@article{yell_li_2023, title={Scalable, Flexible, and Cost-Effective Seedling Grafting}, volume={1}, ISSN={["1940-087X"]}, url={http://dx.doi.org/10.3791/64519}, DOI={10.3791/64519}, abstractNote={Early-stage seedling grafting has become a popular tool in molecular genetics to study root-shoot relationships within plants. Grafting early-stage seedlings of the small model plant, Arabidopsis thaliana, is technically challenging and time consuming due to the size and fragility of its seedlings. A growing collection of published methods describe this technique with varying success rates, difficulty, and associated costs. This paper describes a simple procedure to make an in-house reusable grafting device using silicone elastomer mix, and how to use this device for seedling grafting. At the time of this publication, each reusable grafting device costs only $0.47 in consumable materials to produce. Using this method, beginners can have their first successfully grafted seedlings in less than 3 weeks from start to finish. This highly accessible procedure will allow plant molecular genetics labs to establish seedling grafting as a normal part of their experimental process. Due to the full control users have in the creation and design of these grafting devices, this technique could be easily adjusted for use in larger plants, such as tomato or tobacco, if desired.}, number={191}, journal={JOVE-JOURNAL OF VISUALIZED EXPERIMENTS}, author={Yell, Victoria and Li, Xu}, year={2023}, month={Jan} }
 @article{li_yell_li_2023, title={Two Arabidopsis promoters drive seed-coat specific gene expression in pennycress and camelina}, volume={19}, ISSN={["1746-4811"]}, url={https://doi.org/10.1186/s13007-023-01114-x}, DOI={10.1186/s13007-023-01114-x}, abstractNote={Abstract Background Pennycress and camelina are two important novel biofuel oilseed crop species. Their seeds contain high content of oil that can be easily converted into biodiesel or jet fuel, while the left-over materials are usually made into press cake meals for feeding livestock. Therefore, the ability to manipulate the seed coat encapsulating the oil- and protein-rich embryos is critical for improving seed oil production and press cake quality. Results Here, we tested the promoter activity of two Arabidopsis seed coat genes, AtTT10 and AtDP1 , in pennycress and camelina by using eGFP and GUS reporters. Overall, both promoters show high levels of activities in the seed coat in these two biofuel crops, with very low or no expression in other tissues. Importantly, AtTT10 promoter activity in camelina shows differences from that in Arabidopsis, which highlights that the behavior of an exogenous promoter in closely related species cannot be assumed the same and still requires experimental determination. Conclusion Our work demonstrates that AtTT10 and AtDP1 promoters are suitable for driving gene expression in the outer integument of the seed coat in pennycress and camelina.}, number={1}, journal={PLANT METHODS}, author={Li, Xin and Yell, Victoria and Li, Xu}, year={2023}, month={Dec} }
 @article{chen_li_chapple_dilkes_li_2023, title={UGT76F1 glycosylates an isomer of the C7-necic acid component of pyrrolizidine alkaloids in Arabidopsis thaliana}, volume={4}, ISSN={["1365-313X"]}, url={https://doi.org/10.1111/tpj.16211}, DOI={10.1111/tpj.16211}, abstractNote={Identification of unknown metabolites and their biosynthetic genes is an active research area in plant specialized metabolism. By following a gene-metabolite association from a genome-wide association study of Arabidopsis stem metabolites, we discovered a previously unknown metabolite, 2-hydroxy-2-(1-hydroxyethyl)pentanoic acid glucoside, and demonstrated that UGT76F1 is responsible for its production in Arabidopsis. The chemical structure of the glucoside was determined by a series of analyses including tandem mass spectrometry, acid and base hydrolysis, and NMR spectrometry. T-DNA knockout mutants of UGT76F1 are devoid of the glucoside but accumulate increased levels of the aglycone. 2-hydroxy-2-(1-hydroxyethyl)pentanoic acid is structurally related to the C7-necic acid component of lycopsamine-type pyrrolizidine alkaloids such as trachelantic acid and viridifloric acid. Feeding norvaline greatly enhances the accumulation of 2-hydroxy-2-(1-hydroxyethyl)pentanoic acid glucoside in wild type but not the UGT76F1 knockout mutant plants, providing evidence for an orthologous C7-necic acid biosynthetic pathway in Arabidopsis despite the apparent lack of pyrrolizidine alkaloids.}, journal={PLANT JOURNAL}, author={Chen, Han-Yi and Li, Xin and Chapple, Clint and Dilkes, Brian and Li, Xu}, year={2023}, month={Apr} }
 @article{lu_li_young_li_linder_suchoff_2022, title={Hyperspectral imaging with chemometrics for non-destructive determination of cannabinoids in floral and leaf materials of industrial hemp (Cannabis sativa L.)}, volume={202}, ISSN={["1872-7107"]}, DOI={10.1016/j.compag.2022.107387}, abstractNote={With the passage of the 2018 Farm Bill, industrial hemp (Cannabis sativa L.) has become a legal and economically promising crop commodity for U.S. farmers. There has been a surge of interest in growing industrial hemp for producing cannabinoids, such as cannabidiol (CBD), because of their medical potential. Quantitative determination of cannabinoids in harvested materials (primarily floral tissues) is critical for cannabinoid production and compliance testing. The concentrations of cannabinoids in hemp materials are conventionally determined using wet-chemistry chromatographic methods, which require destructive sampling, and are time-consuming, costly, and thus not suitable for on-site rapid testing. This study presents a novel effort to utilize hyperspectral imaging technology for non-destructive quantification of major cannabinoids, including CBD, THC (tetrahydrocannabinol), CBG (cannabigerol) and their acid forms in fresh floral and leaf materials of industrial hemp on a dry weight basis. Hyperspectral images in the wavelength range of 400–1000 nm were acquired from floral and leaf tissues immediately after harvest from a total of 100 industrial hemp plants of five cultivars at varied growth stages. Linear discriminant analysis showed hyperspectral imaging could identify CBD-rich/poor and THC-legal/illegal flower samples with accuracies of 99% and 97%, respectively. Quantitative models based on full-spectrum PLS (partial least squares) achieved prediction accuracies of RPD (ratio of prediction to deviation) = 2.5 (corresponding R2 = 0.84) for CBD and THC in floral tissues. Similar accuracies were obtained for their acid forms in flower samples. The predictions for CBG and its acid form in floral tissues and all six cannabinoids in leaf tissues were unsatisfactory with noticeably lower RPD values. Consistently improved accuracies were obtained by parsimonious PLS models based on a wavelength selection procedure for minimized variable collinearity. The best RPD values of approximately 2.6 (corresponding R2 = 0.85) were obtained for CBD and THC in floral materials. This study demonstrates the utility of hyperspectral imaging as a potential valuable tool for rapid quantification of cannabinoids in industrial hemp.}, journal={COMPUTERS AND ELECTRONICS IN AGRICULTURE}, author={Lu, Yuzhen and Li, Xu and Young, Sierra and Li, Xin and Linder, Eric and Suchoff, David}, year={2022}, month={Nov} }
 @article{linder_young_li_inoa_suchoff_2022, title={The Effect of Harvest Date on Temporal Cannabinoid and Biomass Production in the Floral Hemp (Cannabis sativa L.) Cultivars BaOx and Cherry Wine}, volume={8}, ISSN={["2311-7524"]}, url={https://doi.org/10.3390/horticulturae8100959}, DOI={10.3390/horticulturae8100959}, abstractNote={The objectives of this study were to model the temporal accumulation of cannabidiol (CBD) and tetrahydrocannabinol (THC) in field-grown floral hemp in North Carolina and establish harvest timing recommendations to minimize non-compliant crop production. Field trials were conducted in 2020 and 2021 with BaOx and Cherry Wine cultivars. Harvest events started two weeks after floral initiation and occurred every two weeks for 12 weeks. Per-plant threshed biomass accumulation exhibited a linear plateau trend. The best fit model for temporal accumulation of THC was a beta growth curve. As harvest date was delayed, total THC concentrations increased until concentrations reached their maximum, then decreased as plants approached senescence. Logistic regression was the best fit model for temporal accumulation of CBD. CBD concentrations increased with later harvest dates. Unlike THC concentrations, there was no decline in total CBD concentrations. To minimize risk, growers should test their crop as early as possible within the USDA’s 30-day compliance window. We observed ‘BaOx’ and ‘Cherry Wine’ exceeding the compliance threshold 50 and 41 days after flower initiation, respectively.}, number={10}, journal={HORTICULTURAE}, author={Linder, Eric R. and Young, Sierra and Li, Xu and Inoa, Shannon Henriquez and Suchoff, David H.}, year={2022}, month={Oct} }
