@article{land_sheppard_doherty_perera_2024, title={Conserved plant transcriptional responses to microgravity from two consecutive spaceflight experiments}, volume={14}, ISSN={["1664-462X"]}, url={http://dx.doi.org/10.3389/fpls.2023.1308713}, DOI={10.3389/fpls.2023.1308713}, abstractNote={IntroductionUnderstanding how plants adapt to the space environment is essential, as plants will be a valuable component of long duration space missions. Several spaceflight experiments have focused on transcriptional profiling as a means of understanding plant adaptation to microgravity. However, there is limited overlap between results from different experiments. Differences in experimental conditions and hardware make it difficult to find a consistent response across experiments and to distinguish the primary effects of microgravity from other spaceflight effects.MethodsPlant Signaling (PS) and Plant RNA Regulation (PRR) were two separate spaceflight experiments conducted on the International Space Station utilizing the European Modular Cultivation System (EMCS). The EMCS provided a lighted environment for plant growth with centrifugal capabilities providing an onboard 1 g control.Results and discussionAn RNA-Seq analysis of shoot samples from PS and PRR revealed a significant overlap of genes differentially expressed in microgravity between the two experiments. Relative to onboard 1 g controls, genes involved in transcriptional regulation, shoot development, and response to auxin and light were upregulated in microgravity in both experiments. Conversely, genes involved in defense response, abiotic stress, Ca++ signaling, and cell wall modification were commonly downregulated in both datasets. The downregulation of stress responses in microgravity in these two experiments is interesting as these pathways have been previously observed as upregulated in spaceflight compared to ground controls. Similarly, we have observed many stress response genes to be upregulated in the 1 g onboard control compared to ground reference controls; however these genes were specifically downregulated in microgravity. In addition, we analyzed the sRNA landscape of the 1 g and microgravity (μ g) shoot samples from PRR. We identified three miRNAs (miR319c, miR398b, and miR8683) which were upregulated in microgravity, while several of their corresponding target genes were found to be downregulated in microgravity. Interestingly, the downregulated target genes are enriched in those encoding chloroplast-localized enzymes and proteins. These results uncover microgravity unique transcriptional changes and highlight the validity and importance of an onboard 1 g control.}, journal={FRONTIERS IN PLANT SCIENCE}, author={Land, Eric S. and Sheppard, James and Doherty, Colleen J. and Perera, Imara Y.}, year={2024}, month={Jan} }
 @article{barcenilla_meyers_castillo-gonzalez_young_min_song_phadke_land_canaday_perera_et al._2023, title={Arabidopsis telomerase takes off by uncoupling enzyme activity from telomere length maintenance in space}, volume={14}, ISSN={["2041-1723"]}, url={https://doi.org/10.1038/s41467-023-41510-4}, DOI={10.1038/s41467-023-41510-4}, abstractNote={AbstractSpaceflight-induced changes in astronaut telomeres have garnered significant attention in recent years. While plants represent an essential component of future long-duration space travel, the impacts of spaceflight on plant telomeres and telomerase have not been examined. Here we report on the telomere dynamics of Arabidopsis thaliana grown aboard the International Space Station. We observe no changes in telomere length in space-flown Arabidopsis seedlings, despite a dramatic increase in telomerase activity (up to 150-fold in roots), as well as elevated genome oxidation. Ground-based follow up studies provide further evidence that telomerase is induced by different environmental stressors, but its activity is uncoupled from telomere length. Supporting this conclusion, genetically engineered super-telomerase lines with enhanced telomerase activity maintain wildtype telomere length. Finally, genome oxidation is inversely correlated with telomerase activity levels. We propose a redox protective capacity for Arabidopsis telomerase that may promote survivability in harsh environments.}, number={1}, journal={NATURE COMMUNICATIONS}, author={Barcenilla, Borja Barbero and Meyers, Alexander D. and Castillo-Gonzalez, Claudia and Young, Pierce and Min, Ji-Hee and Song, Jiarui and Phadke, Chinmay and Land, Eric and Canaday, Emma and Perera, Imara Y. and et al.}, year={2023}, month={Nov} }
 @article{land_canaday_meyers_wyatt_perera_2023, title={Bridging the gap: parallel profiling of ribosome associated and total RNA species can identify transcriptional regulatory mechanisms of plants in spaceflight}, volume={18}, ISSN={["1742-9153"]}, url={https://doi.org/10.1080/17429145.2023.2248173}, DOI={10.1080/17429145.2023.2248173}, abstractNote={As plants are an essential component of sustainable life support systems, long-duration space missions will require a sophisticated understanding of plant adaptations to spaceflight and microgravity. For many years, transcriptional profiling of steady state mRNA abundances has been used as measure of plant adaptations to the space environment. However, measured changes in transcript abundances are often not reflected in corresponding changes in the proteome due regulatory processes governing translation. Translating ribosome affinity purification (TRAP) is a technique which selectively targets ribosome bound mRNAs for isolation and downstream sequencing. Comparing profiles of ribosome associated mRNAs with total mRNAs provides insight into the translatome and may more accurately inform on the cellular responses to the spaceflight environment. Toward that goal, this work describes a methodology developed ahead of the APEx-07 flight mission.}, number={1}, journal={JOURNAL OF PLANT INTERACTIONS}, author={Land, Eric S. and Canaday, Emma and Meyers, Alexander and Wyatt, Sarah and Perera, Imara Y.}, year={2023}, month={Dec} }
 @article{barker_kruse_johnson_saravia-butler_fogle_chang_trane_kinscherf_villacampa_manzano_et al._2023, title={Meta-analysis of the space flight and microgravity response of the Arabidopsis plant transcriptome}, volume={9}, ISSN={["2373-8065"]}, DOI={10.1038/s41526-023-00247-6}, abstractNote={AbstractSpaceflight presents a multifaceted environment for plants, combining the effects on growth of many stressors and factors including altered gravity, the influence of experiment hardware, and increased radiation exposure. To help understand the plant response to this complex suite of factors this study compared transcriptomic analysis of 15 Arabidopsis thaliana spaceflight experiments deposited in the National Aeronautics and Space Administration’s GeneLab data repository. These data were reanalyzed for genes showing significant differential expression in spaceflight versus ground controls using a single common computational pipeline for either the microarray or the RNA-seq datasets. Such a standardized approach to analysis should greatly increase the robustness of comparisons made between datasets. This analysis was coupled with extensive cross-referencing to a curated matrix of metadata associated with these experiments. Our study reveals that factors such as analysis type (i.e., microarray versus RNA-seq) or environmental and hardware conditions have important confounding effects on comparisons seeking to define plant reactions to spaceflight. The metadata matrix allows selection of studies with high similarity scores, i.e., that share multiple elements of experimental design, such as plant age or flight hardware. Comparisons between these studies then helps reduce the complexity in drawing conclusions arising from comparisons made between experiments with very different designs.}, number={1}, journal={NPJ MICROGRAVITY}, author={Barker, Richard and Kruse, Colin P. S. and Johnson, Christina and Saravia-Butler, Amanda and Fogle, Homer and Chang, Hyun-Seok and Trane, Ralph Moller and Kinscherf, Noah and Villacampa, Alicia and Manzano, Aranzazu and et al.}, year={2023}, month={Mar} }
