-radiolabeled porphyrins. DA - 2008/// PY - 2008/// DO - 10.1039/b714127k VL - 32 IS - 3 SP - 436-451 SN - 1369-9261 ER - TY - PAT TI - Methods and intermediates for the synthesis of dipyrrin-substituted porphyrinic macrocycles AU - Yu, L. AU - Muthukumaran, K. AU - Sreedharan, P. AU - Lindsey, J. S. C2 - 2008/// DA - 2008/// PY - 2008/// ER - TY - PAT TI - Metal complexation of 1-acyldipyrromethanes and porphyrins formed therefrom AU - Lindsey, J. S. AU - Muthukumaran, K. AU - Sharada, D. S. AU - Muresan, A. Z. AU - Youngblood, W. J. C2 - 2008/// DA - 2008/// PY - 2008/// ER - TY - PAT TI - Formation of self-assembled monolayers of redox SAMs on silicon for molecular memory applications AU - Bocian, D. F. AU - Kuhr, W. G. AU - Lindsey, J. S. AU - Dabke, R. B. AU - Liu, Z. C2 - 2008/// DA - 2008/// PY - 2008/// ER - TY - JOUR TI - Diverse inhibitors of aflatoxin biosynthesis AU - Holmes, Robert A. AU - Boston, Rebecca S. AU - Payne, Gary A. T2 - APPLIED MICROBIOLOGY AND BIOTECHNOLOGY DA - 2008/3// PY - 2008/3// DO - 10.1007/s00253-008-1362-0 VL - 78 IS - 4 SP - 559-572 SN - 1432-0614 KW - aspergillus KW - secondary metabolism KW - oxidative stress KW - host resistance ER - TY - JOUR TI - Darwinian selection on a selfing locus (vol 306, pg 2081, 2004) AU - Shimizu, K. K. AU - Reininga, J. M. AU - Caicedo, A. L. AU - Mays, C. A. AU - Moore, R. C. AU - Olsen, K. M. AU - Ruzsa, S. AU - Coop, G. AU - Bustamante, C. D. AU - Purugganan, M. D. T2 - Science DA - 2008/// PY - 2008/// VL - 320 IS - 5873 SP - 176-176 ER - TY - PAT TI - Boron complexation strategy for use in manipulating 1-acyldipyrromethanes AU - Lindsey, J. S. AU - Muthukumaran, K. AU - Ptaszek, M. AU - Huma, H. Z. AU - S. C2 - 2008/// DA - 2008/// PY - 2008/// ER - TY - PAT TI - Synthesis of phosphono-substituted porphyrin compounds for attachment to metal oxide surfaces AU - Lindsey, J. S. AU - Loewe, R. S. AU - Muthukumaran, K. AU - Ambroise, A. C2 - 2008/// DA - 2008/// PY - 2008/// ER - TY - JOUR TI - Synthesis and excited-state photodynamics of a chlorin-bacteriochlorin dyad-through-space versus through-bond energy transfer in tetrapyrrole arrays AU - Muthiah, Chinnasamy AU - Kee, Hooi Ling AU - Diers, James R. AU - Fan, Dazhong AU - Ptaszek, Marcin AU - Bocian, David F. AU - Holten, Dewey AU - Lindsey, Jonathan S. T2 - PHOTOCHEMISTRY AND PHOTOBIOLOGY AB - Abstract Understanding energy transfer among hydroporphyrins is of fundamental interest and essential for a wide variety of photochemical applications. Toward this goal, a synthetic free base ethynylphenylchlorin has been coupled with a synthetic free base bromobacteriochlorin to give a phenylethyne‐linked chlorin–bacteriochlorin dyad ( FbC‐pe‐FbB ). The chlorin and bacteriochlorin are each stable toward adventitious oxidation because of the presence of a geminal dimethyl group in each reduced pyrrole ring. A combination of static and transient optical spectroscopic studies indicate that excitation into the Q y band of the chlorin constituent (675 nm) of FbC‐pe‐FbB in toluene results in rapid energy transfer to the bacteriochlorin constituent with a rate of ∼(5 ps) −1 and efficiency of >99%. The excited bacteriochlorin resulting from the energy‐transfer process in FbC‐pe‐FbB has essentially the same fluorescence characteristics as an isolated monomeric reference compound, namely a narrow (12 nm fwhm) fluorescence emission band at 760 nm and a long‐lived (5.4 ns) Q y excited state that exhibits a significant fluorescence quantum yield (Φ f = 0.19). Förster calculations are consistent with energy transfer in FbC‐pe‐FbB occurring predominantly by a through‐space mechanism. The energy‐transfer characteristics of FbC‐pe‐FbB are compared with those previously obtained for analogous phenylethyne‐linked dyads consisting of two porphyrins or two oxochlorins. The comparisons among the sets of dyads are facilitated by density functional theory calculations that elucidate the molecular‐orbital characteristics of the energy donor and acceptor constituents. The electron‐density distributions in the frontier molecular orbitals provide insights into the through‐bond electronic interactions that can also contribute to the energy‐transfer process in the different types of dyads. DA - 2008/// PY - 2008/// DO - 10.1111/j.1751-1097.2007.00258.x VL - 84 IS - 3 SP - 786-801 SN - 1751-1097 ER - TY - JOUR TI - Genomic analysis of closely related astroviruses AU - Strain, Errol AU - Kelley, Laura A. AU - Schultz-Cherry, Stacey AU - Muse, Spencer V. AU - Koci, Matthew D. T2 - JOURNAL OF VIROLOGY AB - ABSTRACT To understand astrovirus biology, it is essential to understand factors associated with its evolution. The current study reports the genomic sequences of nine novel turkey astrovirus (TAstV) type 2-like clinical isolates. This represents, to our knowledge, the largest genomic-length data set available for any one astrovirus type. The comparison of these TAstV sequences suggests that the TAstV species contains multiple subtypes and that recombination events have occurred across the astrovirus genome. In addition, the analysis of the capsid gene demonstrated evidence for both site-specific positive selection and purifying selection. DA - 2008/5// PY - 2008/5// DO - 10.1128/JVI.01993-07 VL - 82 IS - 10 SP - 5099-5103 SN - 0022-538X ER - TY - JOUR TI - From classical genetics to quantitative genetics to systems biology: Modeling epistasis AU - Aylor, D. L. AU - Zeng, Z. B. T2 - PLoS Genetics DA - 2008/// PY - 2008/// VL - 4 IS - 3 ER - TY - JOUR TI - Approximate factorization of multivariate polynomials using singular value decomposition AU - Kaltofen, Erich AU - May, John P. AU - Yang, Zhengfeng AU - Zhi, Lihong T2 - JOURNAL OF SYMBOLIC COMPUTATION AB - We describe the design, implementation and experimental evaluation of new algorithms for computing the approximate factorization of multivariate polynomials with complex coefficients that contain numerical noise. Our algorithms are based on a generalization of the differential forms introduced by W. Ruppert and S. Gao to many variables, and use singular value decomposition or structured total least squares approximation and Gauss–Newton optimization to numerically compute the approximate multivariate factors. We demonstrate on a large set of benchmark polynomials that our algorithms efficiently yield approximate factorizations within the coefficient noise even when the relative error in the input is substantial (10−3). DA - 2008/5// PY - 2008/5// DO - 10.1016/j.jsc.2007.11.005 VL - 43 IS - 5 SP - 359-376 SN - 0747-7171 KW - multivariate polynomial factorization KW - approximate factorization KW - singular value decomposition KW - numerical algebra KW - Gauss-Newton optimization ER - TY - JOUR TI - X-LRT: A likelihood approach to estimate genetic risks and test association with X-linked markers using a case-parents design AU - Zhang, Li AU - Martin, Eden R. AU - Chung, Ren-Hua AU - Li, Yi-Ju AU - Morris, Richard W. T2 - GENETIC EPIDEMIOLOGY AB - Abstract Recently, there has been interest in family‐based tests of association to identify X‐chromosome genes. However, none of the approaches allow for estimation of genetic risks. We propose a likelihood approach to estimate disease‐related marker relative risks and test genotype association using a case‐parents design. The test uses nuclear families with a single affected proband and allows additional siblings and missing parental genotypes. Extension to a haplotype test is based on assumptions of random mating and multiplicative penetrance. We investigate power and type I error rate of the likelihood‐based test, using simulated data and apply our method to marker data from the monoamine oxidase A&B genes in families with Parkinson disease. We show how efficiency with missing parental information can be improved with additional sibling genotype information. Our likelihood approach offers great flexibility for testing different penetrance relationships within and between sexes. In addition, estimation of disease‐related marker relative risks provides a measure of the magnitude of X‐linked genetic effects on complex disorders. Genet. Epidemiol . 