 @article{linder_young_li_inoa_suchoff_2022, title={The Effect of Transplant Date and Plant Spacing on Biomass Production for Floral Hemp (Cannabis sativa L.)}, volume={12}, ISSN={["2073-4395"]}, url={https://doi.org/10.3390/agronomy12081856}, DOI={10.3390/agronomy12081856}, abstractNote={Floral hemp cultivated for the extraction of cannabinoids is a new crop in the United States, and agronomic recommendations are scarce. The objective of this study was to understand the effects of plant spacing and transplant date on floral hemp growth and biomass production. Field trials were conducted in North Carolina in 2020 and 2021 with the floral hemp cultivar BaOx. Transplant date treatments occurred every two weeks from 11 May to 7 July (±1 d). Plant spacing treatments were 0.91, 1.22, 1.52, and 1.83 m between plants. Weekly height and width data were collected throughout the vegetative period, and dry biomass was measured at harvest. Plant width was affected by transplant date and spacing. Plant height was affected by transplant date. Earlier transplant dates resulted in taller, wider plants, while larger plant spacing resulted in wider plants. Individual plant biomass increased with earlier transplant dates and larger plant spacing. On a per-hectare basis, biomass increased with earlier transplant dates and smaller transplant spacing. An economic analysis found that returns were highest with 1.22 m spacing and decreased linearly by a rate of −163.098 USD ha−1 d−1. These findings highlight the importance of earlier transplant timing to maximize harvestable biomass.}, number={8}, journal={AGRONOMY-BASEL}, author={Linder, Eric R. and Young, Sierra and Li, Xu and Inoa, Shannon Henriquez and Suchoff, David H.}, year={2022}, month={Aug} }
 @article{tan_wang_schneider_li_souza_tang_grimm_hsieh_wang_li_et al._2021, title={Comparative Phylogenomic Analysis Reveals Evolutionary Genomic Changes and Novel Toxin Families in Endophytic Liberibacter Pathogens}, volume={9}, ISSN={["2165-0497"]}, DOI={10.1128/Spectrum.00509-21}, abstractNote={Liberibacter pathogens are associated with several severe crop diseases, including citrus Huanglongbing, the most destructive disease to the citrus industry. Currently, no effective cure or treatments are available, and no resistant citrus variety has been found. ABSTRACT Liberibacter pathogens are the causative agents of several severe crop diseases worldwide, including citrus Huanglongbing and potato zebra chip. These bacteria are endophytic and nonculturable, which makes experimental approaches challenging and highlights the need for bioinformatic analysis in advancing our understanding about Liberibacter pathogenesis. Here, we performed an in-depth comparative phylogenomic analysis of the Liberibacter pathogens and their free-living, nonpathogenic, ancestral species, aiming to identify major genomic changes and determinants associated with their evolutionary transitions in living habitats and pathogenicity. Using gene neighborhood analysis and phylogenetic classification, we systematically uncovered, annotated, and classified all prophage loci into four types, including one previously unrecognized group. We showed that these prophages originated through independent gene transfers at different evolutionary stages of Liberibacter and only the SC-type prophage was associated with the emergence of the pathogens. Using ortholog clustering, we vigorously identified two additional sets of genomic genes, which were either lost or gained in the ancestor of the pathogens. Consistent with the habitat change, the lost genes were enriched for biosynthesis of cellular building blocks. Importantly, among the gained genes, we uncovered several previously unrecognized toxins, including new toxins homologous to the EspG/VirA effectors, a YdjM phospholipase toxin, and a secreted endonuclease/exonuclease/phosphatase (EEP) protein. Our results substantially extend the knowledge of the evolutionary events and potential determinants leading to the emergence of endophytic, pathogenic Liberibacter species, which will facilitate the design of functional experiments and the development of new methods for detection and blockage of these pathogens. IMPORTANCE Liberibacter pathogens are associated with several severe crop diseases, including citrus Huanglongbing, the most destructive disease to the citrus industry. Currently, no effective cure or treatments are available, and no resistant citrus variety has been found. The fact that these obligate endophytic pathogens are not culturable has made it extremely challenging to experimentally uncover the genes/proteins important to Liberibacter pathogenesis. Further, earlier bioinformatics studies failed to identify key genomic determinants, such as toxins and effector proteins, that underlie the pathogenicity of the bacteria. In this study, an in-depth comparative genomic analysis of Liberibacter pathogens along with their ancestral nonpathogenic species identified the prophage loci and several novel toxins that are evolutionarily associated with the emergence of the pathogens. These results shed new light on the disease mechanism of Liberibacter pathogens and will facilitate the development of new detection and blockage methods targeting the toxins.}, number={2}, journal={MICROBIOLOGY SPECTRUM}, author={Tan, Yongjun and Wang, Cindy and Schneider, Theresa and Li, Huan and Souza, Robson Francisco and Tang, Xueming and Grimm, Kylie D. Swisher and Hsieh, Tzung-Fu and Wang, Xu and Li, Xu and et al.}, year={2021}, month={Oct} }
 @article{simpson_wunderlich_li_svedin_dilkes_chapple_2021, title={Metabolic source isotopic pair labeling and genome-wide association are complementary tools for the identification of metabolite-gene associations in plants}, volume={33}, ISSN={["1532-298X"]}, url={https://doi.org/10.1093/plcell/koaa046}, DOI={10.1093/plcell/koaa046}, abstractNote={The optimal extraction of information from untargeted metabolomics analyses is a continuing challenge. Here, we describe an approach that combines stable isotope labeling, liquid chromatography- mass spectrometry (LC-MS), and a computational pipeline to automatically identify metabolites produced from a selected metabolic precursor. We identified the subset of the soluble metabolome generated from phenylalanine (Phe) in Arabidopsis thaliana, which we refer to as the Phe-derived metabolome (FDM) In addition to identifying Phe-derived metabolites present in a single wild-type reference accession, the FDM was established in nine enzymatic and regulatory mutants in the phenylpropanoid pathway. To identify genes associated with variation in Phe-derived metabolites in Arabidopsis, MS features collected by untargeted metabolite profiling of an Arabidopsis diversity panel were retrospectively annotated to the FDM and natural genetic variants responsible for differences in accumulation of FDM features were identified by genome-wide association. Large differences in Phe-derived metabolite accumulation and presence/absence variation of abundant metabolites were observed in the nine mutants as well as between accessions from the diversity panel. Many Phe-derived metabolites that accumulated in mutants also accumulated in non-Col-0 accessions and was associated to genes with known or suspected functions in the phenylpropanoid pathway as well as genes with no known functions. Overall, we show that cataloguing a biochemical pathway's products through isotopic labeling across genetic variants can substantially contribute to the identification of metabolites and genes associated with their biosynthesis.}, number={3}, journal={PLANT CELL}, publisher={Oxford University Press (OUP)}, author={Simpson, Jeffrey P. and Wunderlich, Cole and Li, Xu and Svedin, Elizabeth and Dilkes, Brian and Chapple, Clint}, year={2021}, month={Mar}, pages={492–510} }
 @article{wu_la hovary_chen_li_eng_vallejo_qu_dewey_2020, title={An Efficient Stevia rebaudiana Transformation System and In vitro Enzyme Assays Reveal Novel Insights into UGT76G1 Function}, volume={10}, ISSN={["2045-2322"]}, DOI={10.1038/s41598-020-60776-y}, abstractNote={Abstract Stevia rebaudiana (Bertoni) is one of a very few plant species that produce zero calorie, sweet compounds known as steviol glycosides (SG). SGs differ in their sweetness and organoleptic properties depending on the number and positioning of sugar groups on the core steviol backbone. There is great interest of modulating the SG profiles of the Stevia plant to enhance the flavor profile for a given application in the food and beverage industries. Here, we report a highly efficient Agrobacterium-mediated stable transformation system using axillary shoots as the initial explant. Using this system, we generated over 200 transgenic Stevia plants overexpressing a specific isoform of UGT76G1 . By comparing the SG profiles among independent transgenic events, we demonstrated that altering UGT76G1 expression can change the ratios of specific SG species. Furthermore, using recombinant proteins produced in E . coli , we show that two closely related UGT76G1 isoforms differ in their substrate specificities, providing new insights into mechanisms underlying the diversity of SG profiles that are observed across Stevia germplasm. Finally, we found evidence suggesting that alternative and/or aberrant splicing may serve to influence the ability of the plant to produce functional UGT76G1 transcripts, and possibly produce enzyme variants within the plant.}, number={1}, journal={SCIENTIFIC REPORTS}, author={Wu, Qian and La Hovary, Christophe and Chen, Han-Yi and Li, Xu and Eng, Hayde and Vallejo, Veronica and Qu, Rongda and Dewey, Ralph E.}, year={2020}, month={Feb} }