 @article{phillippy_donahue_williams_cridland_perera_gillaspy_2023, title={Regulation of inositol 1,2,4,5,6-pentakisphosphate and inositol hexakisphosphate levels in Gossypium hirsutum by IPK1}, volume={257}, ISSN={["1432-2048"]}, DOI={10.1007/s00425-023-04080-9}, abstractNote={The IPK1 genes, which code for 2-kinases that can synthesize Ins(1,2,4,5,6)P 5  from Ins(1,4,5,6)P 4 , are expressed throughout cotton plants, resulting in the highest Ins(1,2,4,5,6)P 5  concentrations in young leaves and flower buds. Cotton leaves contain large amounts of Ins(1,2,4,5,6)P 5  and InsP 6  compared to plants not in the Malvaceae family. The inositol polyphosphate pathway has been linked to stress tolerance in numerous plant species. Accordingly, we sought to determine why cotton and other Malvaceae have such high levels of these inositol phosphates. We have quantified the levels of InsP 5  and InsP 6  in different tissues of cotton plants and determined the expression of IPK1 (inositol 1,3,4,5,6-pentakisphosphate 2-kinase gene) in vegetative and reproductive tissues. Gossypium hirsutum was found to contain four IPK1 genes that were grouped into two pair (AB, CD) where each pair consists of very similar sequences that were measured together. More IPK1AB is expressed in leaves than in roots, whereas more IPK1CD is expressed in roots than in leaves. Leaves and flower buds have more InsP 5  and InsP 6  than stems and roots. Leaves and roots contain more InsP 5  than InsP 6 , whereas flower buds and stems contain more InsP 6  than InsP 5 . Dark-grown seedlings contain more InsP 5  and InsP 6  than those grown under lights, and the ratio of InsP 5  to InsP 6  is greater in the light-grown seedlings. During 35 days of the life cycle of the third true leaf, InsP 5  and InsP 6  gradually decreased by more than 50%. Silencing IPK1AB and IPK1CD with Cotton Leaf Crumple Virus-induced gene silencing (VIGS) resulted in plants with an intense viral phenotype, reduced IPK1AB expression and lowered amounts of InsP 5 . The results are consistent with Ins(1,2,4,5,6)P 5  synthesis from Ins(1,4,5,6)P 4  by IPK1. This study detailed the central role of IPK1 in cotton inositol polyphosphate metabolism, which has potential to be harnessed to improve the resistance of plants to different kinds of stress.}, number={2}, journal={PLANTA}, author={Phillippy, Brian Q. and Donahue, Janet L. and Williams, Sarah P. and Cridland, Caitlin A. and Perera, Imara Y. and Gillaspy, Glenda E.}, year={2023}, month={Feb} }
 @article{land_cridland_craige_dye_hildreth_helm_gillaspy_perera_2021, title={A Role for Inositol Pyrophosphates in the Metabolic Adaptations to Low Phosphate in Arabidopsis}, volume={11}, ISSN={["2218-1989"]}, url={https://doi.org/10.3390/metabo11090601}, DOI={10.3390/metabo11090601}, abstractNote={Phosphate is a major plant macronutrient and low phosphate availability severely limits global crop productivity. In Arabidopsis, a key regulator of the transcriptional response to low phosphate, phosphate starvation response 1 (PHR1), is modulated by a class of signaling molecules called inositol pyrophosphates (PP-InsPs). Two closely related diphosphoinositol pentakisphosphate enzymes (AtVIP1 and AtVIP2) are responsible for the synthesis and turnover of InsP8, the most implicated molecule. This study is focused on characterizing Arabidopsis vip1/vip2 double mutants and their response to low phosphate. We present evidence that both local and systemic responses to phosphate limitation are dampened in the vip1/vip2 mutants as compared to wild-type plants. Specifically, we demonstrate that under Pi-limiting conditions, the vip1/vip2 mutants have shorter root hairs and lateral roots, less accumulation of anthocyanin and less accumulation of sulfolipids and galactolipids. However, phosphate starvation response (PSR) gene expression is unaffected. Interestingly, many of these phenotypes are opposite to those exhibited by other mutants with defects in the PP-InsP synthesis pathway. Our results provide insight on the nexus between inositol phosphates and pyrophosphates involved in complex regulatory mechanisms underpinning phosphate homeostasis in plants.}, number={9}, journal={METABOLITES}, publisher={MDPI AG}, author={Land, Eric S. and Cridland, Caitlin A. and Craige, Branch and Dye, Anna and Hildreth, Sherry B. and Helm, Rich F. and Gillaspy, Glenda E. and Perera, Imara Y.}, year={2021}, month={Sep} }
 @article{tolsma_torres_richards_perera_doherty_2022, title={Evaluating the Effects of the Circadian Clock and Time of Day on Plant Gravitropic Responses}, volume={2368}, ISBN={["978-1-0716-1676-5"]}, ISSN={["1940-6029"]}, url={https://doi.org/10.1007/978-1-0716-1677-2_19}, DOI={10.1007/978-1-0716-1677-2_19}, abstractNote={Circadian rhythms are regular oscillations of an organism's physiology with a period of approximately 24 h. In the model plant Arabidopsis thaliana, circadian rhythms regulate a suite of physiological processes, including transcription, photosynthesis, growth, and flowering. The circadian clock and external rhythmic factors have extensive control of the underlying biochemistry and physiology. Therefore, it is critical to consider the time of day when performing gravitropism experiments, even if the circadian clock is not a focus of study. We describe the critical factors and methods to be considered and methods to investigate the possible circadian regulation of gravitropic responses.}, journal={PLANT GRAVITROPISM}, publisher={Springer US}, author={Tolsma, Joseph S. and Torres, Jacob J. and Richards, Jeffrey T. and Perera, Imara Y. and Doherty, Colleen J.}, year={2022}, pages={301–319} }
 @article{overbey_saravia-butler_zhang_rathi_fogle_silveira_barker_bass_beheshti_berrios_et al._2021, title={NASA GeneLab RNA-seq consensus pipeline: standardized processing of short-read RNA-seq data}, volume={24}, ISSN={["2589-0042"]}, DOI={10.1016/j.isci.2021.102361}, abstractNote={With the development of transcriptomic technologies, we are able to quantify precise changes in gene expression profiles from astronauts and other organisms exposed to spaceflight. Members of NASA GeneLab and GeneLab-associated analysis working groups (AWGs) have developed a consensus pipeline for analyzing short-read RNA-sequencing data from spaceflight-associated experiments. The pipeline includes quality control, read trimming, mapping, and gene quantification steps, culminating in the detection of differentially expressed genes. This data analysis pipeline and the results of its execution using data submitted to GeneLab are now all publicly available through the GeneLab database. We present here the full details and rationale for the construction of this pipeline in order to promote transparency, reproducibility, and reusability of pipeline data; to provide a template for data processing of future spaceflight-relevant datasets; and to encourage cross-analysis of data from other databases with the data available in GeneLab.}, number={4}, journal={ISCIENCE}, author={Overbey, Eliah G. and Saravia-Butler, Amanda M. and Zhang, Zhe and Rathi, Komal S. and Fogle, Homer and Silveira, Willian A. and Barker, Richard J. and Bass, Joseph J. and Beheshti, Afshin and Berrios, Daniel C. and et al.}, year={2021}, month={Apr} }
 @article{tolsma_ryan_torres_richards_richardson_land_perera_doherty_2021, title={The Circadian-clock Regulates the Arabidopsis Gravitropic Response}, url={https://doi.org/10.2478/gsr-2021-0014}, DOI={10.2478/gsr-2021-0014}, abstractNote={Abstract
               For long-term space missions, it is necessary to understand how organisms respond to changes in gravity. Plant roots are positively gravitropic; the primary root grows parallel to gravity's pull even after being turned away from the direction of gravity. We examined if this gravitropic response varies depending on the time of day reorientation occurs. When plants were reoriented in relation to the gravity vector or placed in simulated microgravity, the magnitude of the root gravitropic response varied depending on the time of day the initial change in gravity occurred. The response was greatest when plants were reoriented at dusk, just before a period of rapid growth, and were minimal just before dawn as the plants entered a period of reduced root growth. We found that this variation in the magnitude of the gravitropic response persisted in constant light (CL) suggesting the variation is circadian-regulated. Gravitropic responses were disrupted in plants with disrupted circadian clocks, including plants overexpressing Circadian-clock Associated 1 (CCA1) and elf3-2, in the reorientation assay and on a 2D clinostat. These findings indicate that circadian-regulated pathways modulate the gravitropic responses, thus, highlighting the importance of considering and recording the time of day gravitropic experiments are performed.}, journal={Gravitational and Space Research}, author={Tolsma, Joseph S. and Ryan, Kaetlyn T. and Torres, Jacob J. and Richards, Jeffrey T. and Richardson, Zach and Land, Eric S. and Perera, Imara Y. and Doherty, Colleen J}, year={2021}, month={Jan} }