2008. © 2008 Wiley‐Liss, Inc. DA - 2008/5// PY - 2008/5// DO - 10.1002/gepi.20311 VL - 32 IS - 4 SP - 370-380 SN - 0741-0395 KW - sex linked KW - maximum-likelihood estimation KW - confidence interval KW - hypothesis test KW - family-based design ER - TY - BOOK TI - Pioneering women in plant pathology DA - 2008/// PY - 2008/// PB - St. Paul, Minn.: APS Press SN - 0890543593 ER - TY - JOUR TI - Measuring and partitioning the high-order linkage disequilibrium by multiple order Markov chains AU - Kim, Yunjung AU - Feng, Sheng AU - Zeng, Zhao-Bang T2 - GENETIC EPIDEMIOLOGY AB - A map of the background levels of disequilibrium between nearby markers can be useful for association mapping studies. In order to assess the background levels of linkage disequilibrium (LD), multilocus LD measures are more advantageous than pairwise LD measures because the combined analysis of pairwise LD measures is not adequate to detect simultaneous allele associations among multiple markers. Various multilocus LD measures based on haplotypes have been proposed. However, most of these measures provide a single index of association among multiple markers and does not reveal the complex patterns and different levels of LD structure. In this paper, we employ non-homogeneous, multiple order Markov Chain models as a statistical framework to measure and partition the LD among multiple markers into components due to different orders of marker associations. Using a sliding window of multiple markers on phased haplotype data, we compute corresponding likelihoods for different Markov Chain (MC) orders in each window. The log-likelihood difference between the lowest MC order model (MC0) and the highest MC order model in each window is used as a measure of the total LD or the overall deviation from the gametic equilibrium for the window. Then, we partition the total LD into lower order disequilibria and estimate the effects from two-, three-, and higher order disequilibria. The relationship between different orders of LD and the log-likelihood difference involving two different orders of MC models are explored. By applying our method to the phased haplotype data in the ENCODE regions of the HapMap project, we are able to identify high/low multilocus LD regions. Our results reveal that the most LD in the HapMap data is attributed to the LD between adjacent pairs of markers across the whole region. LD between adjacent pairs of markers appears to be more significant in high multilocus LD regions than in low multilocus LD regions. We also find that as the multilocus total LD increases, the effects of high-order LD tends to get weaker due to the lack of observed multilocus haplotypes. The overall estimates of first, second, third, and fourth order LD across the ENCODE regions are 64, 23, 9, and 3%. DA - 2008/5// PY - 2008/5// DO - 10.1002/gepi.20305 VL - 32 IS - 4 SP - 301-312 SN - 1098-2272 KW - multilocus linkage disequilibrium KW - multiple order Markov chains KW - partition of multilocus LID ER - TY - JOUR TI - Effects of counterion mobility, surface morphology, and charge screening on the electron-transfer rates of porphyrin monolayers AU - Jiao, Jieying AU - Nordfund, Eric AU - Lindsey, Jonathan S. AU - Bocian, David F. T2 - JOURNAL OF PHYSICAL CHEMISTRY C AB - The standard electron-transfer rate constants (k0) for the oxidation of porphyrin monolayers are reported for a number of solvent/electrolyte systems and electroactive surfaces. The goal is to explain the inverse correlation between the electron-transfer rates and the porphyrin surface concentration (Roth et al., J. Phys. Chem. B 2002, 106, 8639−8648). Each porphyrin is a zinc chelate and contains three meso-mesityl groups and a benzyl alcohol or benzyl thiol for surface attachment. The solvent/electrolyte systems include (i) the organic solvent propylene carbonate containing electrolytes with a common cation and anions of different size/mobility (PF6-, ClO4-, and Cl-) and (ii) neat ionic liquids with a common cation and anions of different size/mobility [(CF3SO2)2N- and (NC)2N-]. The substrates include Si(100), Au(111), and TiN. The k0 values observed using electrolytes with PF6-, ClO4-, and (CF3SO2)2N- counterions are similar to one another, whereas those observed using electrolytes Cl- and (NC)2N- counterions are 2−5 times faster. The faster rates for the latter anions are attributed to their smaller size/higher mobility. The k0 values observed for monolayers on Si(100) and Au(111) are similar to one another; the k0 values for monolayers on TiN are ∼5-fold faster. The faster rates for the TiN substrate are attributed to a rougher surface morphology (as determined via atomic force microscopy measurements) which results in an actual surface concentration that is lower than the concentration based on the geometrical area of a planar substrate. The k0 values determined for mixed monolayers where the electroactive porphyrin is co-deposited with an electroinactive porphyrin are dependent only on the concentration of the redox-active species, not on the total porphyrin concentration. This behavior is consistent with space-charge effects being the principal determinant of the inverse correlation between the electron-transfer rates and the porphyrin surface concentration. The space charge effects can be mitigated, but not eliminated, by using smaller, more mobile counterions and rougher surfaces. DA - 2008/4/17/ PY - 2008/4/17/ DO - 10.1021/jp800123u VL - 112 IS - 15 SP - 6173-6180 SN - 1932-7447 ER - TY - JOUR TI - Vector transmission of Bartonella species with emphasis on the potential for tick transmission AU - Billeter, S. A. AU - Levy, M. G. AU - Chomel, B. B. AU - Breitschwerdt, E. B. T2 - Medical and Veterinary Entomology AB - Bartonella species are gram-negative bacteria that infect erythrocytes, endothelial cells and macrophages, often leading to persistent blood-borne infections. Because of the ability of various Bartonella species to reside within erythrocytes of a diverse number of animal hosts, there is substantial opportunity for the potential uptake of these blood-borne bacteria by a variety of arthropod vectors that feed on animals and people. Five Bartonella species are transmitted by lice, fleas or sandflies. However, Bartonella DNA has been detected or Bartonella spp. have been cultured from numerous other arthropods. This review discusses Bartonella transmission by sandflies, lice and fleas, the potential for transmission by other vectors, and data supporting transmission by ticks. Polymerase chain reaction (PCR) or culture methods have been used to detect Bartonella in ticks, either questing or host-attached, throughout the world. Case studies and serological or molecular surveys involving humans, cats and canines provide indirect evidence supporting transmission of Bartonella species by ticks. Of potential clinical relevance, many studies have proposed co-transmission of Bartonella with other known tick-borne pathogens. Currently, critically important experimental transmission studies have not been performed for Bartonella transmission by many potential arthropod vectors, including ticks. DA - 2008/3// PY - 2008/3// DO - 10.1111/j.1365-2915.2008.00713.x VL - 22 IS - 1 SP - 1-15 J2 - Med Vet Entomol LA - en OP - SN - 0269-283X 1365-2915 UR - http://dx.doi.org/10.1111/j.1365-2915.2008.00713.x DB - Crossref KW - Bartonella species KW - arthropods KW - DNA KW - PCR KW - ticks KW - vector competence KW - vector potential ER - TY - JOUR TI - Transcriptional analysis of Pinus sylvestris roots challenged with the ectomycorrhizal fungus Laccaria bicolor AU - Heller, G. AU - Adomas, A. AU - Li, G. S. AU - Osborne, J. AU - Van Zyl, L. AU - Sederoff, R. AU - Finlay, R. D. AU - Stenlid, J. AU - Asiegbu, F. O. T2 - BMC Plant Biology DA - 2008/// PY - 2008/// VL - 8 ER - TY - JOUR TI - Mapping genes encoding microsomal omega-6 desaturase enzymes and their cosegregation with QTL affecting oleate content in soybean AU - Bachlava, Eleni AU - Dewey, Ralph E. AU - Auclair, Jerome AU - Wang, Sanbao AU - Burton, Joseph W. AU - Cardinal, Andrea J. T2 - CROP SCIENCE AB - The microsomal ω‐6 desaturase enzymes, which catalyze the desaturation of oleic acid to linoleic acid during fatty acid biosynthesis, are encoded by the FAD2‐1 and FAD2‐2 genes in soybean [ Glycine max (L.) Merr.]