 @article{panda_li_wager_chen_li_2020, title={An importin-beta-like protein mediates lignin-modification-induced dwarfism in Arabidopsis}, volume={102}, ISSN={["1365-313X"]}, url={https://doi.org/10.1111/tpj.14701}, DOI={10.1111/tpj.14701}, abstractNote={Perturbation of lignin biosynthesis often results in severe growth and developmental defects in plants, which imposes practical limitations to genetic enhancement of lignocellulosic biomass for biofuel production. Currently, little is known about the cellular and genetic mechanisms of this important phenomenon. Here we show that defects in both cell division and cell expansion underlie the dwarfism of an Arabidopsis lignin mutant ref8, and report the identification of a GROWTH INHIBITION RELIEVED 1 (GIR1) gene from a suppressor screen. GIR1 encodes an importin-beta like protein required for the nuclear import of MYB4, a transcriptional repressor of phenylpropanoid metabolism. Disruption of GIR1 and MYB4 similarly alleviates the cellular defects and growth inhibition in ref8, suggesting that the growth rescue effect of gir1 is likely due to compromised MYB4 transport and function. Importantly, the phenylpropanoid perturbation is not alleviated in gir1 ref8 and myb4 ref8, suggesting that the function of MYB4 in growth inhibition of lignin-modified plants is likely to be distinct from its known role in transcriptional regulation of phenylpropanoid biosynthetic genes. This study also provides evidence that lignin-modification induced dwarfism is not merely due to compromised water transport brought about by lignin deficiency, as gir1 has no effect on the growth inhibition of other lignin mutants that show the collapsed xylem phenotype.}, number={6}, journal={PLANT JOURNAL}, author={Panda, Chinmayee and Li, Xin and Wager, Amanda and Chen, Han-Yi and Li, Xu}, year={2020}, month={Jun}, pages={1281–1293} }
 @article{yang_zhang_luo_liu_shiga_li_kim_rubinelli_overton_subramanyam_et al._2019, title={Overcoming cellulose recalcitrance in woody biomass for the lignin-first biorefinery}, volume={12}, ISSN={1754-6834}, url={http://dx.doi.org/10.1186/s13068-019-1503-y}, DOI={10.1186/s13068-019-1503-y}, abstractNote={Low-temperature swelling of cotton linter cellulose and subsequent gelatinization in trifluoroacetic acid (TFA) greatly enhance rates of enzymatic digestion or maleic acid-AlCl3 catalyzed conversion to hydroxymethylfurfural (HMF) and levulinic acid (LA). However, lignin inhibits low-temperature swelling of TFA-treated intact wood particles from hybrid poplar (Populus tremula × P. alba) and results in greatly reduced yields of glucose or catalytic conversion compared to lignin-free cellulose. Previous studies have established that wood particles from transgenic lines of hybrid poplar with high syringyl (S) lignin content give greater glucose yields following enzymatic digestion.Low-temperature (- 20 °C) treatment of S-lignin-rich poplar wood particles in TFA slightly increased yields of glucose from enzymatic digestions and HMF and LA from maleic acid-AlCl3 catalysis. Subsequent gelatinization at 55 °C resulted in over 80% digestion of cellulose in only 3 to 6 h with high-S-lignin wood, compared to 20-60% digestion in the wild-type poplar hybrid and transgenic lines high in guaiacyl lignin or 5-hydroxy-G lignin. Disassembly of lignin in woody particles by Ni/C catalytic systems improved yields of glucose by enzymatic digestion or catalytic conversion to HMF and LA. Although lignin was completely removed by Ni/C-catalyzed delignification (CDL) treatment, recalcitrance to enzymatic digestion of cellulose from the high-S lines was reduced compared to other lignin variants. However, cellulose still exhibited considerable recalcitrance to complete enzymatic digestion or catalytic conversion after complete delignification. Low-temperature swelling of the CDL-treated wood particles in TFA resulted in nearly complete enzymatic hydrolysis, regardless of original lignin composition.Genetic modification of lignin composition can enhance the portfolio of aromatic products obtained from lignocellulosic biomass while promoting disassembly into biofuel and bioproduct substrates. CDL enhances rates of enzymatic digestion and chemical conversion, but cellulose remains intrinsically recalcitrant. Cold TFA is sufficient to overcome this recalcitrance after CDL treatment. Our results inform a 'no carbon left behind' strategy to convert total woody biomass into lignin, cellulose, and hemicellulose value streams for the future biorefinery.}, number={1}, journal={Biotechnology for Biofuels}, publisher={Springer Science and Business Media LLC}, author={Yang, Haibing and Zhang, Ximing and Luo, Hao and Liu, Baoyuan and Shiga, Tânia M. and Li, Xu and Kim, Jeong Im and Rubinelli, Peter and Overton, Jonathan C. and Subramanyam, Varun and et al.}, year={2019}, month={Jun} }
 @article{reem_chen_hur_zhao_wurtele_li_li_zabotina_2018, title={Comprehensive transcriptome analyses correlated with untargeted metabolome reveal differentially expressed pathways in response to cell wall alterations}, volume={96}, ISSN={["1573-5028"]}, DOI={10.1007/s11103-018-0714-0}, number={4-5}, journal={PLANT MOLECULAR BIOLOGY}, author={Reem, Nathan T. and Chen, Han-Yi and Hur, Manhoi and Zhao, Xuefeng and Wurtele, Eve Syrkin and Li, Xu and Li, Ling and Zabotina, Olga}, year={2018}, month={Mar}, pages={509–529} }
 @article{pais_li_xiang_2018, title={Discovering variation of secondary metabolite diversity and its relationship with disease resistance in Cornus florida
 L.}, volume={8}, ISSN={2045-7758}, url={http://dx.doi.org/10.1002/ece3.4090}, DOI={10.1002/ece3.4090}, abstractNote={Abstract Understanding intraspecific relationships between genetic and functional diversity is a major goal in the field of evolutionary biology and is important for conserving biodiversity. Linking intraspecific molecular patterns of plants to ecological pressures and trait variation remains difficult due to environment‐driven plasticity. Next‐generation sequencing, untargeted liquid chromatography–mass spectrometry (LC‐MS) profiling, and interdisciplinary approaches integrating population genomics, metabolomics, and community ecology permit novel strategies to tackle this problem. We analyzed six natural populations of the disease‐threatened Cornus florida L. from distinct ecological regions using genotype‐by‐sequencing markers and LC‐MS‐based untargeted metabolite profiling. We tested the hypothesis that higher genetic diversity in C. florida yielded higher chemical diversity and less disease susceptibility (screening hypothesis), and we also determined whether genetically similar subpopulations were similar in chemical composition. Most importantly, we identified metabolites that were associated with candidate loci or were predictive biomarkers of healthy or diseased plants after controlling for environment. Subpopulation clustering patterns based on genetic or chemical distances were largely congruent. While differences in genetic diversity were small among subpopulations, we did observe notable similarities in patterns between subpopulation averages of rarefied‐allelic and chemical richness. More specifically, we found that the most abundant compound of a correlated group of putative terpenoid glycosides and derivatives was correlated with tree health when considering chemodiversity. Random forest biomarker and genomewide association tests suggested that this putative iridoid glucoside and other closely associated chemical features were correlated to SNPs under selection.}, number={11}, journal={Ecology and Evolution}, publisher={Wiley}, author={Pais, Andrew L. and Li, Xu and Xiang, Qiu-Yun (Jenny)}, year={2018}, month={May}, pages={5619–5636} }
 @article{wager_li_2018, title={Exploiting natural variation for accelerating discoveries in plant specialized metabolism}, volume={17}, ISSN={["1572-980X"]}, DOI={10.1007/s11101-017-9524-2}, number={1}, journal={PHYTOCHEMISTRY REVIEWS}, author={Wager, Amanda and Li, Xu}, year={2018}, month={Feb}, pages={17–36} }