 @article{sheppard_land_toennisson_doherty_perera_2021, title={Uncovering Transcriptional Responses to Fractional Gravity in Arabidopsis Roots}, volume={11}, ISSN={["2075-1729"]}, url={https://doi.org/10.3390/life11101010}, DOI={10.3390/life11101010}, abstractNote={Although many reports characterize the transcriptional response of Arabidopsis seedlings to microgravity, few investigate the effect of partial or fractional gravity on gene expression. Understanding plant responses to fractional gravity is relevant for plant growth on lunar and Martian surfaces. The plant signaling flight experiment utilized the European Modular Cultivation System (EMCS) onboard the International Space Station (ISS). The EMCS consisted of two rotors within a controlled chamber allowing for two experimental conditions, microgravity (stationary rotor) and simulated gravity in space. Seedlings were grown for 5 days under continuous light in seed cassettes. The arrangement of the seed cassettes within each experimental container results in a gradient of fractional g (in the spinning rotor). To investigate whether gene expression patterns are sensitive to fractional g, we carried out transcriptional profiling of root samples exposed to microgravity or partial g (ranging from 0.53 to 0.88 g). Data were analyzed using DESeq2 with fractional g as a continuous variable in the design model in order to query gene expression across the gravity continuum. We identified a subset of genes whose expression correlates with changes in fractional g. Interestingly, the most responsive genes include those encoding transcription factors, defense, and cell wall-related proteins and heat shock proteins.}, number={10}, journal={LIFE-BASEL}, author={Sheppard, James and Land, Eric S. and Toennisson, Tiffany Aurora and Doherty, Colleen J. and Perera, Imara Y.}, year={2021}, month={Oct} }
 @article{williams_gillaspy_perera_2015, title={Biosynthesis and possible functions of inositol pyrophosphates in plants}, volume={6}, ISSN={["1664-462X"]}, DOI={10.3389/fpls.2015.00067}, abstractNote={Inositol phosphates (InsPs) are intricately tied to lipid signaling, as at least one portion of the inositol phosphate signaling pool is derived from hydrolysis of the lipid precursor, phosphatidyl inositol (4,5) bisphosphate. The focus of this review is on the inositol pyrophosphates, which are a novel group of InsP signaling molecules containing diphosphate or triphosphate chains (i.e., PPx) attached to the inositol ring. These PPx-InsPs are emerging as critical players in the integration of cellular metabolism and stress signaling in non-plant eukaryotes. Most eukaryotes synthesize the precursor molecule, myo-inositol (1,2,3,4,5,6)-hexakisphosphate (InsP6), which can serve as a signaling molecule or as storage compound of inositol, phosphorus, and minerals (referred to as phytic acid). Even though plants produce huge amounts of precursor InsP6 in seeds, almost no attention has been paid to whether PPx-InsPs exist in plants, and if so, what roles these molecules play. Recent work has delineated that Arabidopsis has two genes capable of PP-InsP5 synthesis, and PPx-InsPs have been detected across the plant kingdom. This review will detail the known roles of PPx-InsPs in yeast and animal systems, and provide a description of recent data on the synthesis and accumulation of these novel molecules in plants, and potential roles in signaling.}, journal={FRONTIERS IN PLANT SCIENCE}, author={Williams, Sarah P. and Gillaspy, Glenda E. and Perera, Imara Y.}, year={2015}, month={Feb} }
 @article{phillippy_perera_donahue_gillaspy_2015, title={Certain Malvaceae Plants Have a Unique Accumulation of  myo-Inositol 1,2,4,5,6-Pentakisphosphate}, volume={4}, ISSN={2223-7747}, url={http://dx.doi.org/10.3390/plants4020267}, DOI={10.3390/plants4020267}, abstractNote={Methods used to quantify inositol phosphates in seeds lack the sensitivity and specificity necessary to accurately detect the lower concentrations of these compounds contained in the leaves of many plants. In order to measure inositol hexakisphosphate (InsP6) and inositol pentakisphosphate (InsP5) levels in leaves of different plants, a method was developed to concentrate and pre-purify these compounds prior to analysis. Inositol phosphates were extracted from leaves with diluted HCl and concentrated on small anion exchange columns. Reversed-phase solid phase extraction cartridges were used to remove compounds that give peaks that sometimes interfere during HPLC. The method permitted the determination of InsP6 and InsP5 concentrations in leaves as low as 10 µM and 2 µM, respectively. Most plants analyzed contained a high ratio of InsP6 to InsP5. In contrast, certain members of the Malvaceae family, such as cotton (Gossypium) and some hibiscus (Hibiscus) species, had a preponderance of InsP5. Radiolabeling of cotton seedlings also showed increased amounts of InsP5 relative to InsP6. Why some Malvaceae species exhibit a reversal of the typical ratios of these inositol phosphates is an intriguing question for future research.}, number={2}, journal={Plants}, publisher={MDPI AG}, author={Phillippy, Brian and Perera, Imara and Donahue, Janet and Gillaspy, Glenda}, year={2015}, month={May}, pages={267–283} }
 @inbook{dalal_land_vasani_he_smith_rodriguez-welsh_perera_sederoff_2015, title={Methods for RNA Profiling of Gravi-Responding Plant Tissues}, volume={1309}, ISBN={9781493926961 9781493926978}, ISSN={1064-3745 1940-6029}, url={http://dx.doi.org/10.1007/978-1-4939-2697-8_9}, DOI={10.1007/978-1-4939-2697-8_9}, abstractNote={Plant transcriptional responses to gravity stimulation by reorientation are among the fastest measured in any tissue or species. Upon reorientation, changes in abundance of specific mRNAs can be measured within seconds or minutes, for plastid or nuclear encoded genes, respectively. Identifying fast gravity-induced transcripts has been made possible by the development of high-throughput technology for qualitative and quantitative RNA analysis. RNA profiling has undergone further rapid development due to its enormous potential in basic sciences and medical applications. We describe here the current and most widely used methods to profile the changes in an entire transcriptome by high-throughput sequencing of RNA fractions (RNAseq) and single gene transcript analysis using real-time quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR).}, booktitle={Methods in Molecular Biology}, publisher={Springer New York}, author={Dalal, Jyoti and Land, Eric and Vasani, Naresh and He, Luyan and Smith, Caroline and Rodriguez-Welsh, Maria and Perera, Imara Y. and Sederoff, Heike}, year={2015}, pages={91–117} }
 @article{ma_shatil-cohen_ben-dor_wigoda_perera_im_diminshtein_yu_boss_moshelion_et al._2014, title={Do phosphoinositides regulate membrane water permeability of tobacco protoplasts by enhancing the aquaporin pathway?}, volume={241}, ISSN={0032-0935 1432-2048}, url={http://dx.doi.org/10.1007/s00425-014-2216-x}, DOI={10.1007/s00425-014-2216-x}, abstractNote={Enhancing the membrane content of PtdInsP 2 , the already-recognized protein-regulating lipid, increased the osmotic water permeability of tobacco protoplasts, apparently by increasing the abundance of active aquaporins in their membranes. While phosphoinositides are implicated in cell volume changes and are known to regulate some ion channels, their modulation of aquaporins activity has not yet been reported for any organism. To examine this, we compared the osmotic water permeability (P f) of protoplasts isolated from tobacco (Nicotiana tabacum) cultured cells (NT1) with different (genetically lowered or elevated relative to controls) levels of inositol trisphosphate (InsP3) and phosphatidyl inositol [4,5] bisphosphate (PtdInsP2). To achieve this, the cells were transformed with, respectively, the human InsP3 5-phosphatase ('Ptase cells') or human phosphatidylinositol (4) phosphate 5-kinase ('PIPK cells'). The mean P f of the PIPK cells was several-fold higher relative to that of controls and Ptase cells. Three results favor aquaporins over the membrane matrix as underlying this excessive P f: (1) transient expression of the maize aquaporin ZmPIP2;4 in the PIPK cells increased P f by 12-30 μm s(-1), while in the controls only by 3-4 μm s(-1). (2) Cytosol acidification-known to inhibit aquaporins-lowered the P f in the PIPK cells down to control levels. (3) The transcript of at least one aquaporin was elevated in the PIPK cells. Together, the three results demonstrate the differences between the PIPK cells and their controls, and suggest a hitherto unobserved regulation of aquaporins by phosphoinositides, which could occur through direct interaction or indirect phosphoinositides-dependent cellular effects.}, number={3}, journal={Planta}, publisher={Springer Science and Business Media LLC}, author={Ma, Xiaohong and Shatil-Cohen, Arava and Ben-Dor, Shifra and Wigoda, Noa and Perera, Imara Y. and Im, Yang Ju and Diminshtein, Sofia and Yu, Ling and Boss, Wendy F. and Moshelion, Menachem and et al.}, year={2014}, month={Dec}, pages={741–755} }