. Breeders aim to incorporate the high‐oleate trait into soybean germplasm in order to improve the nutritional value and oxidative stability of soybean oil. The objectives of this study were to map the isoforms of the FAD2‐1 and FAD2‐2 genes and investigate the association of these genetic loci with the oleate phenotype in three populations segregating for oleate content. The populations were grown in replicated multienvironment field trials. According to linkage analysis conducted for two of the populations, FAD2‐1A and FAD2‐1B mapped on Linkage Groups O and I, respectively, while the closely linked FAD2‐2A and FAD2‐2B isoforms mapped on Linkage Group L. Oleate quantitative trait loci with minor effects were detected in the proximity of FAD2‐1B and possibly FAD2‐2B on Linkage Groups I and L. Quantitative trait loci affecting maturity were also detected on chromosomal regions adjacent to the FAD2 genes. The genotyping assays developed for the FAD2‐1A , FAD2‐1B , and FAD2‐2B isoforms, as well as their linked simple sequence repeat markers, can be used in soybean breeding programs for the elevation of oleic acid seed content through marker‐assisted selection. DA - 2008/// PY - 2008/// DO - 10.2135/cropsci2007.07.0381 VL - 48 IS - 2 SP - 640-650 SN - 1435-0653 ER - TY - JOUR TI - Estimating the individual effects of the reduced palmitic acid fap(nc) and fap1 alleles on agronomic traits in two soybean populations AU - Cardinal, Andrea J. AU - Dewey, Ralph E. AU - Burton, Joseph W. T2 - CROP SCIENCE AB - Major fap alleles that reduce palmitate content in soybean [ Glycine max (L.) Merr.] seed oil also can reduce seed yield. One of these alleles, fap nc , has been shown to be a deletion in the GmFATB1a gene. Allele‐specific primers that amplify GmFATB1a can be used to test precisely if the fap nc allele has an effect on agronomic traits. The objectives of this study were to determine if the segregation of the fap nc allele explained a significant amount of genetic variation in several agronomic traits; to determine if the fap1 allele or minor palmitate genes have an effect on agronomic traits; and to confirm if GmFATB1a maps to the distal region on linkage group A1. GmFATB1a ‐specific primers were used to genotype lines from two populations segregating for fap nc , fap1, and fan alleles and modifier genes. The fap nc allele explained a significant portion of the genetic variation in seed yield, plant height, protein content, and stearic acid content in both populations. After removing the effect of fap nc from the model, the genetic correlation between palmitate and yield was significant in one population but not significant between palmitate and height, indicating that fap1 has a small but significant effect on seed yield but no effect on plant height. The fap1 and/or modifier genes significantly affected stearic acid content. GmFATB1a mapped 20 cM distal to Satt684 on linkage group A1. Breeding efforts did not totally eliminate the negative influence of the fap nc allele on seed yield and plant height. DA - 2008/// PY - 2008/// DO - 10.2135/cropsci2007.05.0251 VL - 48 IS - 2 SP - 633-639 SN - 1435-0653 ER - TY - JOUR TI - Design, synthesis, and photophysical characterization of water-soluble chlorins AU - Borbas, K. Eszter AU - Chandrashaker, Vanampally AU - Muthiah, Chinnasamy AU - Kee, Hooi Ling AU - Holten, Dewey AU - Lindsey, Jonathan S. T2 - JOURNAL OF ORGANIC CHEMISTRY AB - The use of chlorins as photosensitizers or fluorophores in a range of biological applications requires facile provisions for imparting high water solubility. Two free base chlorins have been prepared wherein each chlorin bears a geminal dimethyl group in the reduced ring and a water-solubilizing unit at the chlorin 10-position. In one design (FbC1-PO3H2), the water-solubilizing unit is a 1,5-diphosphonopent-3-yl (“swallowtail”) unit, which has previously been used to good effect with porphyrins. In the other design (FbC2-PO3H2), the water-solubilizing unit is a 2,6-bis(phosphonomethoxy)phenyl unit. Two complementary routes were developed for preparing FbC2-PO3H2 that entail introduction of the protected phosphonate moieties either in the Eastern-half precursor to the chlorin or by derivatization of an intact chlorin. Water-solubilization is achieved in the last step of each synthesis upon removal of the phosphonate protecting groups. The chlorins FbC1-PO3H2 and FbC2-PO3H2 are highly water-soluble (>10 mM) as shown by 1H NMR spectroscopy (D2O) and UV−vis absorption spectroscopy. The photophysical properties of the water-soluble chlorins in phosphate-buffered saline solution (pH 7.4) at room temperature were investigated using static and time-resolved absorption and fluorescence spectroscopic techniques. Each chlorin exhibits dominant absorption bands in the blue and the red region (λ = 398, 626 nm), a modest fluorescence yield (Φf ≈ 0.11), a long singlet excited-state lifetime (τ = 7.5 ns), and a high yield of intersystem crossing to give the triplet state (Φisc = 0.9). The properties of the water-soluble chlorins in aqueous media are comparable to those of hydrophobic chlorins in toluene. The high aqueous solubility combined with the attractive photophysical properties make these compounds suitable for a wide range of biomedical applications. DA - 2008/4/18/ PY - 2008/4/18/ DO - 10.1021/jo7026728 VL - 73 IS - 8 SP - 3145-3158 SN - 1520-6904 ER - TY - JOUR TI - Cloning of feline FOXP3 and detection of expression in CD4+CD25+ regulatory T cells AU - Lankford, Susan AU - Petty, Christopher AU - La Voy, Alora AU - Reckling, Stacie AU - Tompkins, Wayne AU - Dean, Gregg A. T2 - VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY AB - Regulatory T cells (Treg) are increased and directly infected by feline immunodeficiency virus (FIV) and likely play a role in other feline autoimmune, neoplastic, and infectious diseases. Phenotypically, Treg are best characterized by surface expression of CD4 and CD25 and intranuclear expression of the forkhead transcription factor Foxp3. Our objective was to clone and sequence feline FOXP3 for the purpose of developing assays to enhance studies of feline Treg. We determined the feline FOXP3 is 1293 nucleotides in length and codes for a protein that shares high homology to other species. A splice variant devoid of exon 2 was also identified. A real-time PCR assay was developed and used to show Foxp3 mRNA expression occurs primarily in CD4+CD25+ T cells. Two cross-reacting antibodies were identified by immunocytochemical staining of HEK293 cells transfected with feline FOXP3. The antibody labeling confirmed the nuclear localization of the protein. A flow cytometric assay was also validated and used to correlate the phenotypic and functional characteristics of feline Treg induced by treatment of lymph node lymphocytes with flagellin or LPS in combination with mitogen