 @article{yang_he_kabahuma_chaya_kelly_borrego_bian_el kasmi_yang_teixeira_et al._2017, title={A gene encoding maize caffeoyl-CoA O-methyltransferase confers quantitative resistance to multiple pathogens}, volume={49}, ISSN={1061-4036 1546-1718}, url={http://dx.doi.org/10.1038/ng.3919}, DOI={10.1038/ng.3919}, abstractNote={Alleles that confer multiple disease resistance (MDR) are valuable in crop improvement, although the molecular mechanisms underlying their functions remain largely unknown. A quantitative trait locus, qMdr9.02, associated with resistance to three important foliar maize diseases-southern leaf blight, gray leaf spot and northern leaf blight-has been identified on maize chromosome 9. Through fine-mapping, association analysis, expression analysis, insertional mutagenesis and transgenic validation, we demonstrate that ZmCCoAOMT2, which encodes a caffeoyl-CoA O-methyltransferase associated with the phenylpropanoid pathway and lignin production, is the gene within qMdr9.02 conferring quantitative resistance to both southern leaf blight and gray leaf spot. We suggest that resistance might be caused by allelic variation at the level of both gene expression and amino acid sequence, thus resulting in differences in levels of lignin and other metabolites of the phenylpropanoid pathway and regulation of programmed cell death.}, number={9}, journal={Nature Genetics}, publisher={Springer Science and Business Media LLC}, author={Yang, Qin and He, Yijian and Kabahuma, Mercy and Chaya, Timothy and Kelly, Amy and Borrego, Eli and Bian, Yang and El Kasmi, Farid and Yang, Li and Teixeira, Paulo and et al.}, year={2017}, month={Jul}, pages={1364–1372} }
 @article{chen_li_2017, title={Identification of a residue responsible for UDP-sugar donor selectivity of a dihydroxybenzoic acid glycosyltransferase from Arabidopsis natural accessions}, volume={89}, ISSN={["1365-313X"]}, DOI={10.1111/tpj.13271}, abstractNote={Summary UDP‐glycosyltransferase (UGT) plays a major role in the diversity and reactivity of plant specialized metabolites by catalyzing the transfer of the sugar moiety from activated UDP‐sugars to various acceptors. Arabidopsis UGT89A2 was previously identified from a genome‐wide association study as a key factor that affects the differential accumulation of dihydroxybenzoic acid (DHBA) glycosides in distinct Arabidopsis natural accessions, including Col‐0 and C24. The in vitro enzyme assays indicate that these distinct metabolic phenotypes reflect the divergence of UGT89A2 enzyme properties in the Col‐0 and C24 accessions. UGT89A2 from Col‐0 is highly selective toward UDP‐xylose as the sugar donor, and the isoform from C24 can utilize both UDP‐glucose and UDP‐xylose but with a higher affinity to the glucose donor. The sequences of the two isozymes only differ at six amino acid residues. Examination of these amino acid residues in more natural accessions revealed a strong correlation between the amino acid polymorphism at position 153 and the DHBA glycoside accumulation pattern. Site‐directed mutagenesis that swapped residue 153 between UGT89A2 from Col‐0 and C24 reversed the UDP‐sugar preferences, indicating that residue 153 plays an important role in determining sugar donor specificity of UGT89A2. This study provides insight into the key amino acid changes that confer sugar donor selectivity on UGTs, and demonstrates the usefulness of natural variation in understanding the structure–function relationship of enzymes involved in specialized metabolism. Significance Statement Knowledge about enzyme structure‐function relationship is important for understanding catalytic mechanism, function prediction, and engineering novel activities. In this work, natural variation in Arabidopsis UGT89A2 guided the discovery that a single amino acid residue change determines UDP‐sugar donor selectivity of the dihydroxybenzoic acid glycosyltransferase, providing new insights into UDP‐xylose specificity of a class of enzymes important to specialized metabolism.}, number={2}, journal={PLANT JOURNAL}, author={Chen, Han-Yi and Li, Xu}, year={2017}, month={Jan}, pages={195–203} }
 @article{jiao_li_yu_yang_li_shen_2017, title={In situ enhancement of surfactin biosynthesis in Bacillus subtilis
 using novel artificial inducible promoters}, volume={114}, ISSN={0006-3592}, url={http://dx.doi.org/10.1002/BIT.26197}, DOI={10.1002/BIT.26197}, abstractNote={Surfactin‐family lipopeptides are green biosurfactants with substantial industrial potential. The major problem prohibiting surfactin use is the low titer of the wild producer, Bacillus subtilis. Using transcriptomic analysis, four strong promoters, PgroE, Pcdd, PrplK, and PsspE, were identified and cloned from the genome of B. subtilis THY‐7, a novel surfactin producer that has been identified from soil with a 0.55 g/L surfactin titer. An optimal promoter, PgroE, was selected to replace the native THY‐7 surfactin synthase (SrfA) promoter through single‐cross homologous recombination; however, the resulting engineered strain containing the PgroE substitution did not synthesize surfactin. The sucrose‐inducible promoters PsacB and PsacP were then substituted in place of PsrfA, and the resulting engineered strains produced 1.09 and 0.22 g/L surfactin, respectively. An artificial, sucrose‐inducible Pg1 promoter was produced through fusion of the PgroE and PsacB ribonucleic antiterminator (RAT), and the engineered strain containing the Pg1‐substitution produced a surfactin titer of 1.44 g/L. An artificial IPTG‐inducible promoter, Pg2, was constructed from a PgroE‐lacO fusion and then substituted for the chromosomal PsrfA locus, and the surfactin titer of the resulting THY‐7/Pg2‐srfA increased to 5.98 g/L. The driving capacity of Pg2 was further improved by the inclusion of two point mutations in the −35 and −10 regions to produce the novel promoter Pg3. Pg3 exhibited super‐strong activity as measured by lacZ reporter gene overexpression (approximately 3000 U). The Pg3‐substitution strain THY‐7/Pg3‐srfA produced up to 9.74 g/L surfactin in a 5 L fermentor. The maximum productivity was 0.30 g/L/h, and the greatest yield reached 0.14 g surfactin/g sucrose. Biotechnol. Bioeng. 2017;114: 832–842. © 2016 Wiley Periodicals, Inc.}, number={4}, journal={Biotechnology and Bioengineering}, publisher={Wiley}, author={Jiao, Song and Li, Xu and Yu, Huimin and Yang, Huan and Li, Xue and Shen, Zhongyao}, year={2017}, month={Apr}, pages={832–842} }
 @article{rathinasabapathy_palatini jackson_thor_buru_esposito_li_pichika_hamzah_komarnytsky_2017, title={Thiazolopyridines Improve Adipocyte Function by Inhibiting 11 Beta-HSD1 Oxoreductase Activity}, volume={2017}, ISSN={2090-9063 2090-9071}, url={http://dx.doi.org/10.1155/2017/3182129}, DOI={10.1155/2017/3182129}, abstractNote={Background. Glucocorticoid excess has been linked to clinical observations associated with the pathophysiology of metabolic syndrome. The intracellular glucocorticoid levels are primarily modulated by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) enzyme that is highly expressed in key metabolic tissues including fat, liver, and the central nervous system. Methods. In this study we synthesized a set of novel tetrahydrothiazolopyridine derivatives, TR-01–4, that specifically target 11β-HSD1 and studied their ability to interfere with the glucocorticoid and lipid metabolism in the 3T3-L1 adipocytes. Results. Based on the docking model and structure-activity relationships, tetrahydrothiazolopyridine derivatives TR-02 and TR-04 showed the highest potency against 11β-HSD1 by dose-dependently inhibiting conversion of cortisone to cortisol (IC50 values of 1.8 μM and 0.095 μM, resp.). Incubation of fat cells with 0.1–10 μM TR-01–4 significantly decreased cortisone-induced lipid accumulation in adipocytes and suppressed 11β-HSD1 mRNA expression. Observed reduction in adipocyte fat stores could be partially explained by decreased expression levels of adipogenic markers (PPAR-γ, aP2) and key enzymes of lipid metabolism, including fatty acid synthase (FAS), hormone sensitive lipase (HSL), and lipoprotein lipase (LPL). Conclusions. The tetrahydrothiazolopyridine moiety served as an active pharmacophore for inhibiting 11β-HSD1 and offered a novel therapeutic strategy to ameliorate metabolic alterations found in obesity and diabetes.}, journal={Journal of Chemistry}, publisher={Hindawi Limited}, author={Rathinasabapathy, Thirumurugan and Palatini Jackson, Kimberly Marie and Thor, Yiwen and Buru, Ayuba Sunday and Esposito, Debora and Li, Xu and Pichika, Mallikarjuna Rao and Hamzah, Ahmad Sazali and Komarnytsky, Slavko}, year={2017}, pages={1–10} }