 @article{hung_aspesi jr_hunter_lomax_perera_2014, title={Phosphoinositide-signaling is one component of a robust plant defense response}, volume={5}, ISSN={1664-462X}, url={http://dx.doi.org/10.3389/fpls.2014.00267}, DOI={10.3389/fpls.2014.00267}, abstractNote={The phosphoinositide pathway and inositol-1,4,5-triphosphate (InsP3) have been implicated in plant responses to many abiotic stresses; however, their role in response to biotic stress is not well characterized. In the current study, we show that both basal defense and systemic acquired resistance responses are affected in transgenic plants constitutively expressing the human type I inositol polyphosphate 5-phosphatase (InsP 5-ptase) which have greatly reduced InsP3 levels. Flagellin induced Ca2+-release as well as the expressions of some flg22 responsive genes were attenuated in the InsP 5-ptase plants. Furthermore, the InsP 5-ptase plants were more susceptible to virulent and avirulent strains of Pseudomonas syringae pv. tomato (Pst) DC3000. The InsP 5-ptase plants had lower basal salicylic acid (SA) levels and the induction of SAR in systemic leaves was reduced and delayed. Reciprocal exudate experiments showed that although the InsP 5-ptase plants produced equally effective molecules that could trigger PR-1 gene expression in wild type plants, exudates collected from either wild type or InsP 5-ptase plants triggered less PR-1 gene expression in InsP 5-ptase plants. Additionally, expression profiles indicated that several defense genes including PR-1, PR-2, PR-5, and AIG1 were basally down regulated in the InsP 5-ptase plants compared with wild type. Upon pathogen attack, expression of these genes was either not induced or showed delayed induction in systemic leaves. Our study shows that phosphoinositide signaling is one component of the plant defense network and is involved in both basal and systemic responses. The dampening of InsP3-mediated signaling affects Ca2+ release, modulates defense gene expression and compromises plant defense responses.}, journal={Frontiers in Plant Science}, publisher={Frontiers Media SA}, author={Hung, Chiu-Yueh and Aspesi Jr, Peter and Hunter, Melissa R. and Lomax, Aaron W. and Perera, Imara Y.}, year={2014}, month={Jun} }
 @article{desai_rangarajan_donahue_williams_land_mandal_phillippy_perera_raboy_gillaspy_2014, title={Two inositol hexakisphosphate kinases drive inositol pyrophosphate synthesis in plants}, volume={80}, ISSN={0960-7412}, url={http://dx.doi.org/10.1111/tpj.12669}, DOI={10.1111/tpj.12669}, abstractNote={SummaryInositol pyrophosphates are unique cellular signaling molecules with recently discovered roles in energy sensing and metabolism. Studies in eukaryotes have revealed that these compounds have a rapid turnover, and thus only small amounts accumulate. Inositol pyrophosphates have not been the subject of investigation in plants even though seeds produce large amounts of their precursor, myo‐inositol hexakisphosphate (InsP6). Here, we report that Arabidopsis and maize InsP6 transporter mutants have elevated levels of inositol pyrophosphates in their seed, providing unequivocal identification of their presence in plant tissues. We also show that plant seeds store a little over 1% of their inositol phosphate pool as InsP7 and InsP8. Many tissues, including, seed, seedlings, roots and leaves accumulate InsP7 and InsP8, thus synthesis is not confined to tissues with high InsP6. We have identified two highly similar Arabidopsis genes, AtVip1 and AtVip2, which are orthologous to the yeast and mammalian VIP kinases. Both AtVip1 and AtVip2 encode proteins capable of restoring InsP7 synthesis in yeast mutants, thus AtVip1 and AtVip2 can function as bonafide InsP6 kinases. AtVip1 and AtVip2 are differentially expressed in plant tissues, suggesting non‐redundant or non‐overlapping functions in plants. These results contribute to our knowledge of inositol phosphate metabolism and will lay a foundation for understanding the role of InsP7 and InsP8 in plants.}, number={4}, journal={The Plant Journal}, publisher={Wiley}, author={Desai, Mintu and Rangarajan, Padma and Donahue, Janet L. and Williams, Sarah P. and Land, Eric S. and Mandal, Mihir K. and Phillippy, Brian Q. and Perera, Imara Y. and Raboy, Victor and Gillaspy, Glenda E.}, year={2014}, month={Oct}, pages={642–653} }
 @inbook{heilmann_perera_2013, title={Measurement of Inositol (1,4,5) Trisphosphate in Plant Tissues by a Competitive Receptor Binding Assay}, ISBN={9781627034005 9781627034012}, ISSN={1064-3745 1940-6029}, url={http://dx.doi.org/10.1007/978-1-62703-401-2_4}, DOI={10.1007/978-1-62703-401-2_4}, abstractNote={The phosphoinositide signaling pathway is important for plant responses to many different stresses. As part of the responses to a stimulus, InsP3 levels may increase rapidly and transiently. The receptor binding assay for InsP3 described here is easy to use and an ideal method to monitor and compare InsP3 levels in multiple samples from large scale experiments. The method is based on competitive binding of InsP3 to the mammalian brain InsP3 specific receptor protein. This chapter describes a protocol for extracting and neutralizing plant samples and performing the receptor binding assay (using a commercially available kit). The protocol described has been used effectively to monitor InsP3 levels in plant tissues of different origin and in response to different stresses.}, booktitle={Methods in Molecular Biology}, publisher={Humana Press}, author={Heilmann, Ingo and Perera, Imara Y.}, year={2013}, pages={33–41} }
 @article{ischebeck_werner_krishnamoorthy_lerche_meijón_stenzel_löfke_wiessner_im_perera_et al._2013, title={Phosphatidylinositol 4,5-Bisphosphate Influences PIN Polarization by Controlling Clathrin-Mediated Membrane Trafficking in Arabidopsis}, volume={25}, ISSN={1040-4651 1532-298X}, url={http://dx.doi.org/10.1105/tpc.113.116582}, DOI={10.1105/tpc.113.116582}, abstractNote={AbstractThe functions of the minor phospholipid phosphatidylinositol-4,5-bisphosphate [PtdIns(4,5)P2] during vegetative plant growth remain obscure. Here, we targeted two related phosphatidylinositol 4-phosphate 5-kinases (PI4P 5-kinases) PIP5K1 and PIP5K2, which are expressed ubiquitously in Arabidopsis thaliana. A pip5k1 pip5k2 double mutant with reduced PtdIns(4,5)P2 levels showed dwarf stature and phenotypes suggesting defects in auxin distribution. The roots of the pip5k1 pip5k2 double mutant had normal auxin levels but reduced auxin transport and altered distribution. Fluorescence-tagged auxin efflux carriers PIN-FORMED (PIN1)–green fluorescent protein (GFP) and PIN2-GFP displayed abnormal, partially apolar distribution. Furthermore, fewer brefeldin A–induced endosomal bodies decorated by PIN1-GFP or PIN2-GFP formed in pip5k1 pip5k2 mutants. Inducible overexpressor lines for PIP5K1 or PIP5K2 also exhibited phenotypes indicating misregulation of auxin-dependent processes, and immunolocalization showed reduced membrane association of PIN1 and PIN2. PIN cycling and polarization require clathrin-mediated endocytosis and labeled clathrin light chain also displayed altered localization patterns in the pip5k1 pip5k2 double mutant, consistent with a role for PtdIns(4,5)P2 in the regulation of clathrin-mediated endocytosis. Further biochemical tests on subcellular fractions enriched for clathrin-coated vesicles (CCVs) indicated that pip5k1 and pip5k2 mutants have reduced CCV-associated PI4P 5-kinase activity. Together, the data indicate an important role for PtdIns(4,5)P2 in the control of clathrin dynamics and in auxin distribution in Arabidopsis.}, number={12}, journal={The Plant Cell}, publisher={American Society of Plant Biologists (ASPB)}, author={Ischebeck, Till and Werner, Stephanie and Krishnamoorthy, Praveen and Lerche, Jennifer and Meijón, Mónica and Stenzel, Irene and Löfke, Christian and Wiessner, Theresa and Im, Yang Ju and Perera, Imara Y. and et al.}, year={2013}, month={Dec}, pages={4894–4911} }
 @inbook{im_brglez_dieck_perera_boss_2013, title={Phosphatidylinositol 4-Kinase and Phosphatidylinositol 4-Phosphate 5-Kinase Assays}, ISBN={9781627034005 9781627034012}, ISSN={1064-3745 1940-6029}, url={http://dx.doi.org/10.1007/978-1-62703-401-2_15}, DOI={10.1007/978-1-62703-401-2_15}, abstractNote={Inositol lipid kinases are perhaps the easiest and most straightforward enzymes in the phosphoinositide pathway to analyze. In addition to monitoring lipid kinase-specific activity, lipid kinase assays can be used to quantify the inositol lipids present in isolated membranes (Jones et al., Methods Mol Biol 462:75–88, 2009). The lipid kinase assays are based on the fact that the more negatively charged phosphorylated lipid products are readily separated from their lipid substrates by thin layer chromatography. We have summarized our current protocols and identified important considerations for working with inositol lipids including different methods for substrate delivery when using recombinant proteins.}, booktitle={Methods in Molecular Biology}, publisher={Humana Press}, author={Im, Yang Ju and Brglez, Irena and Dieck, Catherine and Perera, Imara Y. and Boss, Wendy F.}, year={2013}, pages={163–174} }