or IL2. Together, these studies provide useful tools to further investigate Foxp3 and Tregs in cats. DA - 2008/3/15/ PY - 2008/3/15/ DO - 10.1016/j.vetimm.2007.11.007 VL - 122 IS - 1-2 SP - 159-166 SN - 1873-2534 KW - Foxp3 KW - feline KW - Treg KW - regulatory T cells KW - toll-like receptors ER - TY - JOUR TI - Detection ofBartonella henselaein the Blood of 2 Adult Horses AU - Jones, S.L. AU - Maggi, R. AU - Shuler, J. AU - Alward, A. AU - Breitschwerdt, E.B. T2 - Journal of Veterinary Internal Medicine AB - Bartonella spp. are emerging zoonotic agents that have been found in a wide variety of domestic animals and wildlife and cause a number of clinical syndromes. Bartonella sp. infection has been identified in a growing number of animal species, including cats, rodents, porpoises, and canids, but has not been reported in horses.To document the presence of Bartonella sp. in the blood of horses.One horse with chronic arthropathy and 1 horse with presumptive vasculitis.Blood samples were tested for the presence of Bartonella sp. by a combination of multiplex real-time polymerase chain reaction and enrichment culture technique.Bartonella henselae was isolated or detected in the blood of both horses.Bartonella henselae infection should be investigated as the cause of disease in horses. DA - 2008/3// PY - 2008/3// DO - 10.1111/j.1939-1676.2008.0043.x VL - 22 IS - 2 SP - 495-498 LA - en OP - SN - 0891-6640 1939-1676 UR - http://dx.doi.org/10.1111/j.1939-1676.2008.0043.x DB - Crossref KW - arthropathy KW - bartonellosis KW - purpura hemorrhagica KW - vasculitis ER - TY - JOUR TI - Protein folding: Are we there yet? AU - Clark, A. C. T2 - Archives of Biochemistry and Biophysics DA - 2008/// PY - 2008/// VL - 469 IS - 1 SP - 1-3 ER - TY - JOUR TI - Minimizing frustration by folding in an aqueous environment AU - Mattos, C. AU - Clark, A. C. T2 - Archives of Biochemistry and Biophysics DA - 2008/// PY - 2008/// VL - 469 IS - 1 SP - 118-131 ER - TY - JOUR TI - Effect of prior tillage and soil fertility amendments on dispersal of Phytophthora capsici and infection of pepper AU - Liu, Bo AU - Gumpertz, Marcia L. AU - Hu, Shuijin AU - Ristaino, Jean Beagle T2 - EUROPEAN JOURNAL OF PLANT PATHOLOGY DA - 2008/3// PY - 2008/3// DO - 10.1007/s10658-007-9216-7 VL - 120 IS - 3 SP - 273-287 SN - 1573-8469 UR - http://www.scopus.com/inward/record.url?eid=2-s2.0-38649084121&partnerID=MN8TOARS KW - Phytophthora capsici KW - epidemiology KW - organic amendment KW - physical KW - chemical and biological parameters KW - disease spread ER - TY - JOUR TI - Olfactomedin-2 mediates development of the anterior central nervous system and head structures in zebrafish AU - Lee, Ju-Ahng AU - Anholt, Robert R. H. AU - Cole, Gregory J. T2 - MECHANISMS OF DEVELOPMENT AB - Olfactomedins comprise a diverse family of secreted glycoproteins, which includes noelin, tiarin, pancortin and gliomedin, implicated in development of the nervous system, and the glaucoma-associated protein myocilin. Here we show in zebrafish that olfactomedin-2 (OM2) is a developmentally regulated gene, and that knockdown of protein expression by morpholino antisense oligonucleotides leads to perturbations of nervous system development. Interference with OM2 expression results in impaired development of branchiomotor neurons, specific disruption of the late phase branchiomotor axon guidance, and affects development of the caudal pharyngeal arches, olfactory pits, eyes and optic tectum. Effects of OM2 knockdown on eye development are likely associated with Pax6 signaling in developing eyes, as Pax6.1 and Pax6.2 mRNA expression patterns are altered in the eyes of OM2 morphants. The specific absence of most cartilaginous structures in the pharyngeal arches indicates that the observed craniofacial phenotypes may be due to perturbed differentiation of cranial neural crest cells. Our studies show that this member of the olfactomedin protein family is an important regulator of development of the anterior nervous system. DA - 2008/// PY - 2008/// DO - 10.1016/j.mod.2007.09.009 VL - 125 IS - 1-2 SP - 167-181 SN - 0925-4773 KW - olfactomedin-2 KW - Pax6.1 KW - zebrafish KW - islet-1 KW - branchiomotor neurons KW - cranial neural crest KW - crestin ER - TY - JOUR TI - Detection of two Bartonella tamiae-like sequences in Amblyomma americanum (Acari : Ixodidae) using 16S-23S intergenic spacer region-specific primers AU - Billeter, Sarah A. AU - Miller, Melissa K. AU - Breitschwerdt, Edward B. AU - Levy, Michael G. T2 - JOURNAL OF MEDICAL ENTOMOLOGY AB - Four hundred and sixty-six questing Amblyomma americanum (L.) (Acari: Ixodidae) from Carolina County, VA, and 98 questing A. americanum from Chatham County, NC, were screened by polymerase chain reaction (PCR) for the Bartonella 16S-23S intergenic spacer region. Two amplicons, approximately 270-280 bp, were detected in two ticks from Virginia. Based upon PCR and sequencing, an adult male and adult female tick harbored DNA sequences closely related to Bartonella tamiae (DQ395180). Bartonella DNA was not detected in A. americanum from North Carolina. Potential transmission of Bartonella spp. by A. americanum should be the focus of future experimental studies. DA - 2008/1// PY - 2008/1// DO - 10.1603/0022-2585(2008)45[176:DOTBTS]2.0.CO;2 VL - 45 IS - 1 SP - 176-179 SN - 0022-2585 KW - Bartonella KW - Amblyomma americanum KW - PCR ER - TY - JOUR TI - A compact water-soluble porphyrin bearing an iodoacetamido bioconjugatable site AU - Borbas, K. Eszter AU - Kee, Hooi Ling AU - Holten, Dewey AU - Lindsey, Jonathan S. T2 - ORGANIC & BIOMOLECULAR CHEMISTRY AB - A broad range of applications requires access to porphyrins that are compact, water-soluble, and bioconjugatable. A symmetrically branched hydrocarbon chain (‘swallowtail’) bearing polar end groups imparts high (>10 mM) aqueous solubility upon incorporation at one of the meso positions of a trans-AB-porphyrin. Two such swallowtail-porphyrins (1a, 1b) equipped with a conjugatable group (carboxylic acid, bromophenyl) have been prepared previously. The synthesis of three new water-soluble trans-AB-porphyrins is reported, where each porphyrin bears a diphosphonate-terminated swallowtail group and an amino (2a), acetamido (2b), or iodoacetamido (2c) group. The amine affords considerable versatility for functionalization. The iodoacetamide provides a sulfhydryl-reactive site for bioconjugation. Porphyrins 2a–2c were fully characterized in aqueous solution by 1H NMR spectroscopy (in D2O), ESI-MS, static absorption spectroscopy, and static and time-resolved fluorescence spectroscopy. Porphyrins 2a–2c exhibit characteristic porphyrin absorption and emission bands in aqueous solution, with a strong, sharp absorption band in the blue region (∼401 nm) and emission in the red region (∼624, 686 nm). Porphyrin 2b in aqueous phosphate buffer or phosphate-buffered saline solution exhibits a fluorescence quantum yield of ∼0.04 and an excited singlet-state lifetime of ∼11 ns. Collectively, the facile synthesis, amenability to bioconjugation, large spacing between the main absorption and fluorescence features, and long singlet excited-state lifetime make this molecular design quite attractive for a range of biomedical applications. DA - 2008/// PY - 2008/// DO - 10.1039/b715072e VL - 6 IS - 1 SP - 187-194 SN - 1477-0539 ER - TY - JOUR TI - Independent origins of self-compatibility in Arabidopsis thaliana AU - Shimizu, Kentaro K. AU - Shimizu-Inatsugi, Rie AU - Tsuchimatsu, Takashi AU - Purugganan, Michael D. T2 - MOLECULAR ECOLOGY AB - Abstract The evolution from outcrossing based on self‐incompatibility (SI) to a selfing system is one of the most prevalent transitions in flowering plants. It has been suggested that the loss of SI in Arabidopsis thaliana is associated