 @article{jacobi_yang_li_menze_laurentz_janle_ferruzzi_mccabe_chapple_kirchmaier_et al._2016, title={Impacts on Sirtuin Function and Bioavailability of the Dietary Bioactive Compound Dihydrocoumarin}, volume={11}, ISSN={["1932-6203"]}, DOI={10.1371/journal.pone.0149207}, abstractNote={The plant secondary metabolite and common food additive dihydrocoumarin (DHC) is an inhibitor of the Sirtuin family of NAD+-dependent deacetylases. Sirtuins are key regulators of epigenetic processes that maintain silent chromatin in yeast and have been linked to gene expression, metabolism, apoptosis, tumorogenesis and age-related processes in multiple organisms, including humans. Here we report that exposure to the polyphenol DHC led to defects in several Sirtuin-regulated processes in budding yeast including the establishment and maintenance of Sir2p-dependent silencing by causing disassembly of silent chromatin, Hst1p-dependent repression of meiotic-specific genes during the mitotic cell cycle. As both transient and prolonged exposure to environmental and dietary factors have the potential to lead to heritable alterations in epigenetic states and to modulate additional Sirtuin-dependent phenotypes, we examined the bioavailability and digestive stability of DHC using an in vivo rat model and in vitro digestive simulator. Our analyses revealed that DHC was unstable during digestion and could be converted to melilotic acid (MA), which also caused epigenetic defects, albeit less efficiently. Upon ingestion, DHC was observed primarily in intestinal tissues, but did not accumulate over time and was readily cleared from the animals. MA displayed a wider tissue distribution and, in contrast to DHC, was also detected in the blood plasma, interstitial fluid, and urine, implying that the conversion of DHC to the less bioactive compound, MA, occurred efficiently in vivo.}, number={2}, journal={PLOS ONE}, author={Jacobi, J. L. and Yang, B. and Li, X. and Menze, A. K. and Laurentz, S. M. and Janle, E. M. and Ferruzzi, M. G. and McCabe, G. P. and Chapple, C. and Kirchmaier, A. L. and et al.}, year={2016}, month={Feb} }
 @article{esposito_damsud_wilson_grace_strauch_li_lila_komarnytsky_2015, title={Black Currant Anthocyanins Attenuate Weight Gain and Improve Glucose Metabolism in Diet-Induced Obese Mice with Intact, but Not Disrupted, Gut Microbiome}, volume={63}, ISSN={["1520-5118"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84937064835&partnerID=MN8TOARS}, DOI={10.1021/acs.jafc.5b00963}, abstractNote={Black currant (Ribes nigrum L.) is a rich source of anthocyanins; however, the relationship between their apparently limited bioavailability and significant protection against metabolic pathologies is poorly understood. This study examined the gastrointestinal distribution of black currant anthocyanins and their phenolic acid metabolites in lean and diet-induced obese mice with healthy and antibiotic-disrupted microbiomes. Daily consumption of low- or high-fat diet supplemented with 1% black currant powdered extract (32% anthocyanins) for 8 weeks reduced body weight gain and improved glucose metabolism only in mice with the intact gut microbiome. Administration of antibiotic cocktail resulted in a 16-25-fold increase (P < 0.001) in anthocyanin content of feces, and cyanidin-based anthocyanins showed the largest increase in fecal content upon disruption of gut microbiome (92.3 ± 16.3 vs 4719 ± 158 μg/g feces), indicating their high susceptibility to microbial degradation in the gut. A 3-fold enrichment (P < 0.05) in gallic over protocatechuic acid was observed in the jejunum of both intact and antibiotic-treated animals, suggesting that this effect was likely independent of their gut microbiome status. Taken together, the data clearly demonstrate that gut microbiome and the type of the anthocyanin aglycone moiety can alter the protective effect of anthocyanins against obesity and associated insulin resistance.}, number={27}, journal={JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY}, publisher={American Chemical Society (ACS)}, author={Esposito, Debora and Damsud, Thanakorn and Wilson, Mickey and Grace, Mary H. and Strauch, Renee and Li, Xu and Lila, Mary Ann and Komarnytsky, Slavko}, year={2015}, month={Jul}, pages={6172–6180} }
 @article{strauch_svedin_dilkes_chapple_li_2015, title={Discovery of a novel amino acid racemase through exploration of natural variation in Arabidopsis thaliana}, volume={112}, ISSN={["0027-8424"]}, DOI={10.1073/pnas.1503272112}, abstractNote={Significance We describe how untargeted metabolic profiling and genome-wide association analysis was used in Arabidopsis thaliana to link natural products (secondary metabolites) with genes controlling their production. This powerful approach exposed metabolite–enzyme connections even without prior knowledge of the metabolite identity or the biochemical function of the associated enzyme. Further chemical and genetic analysis synergistically led to the discovery and characterization of a d-amino acid derivative, N-malonyl-d-allo-isoleucine, and a novel amino acid racemase responsible for its biosynthesis. Little is known about d-amino acid metabolism and its natural variation in plants. Additionally, this is the first functional characterization of a eukaryotic member of a large family of phenazine biosynthesis protein phzF-like proteins conserved across all the kingdoms. Plants produce diverse low-molecular-weight compounds via specialized metabolism. Discovery of the pathways underlying production of these metabolites is an important challenge for harnessing the huge chemical diversity and catalytic potential in the plant kingdom for human uses, but this effort is often encumbered by the necessity to initially identify compounds of interest or purify a catalyst involved in their synthesis. As an alternative approach, we have performed untargeted metabolite profiling and genome-wide association analysis on 440 natural accessions of Arabidopsis thaliana. This approach allowed us to establish genetic linkages between metabolites and genes. Investigation of one of the metabolite–gene associations led to the identification of N-malonyl-d-allo-isoleucine, and the discovery of a novel amino acid racemase involved in its biosynthesis. This finding provides, to our knowledge, the first functional characterization of a eukaryotic member of a large and widely conserved phenazine biosynthesis protein PhzF-like protein family. Unlike most of known eukaryotic amino acid racemases, the newly discovered enzyme does not require pyridoxal 5′-phosphate for its activity. This study thus identifies a new d-amino acid racemase gene family and advances our knowledge of plant d-amino acid metabolism that is currently largely unexplored. It also demonstrates that exploitation of natural metabolic variation by integrating metabolomics with genome-wide association is a powerful approach for functional genomics study of specialized metabolism.}, number={37}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, author={Strauch, Renee C. and Svedin, Elisabeth and Dilkes, Brian and Chapple, Clint and Li, Xu}, year={2015}, month={Sep}, pages={11726–11731} }
 @article{wang_he_strauch_olukolu_nielsen_li_balint-kurti_2015, title={Maize Homologs of HCT, a Key Enzyme in Lignin Biosynthesis, Bind the NLR Rp1 Proteins to Modulate the Defense Response}, volume={169}, ISSN={0032-0889 1532-2548}, url={http://dx.doi.org/10.1104/pp.15.00703}, DOI={10.1104/pp.15.00703}, abstractNote={Homologs of hydroxycinnamoyltransferase, involved in lignin biosynthesis, interact directly with leucine-rich receptor proteins to suppress the hypersensitive response. In plants, most disease resistance genes encode nucleotide binding Leu-rich repeat (NLR) proteins that trigger a rapid localized cell death called a hypersensitive response (HR) upon pathogen recognition. The maize (Zea mays) NLR protein Rp1-D21 derives from an intragenic recombination between two NLRs, Rp1-D and Rp1-dp2, and confers an autoactive HR in the absence of pathogen infection. From a previous quantitative trait loci and genome-wide association study, we identified a single-nucleotide polymorphism locus highly associated with variation in the severity of Rp1-D21-induced HR. Two maize genes encoding hydroxycinnamoyltransferase (HCT; a key enzyme involved in lignin biosynthesis) homologs, termed HCT1806 and HCT4918, were adjacent to this single-nucleotide polymorphism. Here, we show that both HCT1806 and HCT4918 physically interact with and suppress the HR conferred by Rp1-D21 but not other autoactive NLRs when transiently coexpressed in Nicotiana benthamiana. Other maize HCT homologs are unable to confer the same level of suppression on Rp1-D21-induced HR. The metabolic activity of HCT1806 and HCT4918 is unlikely to be necessary for their role in suppressing HR. We show that the lignin pathway is activated by Rp1-D21 at both the transcriptional and metabolic levels. We derive a model to explain the roles of HCT1806 and HCT4918 in Rp1-mediated disease resistance.}, number={3}, journal={Plant Physiology}, publisher={American Society of Plant Biologists (ASPB)}, author={Wang, Guan-Feng and He, Yijian and Strauch, Renee and Olukolu, Bode and Nielsen, Dahlia and Li, Xu and Balint-Kurti, Peter}, year={2015}, month={Sep}, pages={pp.00703.2015} }