 @article{smith_desai_land_perera_2013, title={A ROLE FOR LIPID-MEDIATED SIGNALING IN PLANT GRAVITROPISM}, volume={100}, ISSN={["0002-9122"]}, DOI={10.3732/ajb.1200355}, abstractNote={Gravitropism is a universal plant response. It is initiated by the sensing of the primary signal (mass or pressure), which is then converted into chemical signals that are transduced and propagated in a precise spatial and temporal fashion, resulting in a differential growth response. Our thesis is that membrane lipids and lipid‐mediated signaling pathways play critical roles in the initial signaling and in the establishment of polarity. In this review, we highlight results from recent literature and discuss the major questions that remain unanswered.}, number={1}, journal={AMERICAN JOURNAL OF BOTANY}, author={Smith, Caroline M. and Desai, Mintu and Land, Eric S. and Perera, Imara Y.}, year={2013}, month={Jan}, pages={153–160} }
 @article{dieck_boss_perera_2012, title={A Role for Phosphoinositides in Regulating Plant Nuclear Functions}, volume={3}, ISSN={1664-462X}, url={http://dx.doi.org/10.3389/fpls.2012.00050}, DOI={10.3389/fpls.2012.00050}, abstractNote={Nuclear localized inositol phospholipids and inositol phosphates are important for regulating many essential processes in animal and yeast cells such as DNA replication, recombination, RNA processing, mRNA export and cell cycle progression. An overview of the current literature indicates the presence of a plant nuclear phosphoinositide (PI) pathway. Inositol phospholipids, inositol phosphates, and enzymes of the PI pathway have been identified in plant nuclei and are implicated in DNA replication, chromatin remodeling, stress responses and hormone signaling. In this review, the potential functions of the nuclear PI pathway in plants are discussed within the context of the animal and yeast literature. It is anticipated that future research will help shed light on the functional significance of the nuclear PI pathway in plants.}, journal={Frontiers in Plant Science}, publisher={Frontiers Media SA}, author={Dieck, Catherine B. and Boss, Wendy F. and Perera, Imara Y.}, year={2012} }
 @article{boss_sederoff_im_moran_grunden_perera_2010, title={Basal Signaling Regulates Plant Growth and Development}, volume={154}, ISSN={["0032-0889"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-77957739073&partnerID=MN8TOARS}, DOI={10.1104/pp.110.161232}, abstractNote={The term signal transduction refers to the classical paradigm where an external stimulus is sensed and initiates an increase in second messengers. Each second messenger transmits and amplifies the signal by activating a subset of downstream pathways. This complex network of interwoven downstream}, number={2}, journal={PLANT PHYSIOLOGY}, author={Boss, Wendy F. and Sederoff, Heike Winter and Im, Yang Ju and Moran, Nava and Grunden, Amy M. and Perera, Imara Y.}, year={2010}, month={Oct}, pages={439–443} }
 @article{salinas mondragon_kajla_perera_brown_sederoff_2010, title={Role of inositol 1,4,5-triphosphate signalling in gravitropic and phototropic gene expression}, volume={33}, ISSN={0140-7791}, url={http://dx.doi.org/10.1111/j.1365-3040.2010.02204.x}, DOI={10.1111/j.1365-3040.2010.02204.x}, abstractNote={ABSTRACTPlants sense light and gravity to orient their direction of growth. One common component in the early events of both phototropic and gravitropic signal transduction is activation of phospholipase C (PLC), which leads to an increase in inositol 1,4,5‐triphosphate (InsP3) levels. The InsP3 signal is terminated by hydrolysis of InsP3 through inositolpolyphosphate‐5‐phosphatases (InsP 5‐ptases). Arabidopsis plants expressing a heterologous InsP 5‐ptase have low basal InsP3 levels and exhibit reduced gravitropic and phototropic bending. Downstream effects of InsP3‐mediated signalling are not understood. We used comparative transcript profiling to characterize gene expression changes in gravity‐ or light‐stimulated Arabidopsis root apices that were manipulated in their InsP3 metabolism either through inhibition of PLC activity or expression of InsP 5‐ptase. We identified InsP3‐dependent and InsP3‐independent co‐regulated gene sets in response to gravity or light stimulation. Inhibition of PLC activity in wild‐type plants caused similar changes in transcript abundance in response to gravitropic and phototropic stimulation as in the transgenic lines. Therefore, we conclude that changes in gene expression in response to gravitropic and phototropic stimulation are mediated by two signal transduction pathways that vary in their dependence on changes in InsP3.}, number={12}, journal={Plant, Cell & Environment}, publisher={Wiley}, author={Salinas Mondragon, Raul E. and Kajla, Jyoti D. and Perera, Imara Y. and Brown, Christopher S. and Sederoff, Heike Winter}, year={2010}, month={Nov}, pages={2041–2055} }
 @inbook{im_heilmann_perera_2010, title={The Hull of Fame: Lipid Signaling in the Plasma Membrane}, ISBN={9783642134302 9783642134319}, ISSN={1861-1370 1861-1362}, url={http://dx.doi.org/10.1007/978-3-642-13431-9_20}, DOI={10.1007/978-3-642-13431-9_20}, abstractNote={As the outermost barrier to the apoplast, the plasma membrane is critical for sensing and propagating signals that arise at the cell surface. Plasma membrane lipids are important mediators of signaling and the phosphoinositides (PIs), and sphingolipids are the major classes of lipids implicated in plant signaling. In this chapter, we will summarize the major findings on plant PI signaling and on the emerging signaling role of sphingolipids.}, booktitle={The Plant Plasma Membrane}, publisher={Springer Berlin Heidelberg}, author={Im, Yang Ju and Heilmann, Ingo and Perera, Imara Y.}, year={2010}, month={Sep}, pages={437–455} }
 @article{khodakovskaya_sword_wu_perera_boss_brown_sederoff_2010, title={Increasing inositol (1,4,5)-trisphosphate metabolism affects drought tolerance, carbohydrate metabolism and phosphate-sensitive biomass increases in tomato}, volume={8}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-73949085190&partnerID=MN8TOARS}, DOI={10.1111/j.1467-7652.2009.00472.x}, abstractNote={SummaryInositol‐(1,4,5)‐trisphosphate (InsP3) is a second messenger in plants that increases in response to many stimuli. The metabolic consequences of this signalling pathway are not known. We reduced the basal level of InsP3 in tomato (Solanum lycopersicum cv. Micro‐Tom) by expressing the human type I inositol polyphosphate 5‐phosphatase (InsP 5‐ptase) gene. Transgenic lines producing InsP 5‐ptase protein had between 15% and 30% of the basal InsP3 level of control plants. This increased hydrolysis of InsP3 caused dramatic increases in drought tolerance, vegetative biomass and lycopene and hexose concentrations in the fruits. Transcript profiling of root, leaf and fruit tissues identified a small group of genes, including a cell‐wall invertase inhibitor gene, that were differentially regulated in all tissues of the InsP 5‐ptase expressing plants. Significant differences were found in the amounts of carbohydrates and organic phosphate in these plants. Plants with increased hydrolysis of InsP3 in the cytosol also showed increased net CO2‐fixation and sucrose export into sink tissue and storage of hexoses in the source leaves. The increase in biomass was dependent on the supply of inorganic phosphate in the nutrient medium. Uptake and storage of phosphate was increased in the transgene expressing lines. This suggests that in tomato, increased flux through the inositol phosphate pathway uncoupled phosphate sensing from phosphate metabolism. Altering the second messenger, InsP3, revealed multiple coordinated changes in development and metabolism in tomato that have potential for crop improvement.}, number={2}, journal={Plant Biotechnology Journal}, author={Khodakovskaya, M. and Sword, C. and Wu, Q. and Perera, Imara and Boss, W. F. and Brown, C. S. and Sederoff, Heike}, year={2010}, pages={170–183} }
 @inbook{im_phillippy_perera_2009, title={InsP3 in Plant Cells}, ISBN={9783642038723 9783642038730}, ISSN={1861-1370 1861-1362}, url={http://dx.doi.org/10.1007/978-3-642-03873-0_10}, DOI={10.1007/978-3-642-03873-0_10}, abstractNote={D-myo-Inositol 1,4,5-trisphosphate (InsP3) is an important second messenger in eukaryotic cells. Although the phosphoinositide (PI) pathway has been well studied in plants, there is much that is not understood about PI-mediated signaling and there are fundamental differences between the plant and animal models. Many researchers have shown that plants produce InsP3 in response to multiple stimuli and that InsP3-mediated Ca2+ release is a component of plant signaling, although the candidate intracellular target of InsP3 in plants remains elusive. As plants are sessile organisms with multiple back-up systems, the InsP3-mediated signaling pathway may be one of the many signaling pathways in plants and its role may be more significant in specialized cells. This chapter provides an overview of InsP3 metabolism in plants, the current methods of analysis, and a review of the role of InsP3 in plants gathered from recent studies using mutants or transgenic plants with altered PI metabolism.}, booktitle={Lipid Signaling in Plants}, publisher={Springer Berlin Heidelberg}, author={Im, Yang Ju and Phillippy, Brian Q. and Perera, Imara Y.}, year={2009}, month={Oct}, pages={145–160} }