with pseudogene formation at the SCR male component of the S locus. Recent work, however, suggests that alternative alleles with large deletions at the S locus are also present and may be responsible for the evolution of self‐compatibility in this species. We demonstrate that most of these deletion alleles are evolutionarily derived from an S haplotype (haplogroups A) that already possessed the SCR pseudogene. This haplotype and its deletion variants are nearly fixed in Europe. Together with previous transgenic data, these results suggest that the pseudogenization of Ψ SCR1 gene changed the SI phenotype in the majority of A. thaliana accessions, and was a critical step in the evolution of selfing in this species. Two other haplogroups (B and C) were also identified, the former of which contains a novel and possibly functional SCR allele. In contrast to haplogroups A, these two haplogroups are found primarily in Africa and Asia. These results suggest that self‐compatibility, which appears to be fixed in this species, arose multiple times with different genetic bases, and indicates that a species‐specific trait is associated with parallel evolution at the molecular level. DA - 2008/1// PY - 2008/1// DO - 10.1111/j.1365-294X.2007.03605.x VL - 17 IS - 2 SP - 704-714 SN - 1365-294X KW - Arabidopsis thaliana KW - evolutionary genomics KW - molecular adaptation KW - reproductive strategies KW - self-incompatibility KW - S locus ER - TY - JOUR TI - Identification of motifs that are conserved in 12 Drosophila species and regulate midline glia vs. neuron expression AU - Estes, Patricia AU - Fulkerson, Eric AU - Zhang, Yi T2 - GENETICS AB - Functional complexity of the central nervous system (CNS) is reflected by the large number and diversity of genes expressed in its many different cell types. Understanding the control of gene expression within cells of the CNS will help reveal how various neurons and glia develop and function. Midline cells of Drosophila differentiate into glial cells and several types of neurons and also serve as a signaling center for surrounding tissues. Here, we examine regulation of the midline gene, wrapper, required for both neuron-glia interactions and viability of midline glia. We identify a region upstream of wrapper required for midline expression that is highly conserved (87%) between 12 Drosophila species. Site-directed mutagenesis identifies four motifs necessary for midline glial expression: (1) a Single-minded/Tango binding site, (2) a motif resembling a pointed binding site, (3) a motif resembling a Sox binding site, and (4) a novel motif. An additional highly conserved 27 bp are required to restrict expression to midline glia and exclude it from midline neurons. These results suggest short, highly conserved genomic sequences flanking Drosophila midline genes are indicative of functional regulatory regions and that small changes within these sequences can alter the expression pattern of a gene. DA - 2008/2// PY - 2008/2// DO - 10.1534/genetics.107.080440 VL - 178 IS - 2 SP - 787-799 SN - 0016-6731 ER - TY - JOUR TI - Evolutionarily conserved cytogenetic changes in hematological malignancies of dogs and humans - man and his best friend share more than companionship AU - Breen, Matthew AU - Modiano, Jaime F. T2 - CHROMOSOME RESEARCH DA - 2008/3// PY - 2008/3// DO - 10.1007/s10577-007-1212-4 VL - 16 IS - 1 SP - 145-154 SN - 1573-6849 KW - chromosome KW - comparative KW - dog KW - evolution KW - leukemia KW - lymphoma ER - TY - JOUR TI - Characterization of a new family of protein kinases from Arabidopsis containing phosphoinositide 3/4-kinase and ubiquitin-like domains AU - Galvao, Rafaelo M. AU - Kota, Uma AU - Soderblom, Erik J. AU - Goshe, Michael B. AU - Boss, Wendy F. T2 - BIOCHEMICAL JOURNAL AB - At least two of the genes predicted to encode type II PI4K (phosphoinositide 4-kinase) in Arabidopsis thaliana (thale cress), namely AtPI4Kγ4 and AtPI4Kγ7, encode enzymes with catalytic properties similar to those of members of the PIKK (phosphoinositide kinase-related kinase) family. AtPI4Kγ4 and AtPI4Kγ7 undergo autophosphorylation and phosphorylate serine/threonine residues of protein substrates, but have no detectable lipid kinase activity. AtPI4Kγ4 and AtPI4Kγ7 are members of a subset of five putative AtPI4Ks that contain N-terminal UBL (ubiquitin-like) domains. In vitro analysis of AtPI4Kγ4 indicates that it interacts directly with, and phosphorylates, two proteins involved in the ubiquitin–proteasome system, namely UFD1 (ubiquitin fusion degradation 1) and RPN10 (regulatory particle non-ATPase 10). On the basis of the present results, we propose that AtPI4Kγ4 and AtPI4Kγ7 should be designated UbDKγ4 and UbDKγ7 (ubiquitin-like domain kinases γ4 and γ7). These UBL-domain-containing AtPI4Ks correspond to a new PIKK subfamily of protein kinases. Furthermore, UFD1 and RPN10 phosphorylation represents an additional mechanism by which their function can be regulated. DA - 2008/1/1/ PY - 2008/1/1/ DO - 10.1042/bj20070959 VL - 409 SP - 117-127 SN - 1470-8728 KW - phosphoinositide kinase (PIK) KW - phosphorylation KW - proteasome regulatory particle non-ATPase 10 subunit (proteasome RPN10 subunit) KW - protein kinase KW - ubiquitin fusion degradation (UFD) KW - ubiquitin-like domain (UBL domain) ER - TY - JOUR TI - Prevalence of Ehrlichia canis, Anaplasma platys, Babesia canis vogeli, Hepatozoon canis, Bartonella vinsonii berkhoffii, and Rickettsia spp. in dogs from Grenada AU - Yabsley, Michael J. AU - McKibben, John AU - Macpherson, Calum N. AU - Cattan, Peggy R. AU - Cherry, Natalie A. AU - Hegarty, Barbara C. AU - Breitschwerdt, Edward B. AU - O'Connor, Tom AU - Chandrashekar, Ramaswamy AU - Paterson, Tara AU - Perea, Marta Lanza AU - Ball, Geoffrey AU - Friesen, Stanley AU - Goedde, Jill AU - Henderson, Brooke AU - Sylvester, Wayne T2 - VETERINARY PARASITOLOGY AB - To identify the tick-borne pathogens in dogs from Grenada, we conducted a serologic survey for Ehrlichia canis in 2004 (104 dogs) and a comprehensive serologic and molecular survey for a variety of tick-borne pathogens in 2006 (73 dogs). In 2004 and 2006, 44 and 32 dogs (42.3% and 43.8%) were seropositive for E. canis, respectively. In 2006, several tick-borne pathogens were identified by serology and PCR. DNA of E. canis, Anaplasma platys, Babesia canis vogeli, Hepatozoon canis, and Bartonella sp. were identified in 18 (24.7%), 14 (19.2%), 5 (7%), 5 (7%), and 1 (1.4%) dogs, respectively. Six (8.2%) dogs were seropositive for Bartonella vinsonii subsp. berkhoffii. All dogs were seronegative and PCR-negative for Rickettsia spp. Coinfection with two or three pathogens was observed in eight dogs. Partial 16S rRNA E. canis and A. platys sequences were identical to sequences in GenBank. Partial 18S rRNA gene sequences from the Grenadian H. canis were identical to each other and had one possible mismatch (ambiguous base) from H. canis detected from Spain and Brazil. Grenadian B. c. vogeli sequences were identical to B. c. vogeli from Brazil and Japan. All of the detected pathogens are transmitted, or suspected to be transmitted, by Rhipicephalus sanguineus. Results of this study indicate that dogs from Grenada are infected with multiple tick-borne pathogens; therefore, tick-borne diseases should be included as differentials for dogs exhibiting thrombocytopenia, leukopenia, fever, or lethargy. One pathogen, E. canis, is also of potential public health significance. DA - 2008/2/14/ PY - 2008/2/14/ DO - 10.1016/j.vetpar.2007.11.008 VL - 151 IS - 2-4 SP - 279-285 SN - 1873-2550 KW - Anaplasma KW - Babesia KW - Bartonella KW - dog KW - Ehrlichia KW - Hepatozoon KW - Rickettsia KW - Rhipicephalus sanguineus ER - TY - JOUR TI - Infusion of dye molecules into Red clover necrotic mosaic