 @article{kim_ciesielski_donohoe_chapple_li_2014, title={Chemically Induced Conditional Rescue of the Reduced Epidermal Fluorescence8 Mutant of Arabidopsis Reveals Rapid Restoration of Growth and Selective Turnover of Secondary Metabolite Pools}, volume={164}, ISSN={["1532-2548"]}, DOI={10.1104/pp.113.229393}, abstractNote={Activation of the wild-type gene in an Arabidopsis phenylpropanoid mutant at different developmental stages has distinct impacts on metabolic changes and plant growth. The phenylpropanoid pathway is responsible for the biosynthesis of diverse and important secondary metabolites including lignin and flavonoids. The reduced epidermal fluorescence8 (ref8) mutant of Arabidopsis (Arabidopsis thaliana), which is defective in a lignin biosynthetic enzyme p-coumaroyl shikimate 3′-hydroxylase (C3′H), exhibits severe dwarfism and sterility. To better understand the impact of perturbation of phenylpropanoid metabolism on plant growth, we generated a chemically inducible C3′H expression construct and transformed it into the ref8 mutant. Application of dexamethasone to these plants greatly alleviates the dwarfism and sterility and substantially reverses the biochemical phenotypes of ref8 plants, including the reduction of lignin content and hyperaccumulation of flavonoids and p-coumarate esters. Induction of C3′H expression at different developmental stages has distinct impacts on plant growth. Although early induction effectively restored the elongation of primary inflorescence stem, application to 7-week-old plants enabled them to produce new rosette inflorescence stems. Examination of hypocotyls of these plants revealed normal vasculature in the newly formed secondary xylem, presumably restoring water transport in the mutant. The ref8 mutant accumulates higher levels of salicylic acid than the wild type, but depletion of this compound in ref8 did not relieve the mutant’s growth defects, suggesting that the hyperaccumulation of salicylic acid is unlikely to be responsible for dwarfism in this mutant.}, number={2}, journal={PLANT PHYSIOLOGY}, author={Kim, Jeong Im and Ciesielski, Peter N. and Donohoe, Bryon S. and Chapple, Clint and Li, Xu}, year={2014}, month={Feb}, pages={584–595} }
 @article{li_svedin_mo_atwell_dilkes_chapple_2014, title={Exploiting Natural Variation of Secondary Metabolism Identifies a Gene Controlling the Glycosylation Diversity of Dihydroxybenzoic Acids in Arabidopsis thaliana}, volume={198}, ISSN={0016-6731 1943-2631}, url={http://dx.doi.org/10.1534/genetics.114.168690}, DOI={10.1534/genetics.114.168690}, abstractNote={Plant secondary metabolism is an active research area because of the unique and important roles the specialized metabolites have in the interaction of plants with their biotic and abiotic environment, the diversity and complexity of the compounds and their importance to human medicine. Thousands of natural accessions of Arabidopsis thaliana characterized with increasing genomic precision are available, providing new opportunities to explore the biochemical and genetic mechanisms affecting variation in secondary metabolism within this model species. In this study, we focused on four aromatic metabolites that were differentially accumulated among 96 Arabidopsis natural accessions as revealed by leaf metabolic profiling. Using UV, mass spectrometry, and NMR data, we identified these four compounds as different dihydroxybenzoic acid (DHBA) glycosides, namely 2,5-dihydroxybenzoic acid (gentisic acid) 5-O-β-D-glucoside, 2,3-dihydroxybenzoic acid 3-O-β-D-glucoside, 2,5-dihydroxybenzoic acid 5-O-β-D-xyloside, and 2,3-dihydroxybenzoic acid 3-O-β-D-xyloside. Quantitative trait locus (QTL) mapping using recombinant inbred lines generated from C24 and Col-0 revealed a major-effect QTL controlling the relative proportion of xylosides vs. glucosides. Association mapping identified markers linked to a gene encoding a UDP glycosyltransferase gene. Analysis of Transfer DNA (T-DNA) knockout lines verified that this gene is required for DHBA xylosylation in planta and recombinant protein was able to xylosylate DHBA in vitro. This study demonstrates that exploiting natural variation of secondary metabolism is a powerful approach for gene function discovery.}, number={3}, journal={Genetics}, publisher={Genetics Society of America}, author={Li, Xu and Svedin, Elisabeth and Mo, Huaping and Atwell, Susanna and Dilkes, Brian P. and Chapple, Clint}, year={2014}, month={Aug}, pages={1267–1276} }
 @article{rubinelli_chuck_li_meilan_2013, title={Constitutive expression of the Corngrass1 microRNA in poplar affects plant architecture and stem lignin content and composition}, volume={54}, ISSN={0961-9534}, url={http://dx.doi.org/10.1016/j.biombioe.2012.03.001}, DOI={10.1016/j.biombioe.2012.03.001}, abstractNote={A role for microRNAs (miRNAs) in the developmental biology of plants has been established in model annual species, but is poorly understood in perennials, particularly trees. We over-expressed in poplar (genus Populus) a unique miRNA gene from maize belonging to the MIR156 family called Corngrass1 (Cg1) under the control of the cauliflower mosaic virus 35S promoter. Transgenics had significantly greater axillary meristem outgrowth (branching), shorter internode length, and up to a 30% reduction in stem lignin content compared to stem lignin of the wild-type (trunk stem from shoot apex down to node 10). The severity of the phenotype was positively correlated with Cg1 expression level. In addition, the syringyl to guaiacyl ratio (S/G) was lower in 35S:Cg1 lines than in wild-type poplar or a control transgenic line with low abundance of Cg1 transcript. We have demonstrated that over-expression in poplar of a MIR156 class miRNA has dramatic effects on plant architecture, and demonstrated that miRNA over-expression represents a novel approach to altering lignin content and composition in poplar. These transgenic plants may provide a tool for investigating the potential role of MIR156 in regulating developmental processes in poplar. The 35S:Cg1 poplars may also have commercial value as a cellulosic feedstock for biofuel production and in the paper-manufacturing industry.}, journal={Biomass and Bioenergy}, publisher={Elsevier BV}, author={Rubinelli, Peter M. and Chuck, George and Li, Xu and Meilan, Richard}, year={2013}, month={Jul}, pages={312–321} }
 @article{krothapalli_buescher_li_brown_chapple_dilkes_tuinstra_2013, title={Forward Genetics by Genome Sequencing Reveals That Rapid Cyanide Release Deters Insect Herbivory of Sorghum bicolor}, volume={195}, ISSN={0016-6731 1943-2631}, url={http://dx.doi.org/10.1534/genetics.113.149567}, DOI={10.1534/genetics.113.149567}, abstractNote={Whole genome sequencing has allowed rapid progress in the application of forward genetics in model species. In this study, we demonstrated an application of next-generation sequencing for forward genetics in a complex crop genome. We sequenced an ethyl methanesulfonate-induced mutant of Sorghum bicolor defective in hydrogen cyanide release and identified the causal mutation. A workflow identified the causal polymorphism relative to the reference BTx623 genome by integrating data from single nucleotide polymorphism identification, prior information about candidate gene(s) implicated in cyanogenesis, mutation spectra, and polymorphisms likely to affect phenotypic changes. A point mutation resulting in a premature stop codon in the coding sequence of dhurrinase2, which encodes a protein involved in the dhurrin catabolic pathway, was responsible for the acyanogenic phenotype. Cyanogenic glucosides are not cyanogenic compounds but their cyanohydrins derivatives do release cyanide. The mutant accumulated the glucoside, dhurrin, but failed to efficiently release cyanide upon tissue disruption. Thus, we tested the effects of cyanide release on insect herbivory in a genetic background in which accumulation of cyanogenic glucoside is unchanged. Insect preference choice experiments and herbivory measurements demonstrate a deterrent effect of cyanide release capacity, even in the presence of wild-type levels of cyanogenic glucoside accumulation. Our gene cloning method substantiates the value of (1) a sequenced genome, (2) a strongly penetrant and easily measurable phenotype, and (3) a workflow to pinpoint a causal mutation in crop genomes and accelerate in the discovery of gene function in the postgenomic era.}, number={2}, journal={Genetics}, publisher={Genetics Society of America}, author={Krothapalli, Kartikeya and Buescher, Elizabeth M. and Li, Xu and Brown, Elliot and Chapple, Clint and Dilkes, Brian P. and Tuinstra, Mitchell R.}, year={2013}, month={Jul}, pages={309–318} }