 @article{ma_shor_diminshtein_yu_im_perera_lomax_boss_moran_2009, title={Phosphatidylinositol (4,5)Bisphosphate Inhibits K+-Efflux Channel Activity in NT1 Tobacco Cultured Cells}, volume={149}, ISSN={["1532-2548"]}, DOI={10.1104/pp.108.129007}, abstractNote={Abstract
               In the animal world, the regulation of ion channels by phosphoinositides (PIs) has been investigated extensively, demonstrating a wide range of channels controlled by phosphatidylinositol (4,5)bisphosphate (PtdInsP2). To understand PI regulation of plant ion channels, we examined the in planta effect of PtdInsP2 on the K+-efflux channel of tobacco (Nicotiana tabacum), NtORK (outward-rectifying K channel). We applied a patch clamp in the whole-cell configuration (with fixed “cytosolic” Ca2+ concentration and pH) to protoplasts isolated from cultured tobacco cells with genetically manipulated plasma membrane levels of PtdInsP2 and cellular inositol (1,4,5)trisphosphate: “Low PIs” had depressed levels of these PIs, and “High PIs” had elevated levels relative to controls. In all of these cells, K channel activity, reflected in the net, steady-state outward K+ currents (IK), was inversely related to the plasma membrane PtdInsP2 level. Consistent with this, short-term manipulations decreasing PtdInsP2 levels in the High PIs, such as pretreatment with the phytohormone abscisic acid (25 μ  m) or neutralizing the bath solution from pH 5.6 to pH 7, increased IK (i.e. NtORK activity). Moreover, increasing PtdInsP2 levels in controls or in abscisic acid-treated high-PI cells, using the specific PI-phospholipase C inhibitor U73122 (2.5–4 μ  m), decreased NtORK activity. In all cases, IK decreases stemmed largely from decreased maximum attainable NtORK channel conductance and partly from shifted voltage dependence of channel gating to more positive potentials, making it more difficult to activate the channels. These results are consistent with NtORK inhibition by the negatively charged PtdInsP2 in the internal plasma membrane leaflet. Such effects are likely to underlie PI signaling in intact plant cells.}, number={2}, journal={PLANT PHYSIOLOGY}, author={Ma, Xiaohong and Shor, Oded and Diminshtein, Sofia and Yu, Ling and Im, Yang Ju and Perera, Imara and Lomax, Aaron and Boss, Wendy F. and Moran, Nava}, year={2009}, month={Feb}, pages={1127–1140} }
 @misc{perera_hung_moore_stevenson-paulik_boss_2008, title={Transgenic Arabidopsis Plants Expressing the Type 1 Inositol 5-Phosphatase Exhibit Increased Drought Tolerance and Altered Abscisic Acid Signaling}, volume={20}, ISSN={["1040-4651"]}, DOI={10.1105/tpc.108.061374}, abstractNote={AbstractThe phosphoinositide pathway and inositol-1,4,5-trisphosphate (InsP3) are implicated in plant responses to stress. To determine the downstream consequences of altered InsP3-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 Ca2+ responses to a cold or salt stimulus were reduced by ∼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 InsP3 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 InsP3 signal reveals unanticipated interconnections between signaling pathways.}, number={10}, journal={PLANT CELL}, author={Perera, Imara Y. and Hung, Chiu-Yueh and Moore, Candace D. and Stevenson-Paulik, Jill and Boss, Wendy F.}, year={2008}, month={Oct}, pages={2876–2893} }
 @article{im_perera_brglez_davis_stevenson-paulik_phillippy_johannes_allen_boss_2007, title={Increasing plasma membrane phosphatidylinositol(4,5)bisphosphate biosynthesis increases phosphoinositide metabolism in Nicotiana tabacum}, volume={19}, ISSN={["1532-298X"]}, DOI={10.1105/tpc.107.051367}, abstractNote={AbstractA genetic approach was used to increase phosphatidylinositol(4,5)bisphosphate [PtdIns(4,5)P2] biosynthesis and test the hypothesis that PtdInsP kinase (PIPK) is flux limiting in the plant phosphoinositide (PI) pathway. Expressing human PIPKIα in tobacco (Nicotiana tabacum) cells increased plasma membrane PtdIns(4,5)P2 100-fold. In vivo studies revealed that the rate of 32Pi incorporation into whole-cell PtdIns(4,5)P2 increased >12-fold, and the ratio of [3H]PtdInsP2 to [3H]PtdInsP increased 6-fold, but PtdInsP levels did not decrease, indicating that PtdInsP biosynthesis was not limiting. Both [3H]inositol trisphosphate and [3H]inositol hexakisphosphate increased 3-and 1.5-fold, respectively, in the transgenic lines after 18 h of labeling. The inositol(1,4,5)trisphosphate [Ins(1,4,5)P3] binding assay showed that total cellular Ins(1,4,5)P3/g fresh weight was >40-fold higher in transgenic tobacco lines; however, even with this high steady state level of Ins(1,4,5)P3, the pathway was not saturated. Stimulating transgenic cells with hyperosmotic stress led to another 2-fold increase, suggesting that the transgenic cells were in a constant state of PI stimulation. Furthermore, expressing Hs PIPKIα increased sugar use and oxygen uptake. Our results demonstrate that PIPK is flux limiting and that this high rate of PI metabolism increased the energy demands in these cells.}, number={5}, journal={PLANT CELL}, author={Im, Yang Ju and Perera, Imara Y. and Brglez, Irena and Davis, Amanda J. and Stevenson-Paulik, Jill and Phillippy, Brian Q. and Johannes, Eva and Allen, Nina S. and Boss, Wendy F.}, year={2007}, month={May}, pages={1603–1616} }
 @inproceedings{howard_sick_perera_im_winter-sederoff_heber_2007, title={Quality Assessment of Affymetrix GeneChip Data using the EM Algorithm and a Naive Bayes Classifier}, ISBN={1424415098 9781424415090}, url={http://dx.doi.org/10.1109/bibe.2007.4375557}, DOI={10.1109/bibe.2007.4375557}, abstractNote={Recent research has demonstrated the utility of using supervised classification systems for automatic identification of low quality microarray data. However, this approach requires annotation of a large training set by a qualified expert. In this paper we demonstrate the utility of an unsupervised classification technique based on the Expectation-Maximization (EM) algorithm and naive Bayes classification. On our test set, this system exhibits performance comparable to that of an analogous supervised learner constructed from the same training data.}, booktitle={2007 IEEE 7th International Symposium on BioInformatics and BioEngineering}, publisher={IEEE}, author={Howard, Brian E. and Sick, Beate and Perera, Imara and Im, Yang Ju and Winter-Sederoff, Heike and Heber, Steffen}, year={2007}, month={Oct}, pages={145–150} }
 @inbook{boss_davis_im_galvão_perera_2006, place={Boston, MA}, series={Subcellular Biochemistry}, title={Phosphoinositide Metabolism: Towards an Understanding of Subcellular Signaling}, ISBN={9780387275994}, url={http://dx.doi.org/10.1007/0-387-27600-9_8}, DOI={10.1007/0-387-27600-9_8}, booktitle={Biology of Inositols and Phosphoinositides}, publisher={Springer US}, author={Boss, Wendy F. and Davis, Amanda J. and Im, Yang Ju and Galvão, Rafaelo M. and Perera, ImaraY.}, editor={Majumber, A.L. and Biswas, B.B.Editors}, year={2006}, month={Sep}, pages={181–205}, collection={Subcellular Biochemistry} }
 @article{im_davis_perera_johannes_allen_boss_2007, title={The N-terminal membrane occupation and recognition nexus domain of Arabidopsis phosphatidylinositol phosphate kinase 1 regulates enzyme activity}, volume={282}, ISSN={["1083-351X"]}, DOI={10.1074/jbc.M611342200}, abstractNote={The type I B family of phosphatidylinositol phosphate kinases (PIPKs) contain a characteristic region of Membrane Occupation and Recognition Nexus (MORN) motifs at the N terminus. These MORN motifs are not found in PIPKs from other eukaryotes. To understand the impact of the additional N-terminal domain on protein function and subcellular distribution, we expressed truncated and full-length versions of AtPIPK1, one member of this family of PIPKs, in Escherichia coli and in tobacco cells grown in suspension culture. Deletion of the N-terminal MORN domain (amino acids 1–251) of AtPIPK1 increased the specific activity of the remaining C-terminal peptide (ΔMORN) >4-fold and eliminated activation by phosphatidic acid (PtdOH). PtdOH activation could also be eliminated by mutating Pro396 to Ala (P396A) in the predicted linker region between the MORN and the kinase homology domains. AtPIPK1 is product-activated and the MORN domain binds PtdIns(4,5)P2. Adding back the MORN peptide to ΔMORN or to the PtdOH-activated full-length protein increased activity ∼2-fold. Furthermore, expressing the MORN domain in vivo increased the plasma membrane PtdInsP kinase activity. When cells were exposed to hyperosmotic stress, the MORN peptide redistributed from the plasma membrane to a lower phase or endomembrane fraction. In addition, endogenous PtdInsP kinase activity increased in the endomembrane fraction of hyperosmotically stressed cells. We conclude that the MORN peptide can regulate both the function and distribution of the enzyme in a manner that is sensitive to the lipid environment.}, number={8}, journal={JOURNAL OF BIOLOGICAL CHEMISTRY}, author={Im, Yang Ju and Davis, Amanda J. and Perera, Imara Y. and Johannes, Eva and Allen, Nina S. and Boss, Wendy F.}, year={2007}, month={Feb}, pages={5443–5452} }