virus AU - Loo, LiNa AU - Guenther, Richard H. AU - Lommel, Steven A. AU - Franzen, Stefan T2 - CHEMICAL COMMUNICATIONS AB - The Red clover necrotic mosaic viruscapsid is utilized to package and release molecules through reversible depletion and re-addition of divalent cations. DA - 2008/// PY - 2008/// DO - 10.1039/b714748a IS - 1 SP - 88-90 SN - 1359-7345 ER - TY - JOUR TI - In vivo depletion of CD4(+)CD25(+) regulatory T cells in cats AU - Smithberg, S. Rochelle AU - Fogle, Jonathan E. AU - Mexas, Angela M. AU - Reckling, Stacie K. AU - Lankford, Susan M. AU - Tompkins, Mary B. AU - Dean, Gregg A. T2 - JOURNAL OF IMMUNOLOGICAL METHODS AB - To establish a characterized model of regulatory T cell (Treg) depletion in the cat we assessed the kinetics of depletion and rebound in peripheral and central lymphoid compartments after treatment with anti-CD25 antibody as determined by cell surface markers and FOXP3 mRNA expression. An 82% decrease in circulating CD4+CD25+ Tregs was observed by day 11 after treatment. CD4+CD25+ cells were also reduced in the thymus (69%), secondary lymphoid tissues (66%), and gut (67%). Although CD4+CD25+ cells rebound by day 35 post-treatment, FOXP3 levels remain depressed suggesting anti-CD25 antibody treatment has a sustainable diminutive effect on the Treg population. To determine whether CD25+ Treg depletion strategies also deplete activated CD25+ effector cells, cats were immunized with feline immunodeficiency virus (FIV) p24-GST recombinant protein, allowing them to develop a measurable memory response, prior to depletion with anti-CD25 antibody. Anti-FIV p24-GST effector cell activity in peripheral blood after depletion was sustained as determined by antigen-specific T cell proliferation and humoral responses against FIV p24-GST with an ELISA for antigen-specific feline IgG. Furthermore, development of an anti-mouse response in Treg-depleted cats was similar to control levels indicating the retained capacity to respond to a novel antigen. We conclude that despite alterations in CD25+ cell levels during depletion, the feline immune system remains functional. We demonstrate here a model for the study of disease pathogenesis in the context of reduced numbers of immunosuppressive CD4+CD25+ Tregs throughout the feline immune system. DA - 2008/1/1/ PY - 2008/1/1/ DO - 10.1016/j.jim.2007.09.015 VL - 329 IS - 1-2 SP - 81-91 SN - 0022-1759 KW - regulatory T cell KW - FOXP3 KW - feline immunodeficiency virus KW - monoclonal antibody ER - TY - JOUR TI - Expression of the bacteriophage T4 lysozyme gene in tall fescue confers resistance to gray leaf spot and brown patch diseases AU - Dong, Shujie AU - Shew, H. David AU - Tredway, Lane P. AU - Lu, Jianli AU - Sivamani, Elumalai AU - Miller, Eric S. AU - Qu, Rongda T2 - TRANSGENIC RESEARCH DA - 2008/2// PY - 2008/2// DO - 10.1007/s11248-007-9073-3 VL - 17 IS - 1 SP - 47-57 SN - 1573-9368 KW - fungal resistance KW - Magnaporthe grisea KW - Rhizoctonia solani KW - T4 lysozyme KW - tall fescue ER - TY - JOUR TI - Concurrent Hepatic Copper Toxicosis and Fanconi's Syndrome in a Dog AU - Hill, T.L. AU - Breitschwerdt, E.B. AU - Cecere, T. AU - Vaden, S. T2 - Journal of Veterinary Internal Medicine AB - A3-year-old male castrated West Highland White Terrier was presented to the Veterinary Teaching Hospital at North Carolina State University with a 1-week history of intermittent vomiting, polyuria and polydypsia, progressive anorexia, and lethargy. No treatment was administered by the primary veterinarian before referral. The dog was adopted as a puppy and had lived exclusively in North Carolina. The dog received no other medications and had no previous illnesses, except for a transient decrease in appetite 1 year earlier. On physical examination, the dog weighed 8.2 kg, had a body condition score of 6/9, but was approximately 5% dehydrated. Hematologic abnormalities included neutrophilia (14,293 cells/μL, reference range 2,529–12,876 cells/μL) with a left shift (642 band neutrophils/μL) and immature granulocytes (161/μL). Biochemical abnormalities included increased ALT activity (456 U/L; reference range, 16–73 U/L), hyperbilirubinemia (0.3 mg/dL; reference range, 0–0.2 mg/dL), hypokalemia (3.8 mEq/L; reference range, 3.9–5.2 mEq/L), hyperchloremia (121 mEq/L; reference range, 104–117 mEq/L), and increased lipase activity (715 U/L; reference range, 22–216 U/L). Urinalysis findings included a specific gravity of 1.022, pH of 8, 2+ proteinuria, 2+ ketones, 2+ blood, 4+ glucose, 0–2 coarse granular casts/hpf, 0–5 white cells/hpf, and 5–10 red cells/hpf. The urine protein : creatinine ratio was 1.7. A urine sample submitted for bacterial culture yielded no growth. There was mild metabolic acidemia (blood pH 7.26; reference range, 7.36–7.47), bicarbonate 16.2 mEq/L (reference range, 19.8–26.2 mEq/L), and PCO2 36 mmHg (reference range, 30–40 mmHg), on a venous blood sample. Abdominal ultrasonographic findings included periportal lymph node enlargement, microhepatica, and mildly hyperechoic kidneys bilaterally. Fasting and postprandial serum bile acid concentrations were within reference range. The dog was treated with an IV infusion of lactated Ringer's solution containing 30 mEq/L KCl for dehydration, hypokalemia, and metabolic acidosis. Within 48 hours of hospitalization, sodium bicarbonate at 3 mEq/kg/d was administered as a continuous rate infusion to correct the persistent metabolic acidosis. In addition, the dog was treated with a metoclopramide constant rate infusion of 1 mg/kg/d and famotidine 0.5 mg/kg IV q12h. During the 1st 48 hours of hospitalization, the dog remained anorexic and frequently vomited bile; dolasetron 0.6 mg/kg IV q24h and sucralfate 500 mg PO q8h were administered. The dog was supplemented nutritionally by nasoesophageal feeding a liquid diet (Perativea), the continuous rate of which was adjusted to minimize vomiting. Serial assessment of urine dipstick tests and serum glucose measurements indicated that the dog was persistently ketonuric and glucosuric, with normal serum glucose concentrations. Despite a metabolic acidosis, urine pH ranged from 7.0 to 8.0. The findings of proteinuria, glucosuria with normoglycemia, and hyperchloremic metabolic acidosis with alkaline urine supported a diagnosis of Fanconi's syndrome. Results of a urine metabolic profile, performed at the University of Pennsylvania, also were consistent with Fanconi's syndrome with severe generalized amino aciduria and marked glucosuria. By the 4th day of hospitalization, the dog became febrile (103.6°F). An abdominal ultrasound examination indicated mild thickening of the gallbladder wall. Antibiotic therapy was initiated with ampicillin and sulbactam (Unasyn,b 30 mg/kg IV q8h); the fever resolved within 36 hours. Because of the dog's refractory vomiting and unknown underlying hepatic pathology, an exploratory laparotomy was performed on the 5th day of hospitalization. Biopsy specimens were taken of the liver, stomach, duodenum, jejunum, and left kidney. The gallbladder was aspirated and 1 aliquot of bile was submitted for cytologic analysis; another aliquot was submitted for aerobic microbial culture. Gastrostomy and jejunostomy tubes were placed. Recovery from surgery was uneventful. Perativea liquid diet was administered continuously through the jejunostomy tube throughout the remainder of hospitalization without incident. The volume administered was gradually increased to basal energy requirements. Ketonuria resolved within 24 hours after feeding basal energy requirements. Dexamethasone (0.08 mg/kg IV q24h), s-adenosylmethionine (22 mg/kg PO q24h), and ursodeoxycholic acid (15 mg/kg PO q24h) were administered on day 5 postoperatively. Within 24 hours, steady improvement was observed; the vomiting ceased, the