 @article{venkata_lauter_li_chapple_krupke_johal_moose_2013, title={crw1 - A Novel Maize Mutant Highly Susceptible to Foliar Damage by the Western Corn Rootworm Beetle}, volume={8}, ISSN={["1932-6203"]}, DOI={10.1371/journal.pone.0071296}, abstractNote={Western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), is the most destructive insect pest of corn (Zea mays L.) in the United States. The adult WCR beetles derive their nourishment from multiple sources including corn pollen and silks as well as the pollen of alternate hosts. Conversely, the corn foliage is largely neglected as a food source by WCR beetles, leading to a perception of a passive interaction between the two. We report here a novel recessive mutation of corn that was identified and named after its foliar susceptibility to corn rootworm beetles (crw1). The crw1 mutant under field conditions was exceptionally susceptible to foliar damage by WCR beetles in an age-specific manner. It exhibits pleiotropic defects on cell wall biochemistry, morphology of leaf epidermal cells and lower structural integrity via differential accumulation of cell wall bound phenolic acids. These findings indicate that crw1 is perturbed in a pathway that was not previously ascribed to WCR susceptibility, as well as implying the presence of an active mechanism(s) deterring WCR beetles from devouring corn foliage. The discovery and characterization of this mutant provides a unique opportunity for genetic analysis of interactions between maize and adult WCR beetles and identify new strategies to control the spread and invasion of this destructive pest.}, number={8}, journal={PLOS ONE}, author={Venkata, Bala Puchakayala and Lauter, Nick and Li, Xu and Chapple, Clint and Krupke, Christian and Johal, Gurmukh and Moose, Stephen}, year={2013}, month={Aug} }
 @article{weng_akiyama_bonawitz_li_ralph_chapple_2010, title={Convergent Evolution of Syringyl Lignin Biosynthesis via Distinct Pathways in the Lycophyte Selaginella and Flowering Plants}, volume={22}, ISSN={1040-4651 1532-298X}, url={http://dx.doi.org/10.1105/tpc.109.073528}, DOI={10.1105/tpc.109.073528}, abstractNote={This study shows that the independent origin of syringyl lignin in the lycophyte Selaginella involved the elaboration of a biochemical pathway that bypasses four steps of the canonical lignin biosynthetic pathway established in flowering plants. Phenotypic convergence in unrelated lineages arises when different organisms adapt similarly under comparable selective pressures. In an apparent example of this process, syringyl lignin, a fundamental building block of plant cell walls, occurs in two major plant lineages, lycophytes and angiosperms, which diverged from one another more than 400 million years ago. Here, we show that this convergence resulted from independent recruitment of lignin biosynthetic cytochrome P450-dependent monooxygenases that route cell wall monomers through related but distinct pathways in the two lineages. In contrast with angiosperms, in which syringyl lignin biosynthesis requires two phenylpropanoid meta-hydroxylases C3′H and F5H, the lycophyte Selaginella employs one phenylpropanoid dual meta-hydroxylase to bypass several steps of the canonical lignin biosynthetic pathway. Transgenic expression of the Selaginella hydroxylase in Arabidopsis thaliana dramatically reroutes its endogenous lignin biosynthetic pathway, yielding a novel lignin composition not previously identified in nature. Our findings demonstrate a unique case of convergent evolution via distinct biochemical strategies and suggest a new way to genetically reconstruct lignin biosynthesis in higher plants.}, number={4}, journal={The Plant Cell}, publisher={American Society of Plant Biologists (ASPB)}, author={Weng, Jing-Ke and Akiyama, Takuya and Bonawitz, Nicholas D. and Li, Xu and Ralph, John and Chapple, Clint}, year={2010}, month={Apr}, pages={1033–1045} }
 @article{li_ximenes_kim_slininger_meilan_ladisch_chapple_2010, title={Lignin monomer composition affects Arabidopsis cell-wall degradability after liquid hot water pretreatment}, volume={3}, ISSN={1754-6834}, url={http://dx.doi.org/10.1186/1754-6834-3-27}, DOI={10.1186/1754-6834-3-27}, abstractNote={Abstract Background Lignin is embedded in the plant cell wall matrix, and impedes the enzymatic saccharification of lignocellulosic feedstocks. To investigate whether enzymatic digestibility of cell wall materials can be improved by altering the relative abundance of the two major lignin monomers, guaiacyl (G) and syringyl (S) subunits, we compared the degradability of cell wall material from wild-type Arabidopsis thaliana with a mutant line and a genetically modified line, the lignins of which are enriched in G and S subunits, respectively. Results Arabidopsis tissue containing G- and S-rich lignins had the same saccharification performance as the wild type when subjected to enzyme hydrolysis without pretreatment. After a 24-hour incubation period, less than 30% of the total glucan was hydrolyzed. By contrast, when liquid hot water (LHW) pretreatment was included before enzyme hydrolysis, the S-lignin-rich tissue gave a much higher glucose yield than either the wild-type or G-lignin-rich tissue. Applying a hot-water washing step after the pretreatment did not lead to a further increase in final glucose yield, but the initial hydrolytic rate was doubled. Conclusions Our analyses using the model plant A. thaliana revealed that lignin composition affects the enzymatic digestibility of LHW pretreated plant material. Pretreatment is more effective in enhancing the saccharification of A. thaliana cell walls that contain S-rich lignin. Increasing lignin S monomer content through genetic engineering may be a promising approach to increase the efficiency and reduce the cost of biomass to biofuel conversion.}, number={1}, journal={Biotechnology for Biofuels}, publisher={Springer Science and Business Media LLC}, author={Li, Xu and Ximenes, Eduardo and Kim, Youngmi and Slininger, Mary and Meilan, Richard and Ladisch, Michael and Chapple, Clint}, year={2010}, pages={27} }
 @article{li_ilarslan_brachova_qian_li_che_wurtele_nikolau_2010, title={Reverse-Genetic Analysis of the Two Biotin-Containing Subunit Genes of the Heteromeric Acetyl-Coenzyme A Carboxylase in Arabidopsis Indicates a Unidirectional Functional Redundancy}, volume={155}, ISSN={0032-0889 1532-2548}, url={http://dx.doi.org/10.1104/pp.110.165910}, DOI={10.1104/pp.110.165910}, abstractNote={The heteromeric acetyl-coenzyme A carboxylase catalyzes the first and committed reaction of de novo fatty acid biosynthesis in plastids. This enzyme is composed of four subunits: biotin carboxyl-carrier protein (BCCP), biotin carboxylase, α-carboxyltransferase, and β-carboxyltransferase. With the exception of BCCP, single-copy genes encode these subunits in Arabidopsis (Arabidopsis thaliana). Reverse-genetic approaches were used to individually investigate the physiological significance of the two paralogous BCCP-coding genes, CAC1A (At5g16390, codes for BCCP1) and CAC1B (At5g15530, codes for BCCP2). Transfer DNA insertional alleles that completely eliminate the accumulation of BCCP2 have no perceptible effect on plant growth, development, and fatty acid accumulation. In contrast, transfer DNA insertional null allele of the CAC1A gene is embryo lethal and deleteriously affects pollen development and germination. During seed development the effect of the cac1a null allele first becomes apparent at 3-d after flowering, when the synchronous development of the endosperm and embryo is disrupted. Characterization of CAC1A antisense plants showed that reducing BCCP1 accumulation to 35% of wild-type levels, decreases fatty acid accumulation and severely affects normal vegetative plant growth. Detailed expression analysis by a suite of approaches including in situ RNA hybridization, promoter:reporter transgene expression, and quantitative western blotting reveal that the expression of CAC1B is limited to a subset of the CAC1A-expressing tissues, and CAC1B expression levels are only about one-fifth of CAC1A expression levels. Therefore, a likely explanation for the observed unidirectional redundancy between these two paralogous genes is that whereas the BCCP1 protein can compensate for the lack of BCCP2, the absence of BCCP1 cannot be tolerated as BCCP2 levels are not sufficient to support heteromeric acetyl-coenzyme A carboxylase activity at a level that is required for normal growth and development.}, number={1}, journal={Plant Physiology}, publisher={American Society of Plant Biologists (ASPB)}, author={Li, Xu and Ilarslan, Hilal and Brachova, Libuse and Qian, Hui-Rong and Li, Ling and Che, Ping and Wurtele, Eve Syrkin and Nikolau, Basil J.}, year={2010}, month={Oct}, pages={293–314} }