 @article{perera_hung_brady_muday_boss_2006, title={A universal role for inositol 1,4,5-trisphosphate-mediated signaling in plant gravitropism}, volume={140}, ISSN={["1532-2548"]}, DOI={10.1104/pp.105.075119}, abstractNote={Abstract
               Inositol 1,4,5-trisphosphate (InsP3) has been implicated in the early signaling events of plants linking gravity sensing to the initiation of the gravitropic response. However, at present, the contribution of the phosphoinositide signaling pathway in plant gravitropism is not well understood. To delineate the role of InsP3 in plant gravitropism, we generated Arabidopsis (Arabidopsis thaliana) plants constitutively expressing the human type I inositol polyphosphate 5-phosphatase (InsP 5-ptase), an enzyme that specifically hydrolyzes InsP3. The transgenic plants show no significant differences in growth and life cycle compared to wild-type plants, although basal InsP3 levels are reduced by greater than 90% compared to wild-type plants. With gravistimulation, InsP3 levels in inflorescence stems of transgenic plants show no detectable change, whereas in wild-type plant inflorescences, InsP3 levels increase approximately 3-fold within the first 5 to 15 min of gravistimulation, preceding visible bending. Furthermore, gravitropic bending of the roots, hypocotyls, and inflorescence stems of the InsP 5-ptase transgenic plants is reduced by approximately 30% compared with the wild type. Additionally, the cold memory response of the transgenic plants is attenuated, indicating that InsP3 contributes to gravisignaling in the cold. The transgenic roots were shown to have altered calcium sensitivity in controlling gravitropic response, a reduction in basipetal indole-3-acetic acid transport, and a delay in the asymmetric auxin-induced β-glucuronidase expression with gravistimulation as compared to the controls. The compromised gravitropic response in all the major axes of growth in the transgenic Arabidopsis plants reveals a universal role for InsP3 in the gravity signal transduction cascade of plants.}, number={2}, journal={PLANT PHYSIOLOGY}, author={Perera, IY and Hung, CY and Brady, S and Muday, GK and Boss, WF}, year={2006}, month={Feb}, pages={746–760} }
 @article{perera_davis_galanopilou_im_boss_2005, title={Characterization and comparative analysis of Arabidopsis phosphatidylinositol phosphate 5-kinase 10 reveals differences in Arabidopsis and human phosphatidylinositol phosphate kinases}, volume={579}, ISSN={["1873-3468"]}, DOI={10.1016/j.febslet.2005.05.018}, abstractNote={
Arabidopsis phosphatidylinositol phosphate (PtdInsP) kinase 10 (AtPIPK10; At4g01190) is shown to be a functional enzyme of the subfamily A, type I AtPtdInsP kinases. It is biochemically distinct from AtPIPK1 (At1g21980), the only other previously characterized AtPtdInsP kinase which is of the B subfamily. AtPIPK10 has the same K
m, but a 10‐fold lower V
max than AtPIPK1 and it is insensitive to phosphatidic acid. AtPIPK10 transcript is most abundant in inflorescence stalks and flowers, whereas AtPIPK1 transcript is present in all tissues. Comparative analysis of recombinant AtPIPK10 and AtPIPK1 with recombinant HsPIPKIα reveals that the Arabidopsis enzymes have roughly 200‐ and 20‐fold lower V
max/K
m, respectively. These data reveal one explanation for the longstanding mystery of the relatively low phosphatidylinositol‐(4,5)‐bisphosphate:phosphatidylinositol‐4‐phosphate ratio in terrestrial plants.}, number={16}, journal={FEBS LETTERS}, author={Perera, IY and Davis, AJ and Galanopilou, D and Im, YJ and Boss, WF}, year={2005}, month={Jun}, pages={3427–3432} }
 @article{davis_perera_boss_2004, title={Cyclodextrins enhance recombinant phosphatidylinositol phosphate kinase activity}, volume={45}, ISSN={["1539-7262"]}, DOI={10.1194/jlr.D400005-JLR200}, abstractNote={Inositol lipid kinases have been studied extensively in both plant and animal systems. However, major limitations for in vitro studies of recombinant lipid kinases are the low specific activity and instability of the purified proteins. Our goal was to determine if cyclodextrins would provide an effective substrate delivery system and enhance the specific activity of lipid kinases. For these studies, we have used recombinant Arabidopsis thaliana phosphatidylinositol phosphate kinase 1 (At PIPK1). At PIPK1 was produced as a fusion protein with glutathione-S-transferase and purified on glutathione-Sepharose beads. A comparison of lipid kinase activity using substrate prepared in α-, β-, or γ-cyclodextrin indicated that β-cyclodextrin was most effective and enhanced lipid kinase activity 6-fold compared with substrate prepared in Triton X-100-mixed micelles. We have optimized reaction conditions and shown that product can be recovered from the cyclodextrin-treated recombinant protein, which reveals a potential method for automating the assay for pharmacological screening.}, number={9}, journal={JOURNAL OF LIPID RESEARCH}, author={Davis, AJ and Perera, IY and Boss, WF}, year={2004}, month={Sep}, pages={1783–1789} }
 @article{padmanaban_lin_perera_kawamura_sze_2004, title={Differential Expression of Vacuolar H+-ATPase Subunit c Genes in Tissues Active in Membrane Trafficking and Their Roles in Plant Growth as Revealed by RNAi}, volume={134}, ISSN={0032-0889 1532-2548}, url={http://dx.doi.org/10.1104/pp.103.034025}, DOI={10.1104/pp.103.034025}, abstractNote={Abstract
               Acidification of intracellular compartments by the vacuolar-type H+-ATPases (VHA) is known to energize ion and metabolite transport, though cellular processes influenced by this activity are poorly understood. At least 26 VHA genes encode 12 subunits of the V1Vo-ATPase complex in Arabidopsis, and how the expression, assembly, and activity of the pump are integrated into signaling networks that govern growth and adaptation are largely unknown. The role of multiple VHA-c genes encoding the 16-kD subunit of the membrane Vo sector was investigated. Expression of VHA-c1, monitored by promoter-driven β-glucuronidase (GUS) activity was responsive to light or dark in an organ-specific manner. VHA-c1 expression in expanding cotyledons, hypocotyls of etiolated seedlings, and elongation zone of roots supported a role for V-ATPase in cell enlargement. Mutants reduced in VHA-c1 transcript using dsRNA-mediated interference showed reduction in root growth relative to wild-type seedlings. In contrast, VHA-c3 promoter::GUS expression was undetectable in most organs of seedlings, but strong in the root cap. Interestingly, dsRNA-mediated mutants of vha-c3 also showed reduced root length and decreased tolerance to moderate salt stress. The results suggest that V-ATPase functions in the root cap influenced root growth. Expression of VHA-c1 and VHA-c3 in tissues with active membrane flow, including root cap, vascular strands, and floral style would support a model for participation of the Vo sector and V1Vo-ATPase in membrane trafficking and fusion. Two VHA-c genes are thus differentially expressed to support growth in expanding cells and to supply increased demand for V-ATPase in cells with active exocytosis.}, number={4}, journal={Plant Physiology}, publisher={American Society of Plant Biologists (ASPB)}, author={Padmanaban, Senthilkumar and Lin, Xiaoying and Perera, Imara and Kawamura, Yukio and Sze, Heven}, year={2004}, month={Mar}, pages={1514–1526} }
 @inbook{heilmann_perera_boss_2002, title={An Evolutionary Perspective of Phosphoinositide Signaling During Osmotic Stress}, booktitle={Intracellular Signalling in Plant and Animal Systems}, author={Heilmann, I. and Perera, I.Y. and Boss, W.F.}, editor={Kravets, V.Editor}, year={2002} }
 @article{perera_love_heilmann_thompson_boss_2002, title={Up-regulation of phosphoinositide metabolism in tobacco cells constitutively expressing the human type I inositol polyphosphate 5-phosphatase}, volume={129}, ISSN={["1532-2548"]}, DOI={10.1104/pp.003426}, abstractNote={Abstract