dog was more active and alert, and began eating. IV bicarbonate supplementation was necessary to maintain a near normal serum pH. The bile fluid was cytologically unremarkable and bacterial growth was not observed. Lymphofollicular gastritis, which was attributed to refractory vomiting, and mild lymphocytic enteritis were observed histologically on full-thickness stomach and intestinal biopsy specimens. Centrilobular pyogranulomatous hepatitis, characterized by infiltrates of neutrophils, macrophages, and fewer lymphocytes and plasma cells associated with single cell hepatocyte necrosis, was present in liver wedge biopsy specimens. Abundant copper accumulation was present within centrilobular hepatocytes and macrophages (Fig 1). Copper was quantified at 1186 ppm dry weight. Histopathologic abnormalities in the kidney included diffuse mild to moderate tubular atrophy, multifocal tubular epithelial vacuolation, and tubular regeneration. Additionally, there was evidence of abnormal copper accumulation within vacuolated renal tubular epithelium (Fig 2). Liver, dog. (A) Centrilobular zone with individual hepatocellular necrosis (arrow) and infiltrates of macrophages, neutrophils, lymphocytes, and plasma cells. H&E stain. Scale bar = 50 μm. (B) Same section as A with numerous well-demarcated, red–brown copper-containing granules present in the cytoplasm of macrophages and hepatocytes. Rhodanine stain. Scale bar = 50 μm. Kidney, dog. (A) Tubules lined by plump vacuolated epithelial cells (arrow) or mildly atrophied epithelium (arrowhead). H&E stain. Scale bar = 20 μm. (B) Renal tubular epithelial cells containing multiple cytoplasmic well-demarcated, red–brown copper-containing granules. Rhodanine stain. Scale bar = 20 μm. The dog gradually was transitioned to oral medications administered through the gastrotomy tube, including 971 mg of bicarbonate q12h, s-adenosylmethionine, ursodeoxycholic acid, prednisone 0.6 mg/kg q12h that was tapered over the next 4 weeks, famotidine 0.6 mg/kg q12h for 2 weeks, amoxicillin–clavulanic acid 12 mg/kg q12h for 2 weeks, ciprofloxacin 8 mg/kg q12h for 2 weeks, and Marinc 1 medium dog tablet daily for 2 months. d-Penicillamine was given at a dosage of 10 mg/kg PO q12h for 3 months, with no reported adverse effects. After 2 weeks of treatment, the dog was no longer proteinuric, glucosuric, or ketonuric. Venous pH was 7.36 and bicarbonate was 26.6 mEq/L. ALT activity had decreased from 456 to 81 U/L. Ursodeoxycholic acid, s-adenosylmethionine, Marin, bicarbonate, and d-penicillamine therapy were continued. Monthly venous blood gas analysis disclosed normal pH while receiving oral bicarbonate supplementation. At the 3-month recheck examination, the bicarbonate dosage was reduced by half and d-penicillamine was discontinued. Repeat biopsies of the liver and kidney were declined by the owner. Zinc acetate was prescribed (10 mg/kg orally q12h); s-adenosylmethionine, ursodeoxycholic acid, and Marin were continued. Three weeks after decreasing the oral bicarbonate supplementation, venous pH was 7.4 and bicarbonate was 27.9 mEq/L and bicarbonate supplementation was discontinued, but zinc acetate supplementation was continued indefinitely. Thirteen months after initial diagnosis and treatment, the dog was doing well with no clinical signs reported. Copper storage diseases (CSDs) have been described in several breeds, including the Bedlington Terrier, West Highland White Terrier, Skye Terrier, Doberman Pinscher, Labrador Retriever, and Dalmatian, as well as other species, including humans, rats, and sheep. With CSD, copper accumulates in the liver, leading to hepatitis and eventually cirrhosis of the hepatic parenchyma.1 As yet, the genetic basis of CSD has been elucidated only in the Bedlington Terrier, where it is related to a defect in the MURR-1 gene.2 CSD in the Bedlington Terrier results in copper accumulation in the liver as well as renal cortical tissue in some patients.3 We describe a West Highland White Terrier with CSDand concurrent Fanconi's syndrome, which resolved after copper chelation therapy. A genetic mutation that causes abnormal copper accumulation has yet to be identified in West Highland White Terrier. Unlike the Bedlington Terrier, there does not seem to be a correlation between age and hepatic copper concentration in the West Highland White Terrier. In addition, the total hepatic copper concentration is lower in the majority of affected West Highland White Terriers; most West Highland White Terriers have copper concentrations <1,500 ppm dry weight.4, 5 Copper concentrations up to 500 ppm dry weight may occur in normal dogs.6 Thornburg describes the histopathologic lesions of copper toxicosis in West Highland White Terriers as being characterized by multifocal centrilobular hepatitis and cirrhosis. In this dog, the findings were consistent with previous reports, including multifocal centrilobular hepatitis and a copper concentration of 1,186 ppm. It is unclear whether copper toxicosis was the direct cause of Fanconi's syndrome in this dog, because several medical therapies, including antibiotics, steroids, and penicillamine, may have contributed to resolution of Fanconi's syndrome regardless of treating copper toxicosis. The presence of copper in the renal tubules and resolution of Fanconi's syndrome after copper chelation therapy suggest that copper toxicosis may be a cause of Fanconi's syndrome in dogs. In other reports, dogs with copper toxicosis had abnormalities indicative of proximal tubular dysfunction.7, 8 In a study of 10 Dalmatians with copper toxicosis, 3 dogs had proteinuria without pyuria, 2 had glucosuria with normoglycemia, and 1 had renal tubular necrosis with granular casts.7 In a separate report, a 1.5-year-old Dalmatian had copper toxicosis with a positive metabolic screen for Fanconi's syndrome.8 This dog was euthanized because the dog continued to decline clinically. In the dog reported here, glucosuria, metabolic acidosis, amino aciduria, and copper accumulation in the renal tubules were documented with resolution of Fanconi's syndrome after copper chelation therapy. Wilson's disease is a CSD in humans in which copper accumulates to toxic concentrations in the liver and secondarily in the central nervous system and kidneys because of a mutation in the ATP7B gene, which normally allows for copper excretion into the bile and for production of ceruloplasmin. As a result, patients with Wilson's disease have copper accumulation in the liver and in other tissues, including the brain, kidney, red blood cells, and eye. Neurologic signs develop including speech disorders and dysphagia, abnormal and uncoordinated gait, and tremors.9 Renal complications, including urolithiasis and Fanconi's syndrome, have been reported in human patients with Wilson's disease.10, 11 As is reported in association with Wilson's disease in human patients, this dog may represent a subset of patients with copper storage disease that have concurrent renal tubular dysfunction in association with copper accumulation in the proximal tubular epithelium. Several types of proximal renal tubular dysfunction have been described in association with Wilson's disease in humans: failure of renal acidification, amino aciduria, glucosuria, and phosphaturia.11-15 Resolution of proximal tubular dysfunction also may occur after treatment for copper toxicosis with penicillamine.11, 13-15 A renal biopsy in 1 patient, with prior documentation of renal tubular copper accumulation, demonstrated normal proximal tubular ultrastructure 2 years after initiation of penicillamine therapy. Penicillamine therapy was discontinued because of adverse effects, including glomerulonephritis and systemic lupus erythematosus. Eighteen months later, the patient again showed