 @article{li_bergelson_chapple_2010, title={The ARABIDOPSIS Accession Pna-10 Is a Naturally Occurring sng1 Deletion Mutant}, volume={3}, ISSN={1674-2052}, url={http://dx.doi.org/10.1093/mp/ssp090}, DOI={10.1093/mp/ssp090}, abstractNote={Sinapoylmalate is the major sinapate ester found in leaves of Arabidopsis thaliana, where it plays an important role in UV-B protection. Metabolic profiling of rosette leaves from 96 Arabidopsis accessions revealed that the Pna-10 accession accumulates sinapoylglucose instead of sinapoylmalate. This unique leaf sinapate ester profile is similar to that of the previously characterized sinapoylglucose accumulator1 (sng1) mutants. SNG1 encodes sinapoylglucose:malate sinapoyltransferase (SMT), a serine carboxypeptidase-like (SCPL) enzyme that catalyzes the conversion of sinapoylglucose to sinapoylmalate. In the reference Columbia genome, the SNG1 gene is located in a cluster of five SCPL genes on Chromosome II. PCR and sequencing analysis of the same genomic region in the Pna-10 accession revealed a 13-kb deletion that eliminates the SNG1 gene (At2g22990) and the gene encoding sinapoylglucose:anthocyanin sinapoyltransferase (SAT) (At2g23000). In addition to its sinapoylmalate-deficient phenotype, and consistent with the loss of SAT, Pna-10 is unable to accumulate sinapoylated anthocyanins. Interestingly, the Pna-17 accession, collected from the same location as Pna-10, has no such deletion. Further analysis of 135 lines collected from the same location as Pna-10 and Pna-17 revealed that four more lines contain the deletion found in Pna-10 accession, suggesting that either the deletion found in Pna-10 is a recent event that has not yet been eliminated through selection or that sinapoylmalate is dispensable for the growth of Arabidopsis under field conditions.}, number={1}, journal={Molecular Plant}, publisher={Elsevier BV}, author={Li, X.u. and Bergelson, Joy and Chapple, Clint}, year={2010}, month={Jan}, pages={91–100} }
 @article{li_bonawitz_weng_chapple_2010, title={The Growth Reduction Associated with Repressed Lignin Biosynthesis in Arabidopsis thaliana Is Independent of Flavonoids}, volume={22}, ISSN={1040-4651 1532-298X}, url={http://dx.doi.org/10.1105/tpc.110.074161}, DOI={10.1105/tpc.110.074161}, abstractNote={This study reexamines the relationship between flavonoid accumulation and the growth reduction exhibited by Arabidopsis plants that are defective in lignin biosynthesis. It shows that flavonoids are not responsible for the observed growth inhibition, contrary to a previous report. Defects in phenylpropanoid biosynthesis arising from deficiency in hydroxycinnamoyl CoA:shikimate hydroxycinnamoyl transferase (HCT) or p-coumaroyl shikimate 3′-hydroxylase (C3′H) lead to reduced lignin, hyperaccumulation of flavonoids, and growth inhibition in Arabidopsis thaliana. It was previously reported that flavonoid-mediated inhibition of auxin transport is responsible for growth reduction in HCT-RNA interference (RNAi) plants. This conclusion was based on the observation that simultaneous RNAi silencing of HCT and chalcone synthase (CHS), an enzyme essential for flavonoid biosynthesis, resulted in less severe dwarfing than silencing of HCT alone. In an attempt to extend these results using a C3′H mutant (ref8) and a CHS null mutant (tt4-2), we found that the growth phenotype of the ref8 tt4-2 double mutant, which lacks flavonoids, is indistinguishable from that of ref8. Moreover, using RNAi, we found that the relationship between HCT silencing and growth inhibition is identical in both the wild type and tt4-2. We conclude from these results that the growth inhibition observed in HCT-RNAi plants and the ref8 mutant is independent of flavonoids. Finally, we show that expression of a newly characterized gene bypassing HCT and C3′H partially restores both lignin biosynthesis and growth in HCT-RNAi plants, demonstrating that a biochemical pathway downstream of coniferaldehyde, probably lignification, is essential for normal plant growth.}, number={5}, journal={The Plant Cell}, publisher={American Society of Plant Biologists (ASPB)}, author={Li, Xu and Bonawitz, Nicholas D. and Weng, Jing-Ke and Chapple, Clint}, year={2010}, month={May}, pages={1620–1632} }
 @article{li_chapple_2010, title={Understanding Lignification: Challenges Beyond Monolignol Biosynthesis}, volume={154}, ISSN={0032-0889 1532-2548}, url={http://dx.doi.org/10.1104/pp.110.162842}, DOI={10.1104/pp.110.162842}, abstractNote={Lignin, a major component of vascular plant cell wall, provides mechanical support for plants to stand upright and enables xylems to withstand the negative pressure generated during water transport. Although important for plant growth, the presence of lignin limits access to cell wall}, number={2}, journal={Plant Physiology}, publisher={American Society of Plant Biologists (ASPB)}, author={Li, Xu and Chapple, Clint}, year={2010}, month={Oct}, pages={449–452} }
 @article{weng_li_bonawitz_chapple_2008, title={Emerging strategies of lignin engineering and degradation for cellulosic biofuel production}, volume={19}, ISSN={0958-1669}, url={http://dx.doi.org/10.1016/j.copbio.2008.02.014}, DOI={10.1016/j.copbio.2008.02.014}, abstractNote={Ethanol and other biofuels produced from lignocellulosic biomass represent a renewable, more carbon-balanced alternative to both fossil fuels and corn-derived or sugarcane-derived ethanol. Unfortunately, the presence of lignin in plant cell walls impedes the breakdown of cell wall polysaccharides to simple sugars and the subsequent conversion of these sugars to usable fuel. Recent advances in the understanding of lignin composition, polymerization, and regulation have revealed new opportunities for the rational manipulation of lignin in future bioenergy crops, augmenting the previous successful approach of manipulating lignin monomer biosynthesis. Furthermore, recent studies on lignin degradation in nature may provide novel resources for the delignification of dedicated bioenergy crops and other sources of lignocellulosic biomass.}, number={2}, journal={Current Opinion in Biotechnology}, publisher={Elsevier BV}, author={Weng, Jing-Ke and Li, Xu and Bonawitz, Nicholas D and Chapple, Clint}, year={2008}, month={Apr}, pages={166–172} }
 @article{li_weng_chapple_2008, title={Improvement of biomass through lignin modification}, volume={54}, ISSN={0960-7412 1365-313X}, url={http://dx.doi.org/10.1111/j.1365-313x.2008.03457.x}, DOI={10.1111/j.1365-313x.2008.03457.x}, abstractNote={Lignin, a major component of the cell wall of vascular plants, has long been recognized for its negative impact on forage quality, paper manufacturing, and, more recently, cellulosic biofuel production. Over the last two decades, genetic and biochemical analyses of brown midrib mutants of maize, sorghum and related grasses have advanced our understanding of the relationship between lignification and forage digestibility. This work has also inspired genetic engineering efforts aimed at generating crops with altered lignin, with the expectation that these strategies would enhance forage digestibility and/or pulping efficiency. The knowledge gained from these bioengineering efforts has greatly improved our understanding of the optimal lignin characteristics required for various applications of lignocellulosic materials while also contributing to our understanding of the lignin biosynthetic pathway. The recent upswing of interest in cellulosic biofuel production has become the new focus of lignin engineering. Populus trichocarpa and Brachypodium distachyon are emerging as model systems for energy crops. Lignin research on these systems, as well as on a variety of proposed energy crop species, is expected to shed new light on lignin biosynthesis and its regulation in energy crops, and lead to rational genetic engineering approaches to modify lignin for improved biofuel production.}, number={4}, journal={The Plant Journal}, publisher={Wiley}, author={Li, Xu and Weng, Jing-Ke and Chapple, Clint}, year={2008}, month={May}, pages={569–581} }
 @article{weng_li_stout_chapple_2008, title={Independent origins of syringyl lignin in vascular plants}, volume={105}, ISSN={0027-8424 1091-6490}, url={http://dx.doi.org/10.1073/pnas.0801696105}, DOI={10.1073/pnas.0801696105}, abstractNote={Lycophytes arose in the early Silurian (≈400 Mya) and represent a major lineage of vascular plants that has evolved in parallel with the ferns, gymnosperms, and angiosperms. A hallmark of vascular plants is the presence of the phenolic lignin heteropolymer in xylem and other sclerified cell types. Although syringyl lignin is often considered to be restricted in angiosperms, it has been detected in lycophytes as well. Here we report the characterization of a cytochrome P450-dependent monooxygenase from the lycophyte Selaginella moellendorffii. Gene expression data, cross-species complementation experiments, and in vitro enzyme assays indicate that this P450 is a ferulic acid/coniferaldehyde/coniferyl alcohol 5-hydroxylase (F5H), and is capable of diverting guaiacyl-substituted intermediates into syringyl lignin biosynthesis. Phylogenetic analysis indicates that the Selaginella F5H represents a new family of plant P450s and suggests that it has evolved independently of angiosperm F5Hs.}, number={22}, journal={Proceedings of the National Academy of Sciences}, publisher={Proceedings of the National Academy of Sciences}, author={Weng, J.-K. and Li, X. and Stout, J. and Chapple, C.}, year={2008}, month={May}, pages={7887–7892} }