signs of Wilson's disease, and a renal biopsy was repeated. Electron-dense bodies, consistent with copper-bound metalloprotein, were evident in the subapical cytoplasm of the tubular cells, although copper quantification was not performed to confirm increased copper concentrations in the kidney.11 The effect of copper loading has been examined in the rat kidney as a model for copper toxicosis in humans and other species. Rats supplemented with excessive copper, either by injection or by dietary supplementation, develop copper staining in the liver and in the proximal convoluted tubular epithelium.16-18 Haywood demonstrated an increase in copper content of the kidney in rats fed excess copper as well as copper staining granules confined to the proximal tubule. There also were degenerative changes of the tubular cells as well as copper-staining debris in the tubular lumen, suggesting active exocytosis of copper-bound metallothionein.17 A later report of copper loading in the rat kidney described increased copper in renal tubular lysosomes. With time and increased copper concentrations, there was progressive nuclear degeneration in proximal tubular cells and disruption of the mitochondrial membrane.18 These findings are consistent with the observation that copper acts as a prooxidant, disrupting cell membranes and damaging DNA. Eventually, the rats extruded copper-stained lysosomes and copper-laden pinocytotic vesicles into the tubular lumen. After this time point, the copper concentration in the kidney began to decrease and the tubular epithelium recovered to nearly normal. The localization of copper to the renal tubular epithelium, later exocytosis of copper-bound organelles, and recovery of the tubular epithelium suggest a mechanism by which the rat seems able to cope with increased dietary copper intake. Copper also may alter the function of Na-K-ATPase in the proximal tubular epithelium. In vitro, for both rat kidney tissue homogenate and rat synaptic plasma membrane, copper has an inhibitory effect on the function of this important enzyme in a concentration-dependent manner.19, 20 As copper stores accumulate in the canine liver, the kidney may attempt exocytosis of excess copper as occurs in the copper-loaded rat. Exocytosis of copper-bound organelles in the canine and human kidney may be less effective than in the rat, because tubular debris is not described in any published reports of copper-stained kidneys from rats. Because copper accumulates in the kidney as well as the liver, it then may have several effects that ultimately lead to proximal tubular dysfunction and Fanconi's syndrome: necrosis and apoptosis of epithelial cells because copper acts as a pro-oxidant, disrupting mitochondrial membranes and DNA, inducing inflammation that may affect epithelial function, and inhibiting the function of Na-K-ATPase in a concentration-dependent manner that would alter transport mechanisms in the proximal tubule. These effects all may lead to decreased reabsorption of glucose, amino acids, phosphate, and bicarbonate from the tubular lumen. These may be the mechanisms by which copper toxicosis in this West Highland White Terrier resulted in proximal tubular dysfunction characterized as Fanconi's syndrome. Few controlled studies are available in human and canine medicine that describe renal pathology with copper toxicosis or the effect of copper chelation therapy. Additional study is needed to definitively confirm a link between Fanconi's syndrome and copper storage disease in dogs as suggested by this dog, and to elucidate the nature of that association. In dogs with suspected copper storage disease and evidence of tubular dysfunction that are undergoing liver biopsy, it may be warranted to perform a renal biopsy to allow for histopathologic examination of the renal parenchyma, as well as renal copper quantification, which was not performed here. Sequential urine metabolic screening or urinalyses with protein quantification may be a useful diagnostic and therapeutic monitoring tool in those patients with evidence of tubular dysfunction. Additionally, liver and kidney biopsies after 3 months of d-penicillamine therapy may have been informative to document the histological response to treatment. The authors gratefully acknowledge the assistance of the veterinarians involved in the treatment and referral of the dog. Our appreciation is extended to Dr Herman Jeffers, Dr Karyn Harrell, and Dr Sally Bissett. aPerative Specialized Nutrition, Abbot Laboratories, Ross Products Division, Columbus, OH bUnasyn, Pfizer Roerig, Pfizer Inc, New York cMarin, Nutramax Laboratories Inc, Edgewood, MD DA - 2008/1// PY - 2008/1// DO - 10.1111/j.1939-1676.2007.0040.x VL - 22 IS - 1 SP - 219-222 LA - en OP - SN - 0891-6640 1939-1676 UR - http://dx.doi.org/10.1111/j.1939-1676.2007.0040.x DB - Crossref ER - TY - JOUR TI - CYP82E4-mediated nicotine to nornicotine conversion in tobacco is regulated by a senescence-specific signaling pathway AU - Chakrabarti, Manohar AU - Bowen, Steven W. AU - Coleman, Nicholas P. AU - Meekins, Karen M. AU - Dewey, Ralph E. AU - Siminszky, Balazs T2 - PLANT MOLECULAR BIOLOGY DA - 2008/3// PY - 2008/3// DO - 10.1007/s11103-007-9280-6 VL - 66 IS - 4 SP - 415-427 SN - 1573-5028 KW - alkaloid KW - cytochrome P450 KW - SAG KW - senescence-associated gene KW - stress KW - transcriptional regulation ER - TY - JOUR TI - Bartonella DNA in the Blood and Lymph Nodes of Golden Retrievers with Lymphoma and in Healthy Controls AU - Duncan, A.W. AU - Marr, H.S. AU - Birkenheuer, A.J. AU - Maggi, R.G. AU - Williams, L.E. AU - Correa, M.T. AU - Breitschwerdt, E.B. T2 - Journal of Veterinary Internal Medicine AB - Background: Although lymphoma is the most common neoplastic process reported in dogs, its precise etiology is unknown. Golden Retrievers are more likely to develop lymphoma, suggesting a breed predisposition; however, other factors, including environment, immunity, and infection, are likely contributors to oncogenesis. Hypothesis: We hypothesized that the development of lymphoma in Golden Retrievers may be associated with vector‐borne infections, specifically Bartonella , Anaplasma , or Ehrlichia species infections. Animals: Golden Retrievers with lymphoma and healthy Golden Retrievers from across the United States were recruited for study participation. Methods: A matched, case‐control study was performed to determine the association of lymphoma and the presence of Bartonella, Anaplasma , and Ehrlichia species in serum, blood, and lymph node aspirates. Results: Using PCR analyses and DNA sequencing, single and coinfections with Bartonella henselae , Bartonella elizabethae, Bartonella quintana , and/or Bartonella vinsonii ( berkhoffii ) were detected in the blood and lymph node aspirates of Golden Retrievers with lymphoma (5/28 dogs, 18%) and in healthy Golden Retrievers (10/56 dogs, 18%); no Anaplasma or Ehrlichia DNA was detected in any dog. When compared with dogs with lymphoma, a higher ( P <.001) proportion of healthy Golden Retrievers were receiving monthly acaricide treatments (2.6 times higher). Conclusions and Clinical Importance: Bartonella DNA can be detected in blood and lymph nodes; importantly, in this report, Bartonella was detected in the same proportion of clinically healthy dogs and dogs with lymphoma. Longitudinal studies should be conducted to determine the mode of transmission of Bartonella in dogs, whether lymphatic infection is persistent, or whether these bacteria may contribute to the development of lymphoma. DA - 2008/1// PY - 2008/1// DO - 10.1111/j.1939-1676.2007.0018.x VL - 22 IS - 1 SP - 89–95 SN - 0891-6640 1939-1676 UR - http://dx.doi.org/10.1111/j.1939-1676.2007.0018.x KW - bacteria KW - cancer KW - dog KW - tick KW - vector-borne ER -