               To evaluate the impact of suppressing inositol 1,4,5-trisphosphate (InsP3) in plants, tobacco (Nicotiana tabacum) cells were transformed with the human type I inositol polyphosphate 5-phosphatase (InsP 5-ptase), an enzyme which specifically hydrolyzes InsP3. The transgenic cell lines showed a 12- to 25-fold increase in InsP 5-ptase activity in vitro and a 60% to 80% reduction in basal InsP3 compared with wild-type cells. Stimulation with Mas-7, a synthetic analog of the wasp venom peptide mastoparan, resulted in an approximately 2-fold increase in InsP3 in both wild-type and transgenic cells. However, even with stimulation, InsP3 levels in the transgenic cells did not reach wild-type basal values, suggesting that InsP3 signaling is compromised. Analysis of whole-cell lipids indicated that phosphatidylinositol 4,5-bisphosphate (PtdInsP2), the lipid precursor of InsP3, was greatly reduced in the transgenic cells. In vitro assays of enzymes involved in PtdInsP2 metabolism showed that the activity of the PtdInsP2-hydrolyzing enzyme phospholipase C was not significantly altered in the transgenic cells. In contrast, the activity of the plasma membrane PtdInsP 5 kinase was increased by approximately 3-fold in the transgenic cells. In vivo labeling studies revealed a greater incorporation of 32P into PtdInsP2 in the transgenic cells compared with the wild type, indicating that the rate of PtdInsP2 synthesis was increased. These studies show that the constitutive expression of the human type I InsP 5-ptase in tobacco cells leads to an up-regulation of the phosphoinositide pathway and highlight the importance of PtdInsP2 synthesis as a regulatory step in this system.}, number={4}, journal={PLANT PHYSIOLOGY}, author={Perera, IY and Love, J and Heilmann, I and Thompson, WF and Boss, WF}, year={2002}, month={Aug}, pages={1795–1806} }
 @article{perera_hilmann_chang_boss_kaufman_2001, title={A role for inositol 1,4,5-trisphosphate in gravitropic signaling and the retention of cold-perceived gravistimulation of oat shoot pulvini}, volume={125}, ISSN={["1532-2548"]}, DOI={10.1104/pp.125.3.1499}, abstractNote={Abstract
               Plants sense positional changes relative to the gravity vector. To date, the signaling processes by which the perception of a gravistimulus is linked to the initiation of differential growth are poorly defined. We have investigated the role of inositol 1,4,5-trisphosphate (InsP3) in the gravitropic response of oat (Avena sativa) shoot pulvini. Within 15 s of gravistimulation, InsP3 levels increased 3-fold over vertical controls in upper and lower pulvinus halves and fluctuated in both pulvinus halves over the first minutes. Between 10 and 30 min of gravistimulation, InsP3 levels in the lower pulvinus half increased 3-fold over the upper. Changes in InsP3 were confined to the pulvinus and were not detected in internodal tissue, highlighting the importance of the pulvinus for both graviperception and response. Inhibition of phospholipase C blocked the long-term increase in InsP3, and reduced gravitropic bending by 65%. Short-term changes in InsP3 were unimpaired by the inhibitor. Gravitropic bending of oat plants is inhibited at 4°C; however, the plants retain the information of a positional change and respond at room temperature. Both short- and long-term changes in InsP3 were present at 4°C. We propose a role for InsP3 in the establishment of tissue polarity during the gravitropic response of oat pulvini. InsP3 may be involved in the retention of cold-perceived gravistimulation by providing positional information in the pulvini prior to the redistribution of auxin.}, number={3}, journal={PLANT PHYSIOLOGY}, author={Perera, IK and Hilmann, I and Chang, SC and Boss, WF and Kaufman, PB}, year={2001}, month={Mar}, pages={1499–1507} }
 @inproceedings{boss_perera_love_heilmann_2001, title={Altering phosphoinositide metabolism by expressing human type I inositol polyphosphate 5 ' phosphatase in tobacco cells}, volume={12}, number={2001 Nov}, booktitle={Molecular Biology of the Cell}, author={Boss, W. F. and Perera, I. Y. and Love, J. and Heilmann, I.}, year={2001}, pages={820} }
 @article{heilmann_perera_gross_boss_2001, title={Plasma membrane phosphatidylinositol 4,5-bisphosphate levels decrease with time in culture}, volume={126}, ISSN={["1532-2548"]}, DOI={10.1104/pp.126.4.1507}, abstractNote={Abstract
               During the stationary phase of growth, after 7 to 12 d in culture, the levels of phosphatidylinositol 4,5-bisphosphate (PtdInsP2) decreased by 75% in plasma membranes of the red alga Galdieria sulphuraria. Concomitant with the decrease in PtdInsP2 levels in plasma membranes, there was an increase in PtdInsP2 in microsomes, suggesting that the levels of plasma membrane PtdInsP2 are regulated differentially. The decline of PtdInsP2 in plasma membranes was accompanied by a 70% decrease in the specific activity of PtdInsP kinase and by reduced levels of protein cross-reacting with antisera against a conserved PtdInsP kinase domain. Upon osmotic stimulation, the loss of PtdInsP2from the plasma membrane increased from 10% in 7-d-old cells to 60% in 12-d-old cells, although the levels of inositol 1,4,5-trisphosphate (InsP3) produced in whole cells were roughly equal at both times. When cells with low plasma membrane PtdInsP2 levels were osmotically stimulated, a mild osmotic stress (12.5 mm KCl) activated PtdInsP kinase prior to InsP3 production, whereas in cells with high plasma membrane PtdInsP2, more severe stress (250 mm KCl) was required to induce an increase in PtdInsP kinase activity. The differential regulation of a plasma membrane signaling pool of PtdInsP2 is discussed with regard to the implications for understanding the responsive state of cells.}, number={4}, journal={PLANT PHYSIOLOGY}, author={Heilmann, I and Perera, IY and Gross, W and Boss, WF}, year={2001}, month={Aug}, pages={1507–1518} }
 @article{heilmann_shin_huang_perera_davies_2001, title={Transient dissociation of polyribosomes and concurrent recruitment of calreticulin and calmodulin transcripts in gravistimulated maize pulvini}, volume={127}, ISSN={["1532-2548"]}, DOI={10.1104/pp.127.3.1193}, abstractNote={In plants, sugars are the main respiratory substrates and important signaling molecules in the regulation of carbon metabolism. Sugar signaling studies suggested that sugar sensing involves several key components, among them hexokinase (HXK). Although the sensing mechanism of HXK is unknown, several experiments support the hypothesis that hexose phosphorylation is a determining factor. Glucose (Glc) analogs transported into cells but not phosphorylated are frequently used to test this hypothesis, among them 3-O-methyl-Glc (3-OMG). The aim of the present work was to investigate the effects and fate of 3-OMG in heterotrophic plant cells. Measurements of respiration rates, protein and metabolite contents, and protease activities and amounts showed that 3-OMG is not a respiratory substrate and does not contribute to biosynthesis. Proteolysis and lipolysis are induced in 3-OMG-fed maize (Zea mays L. cv DEA) roots in the same way as in sugar-starved organs. However, contrary to the generally accepted idea, phosphorous and carbon nuclear magnetic resonance experiments and enzymatic assays prove that 3-OMG is phosphorylated to 3-OMG-6-phosphate, which accumulates in the cells. Insofar as plant HXK is involved in sugar sensing, these findings are discussed on the basis of the kinetic properties because the catalytic efficiency of HXK isolated from maize root tips is five orders of magnitude lower for 3-OMG than for Glc and Man.}, number={3}, journal={PLANT PHYSIOLOGY}, author={Heilmann, I and Shin, J and Huang, J and Perera, IY and Davies, E}, year={2001}, month={Nov}, pages={1193–1203} }
 @article{stevenson_perera_heilmann_persson_boss_2000, title={Inositol signaling and plant growth}, volume={5}, ISSN={1360-1385}, url={http://dx.doi.org/10.1016/s1360-1385(00)01652-6}, DOI={10.1016/s1360-1385(00)01652-6}, abstractNote={Living organisms have evolved to contain a wide variety of receptors and signaling pathways that are essential for their survival in a changing environment. Of these, the phosphoinositide pathway is one of the best conserved. The ability of the phosphoinositides to permeate both hydrophobic and hydrophilic environments, and their diverse functions within cells have contributed to their persistence in nature. In eukaryotes, phosphoinositides are essential metabolites as well as labile messengers that regulate cellular physiology while traveling within and between cells. The stereospecificity of the six hydroxyls on the inositol ring provides the basis for the functional diversity of the phosphorylated isomers that, in turn, generate a selective means of intracellular and intercellular communication for coordinating cell growth. Although such complexity presents a difficult challenge for bench scientists, it is ideal for the regulation of cellular functions in living organisms.}, number={6}, journal={Trends in Plant Science}, publisher={Elsevier BV}, author={Stevenson, Jill M and Perera, Imara Y and Heilmann, Ingo and Persson, Staffan and Boss, Wendy F}, year={2000}, month={Jun}, pages={252–258} }
 @article{heilmann_perera_gross_boss_1999, title={Changes in phosphoinositide metabolism with days in culture affect signal transduction pathways in Galdieria sulphuraria}, volume={119}, ISSN={["0032-0889"]}, DOI={10.1104/pp.119.4.1331}, abstractNote={Abstract
               The metabolism of phosphatidylinositol-4,5-bisphosphate (PIP2) changed during the culture period of the thermoacidophilic red alga Galdieria sulphuraria. Seven days after inoculation, the amount of PIP2 in the cells was 910 ± 100 pmol g−1fresh weight; by 12 d, PIP2 levels increased to 1200 ± 150 pmol g−1 fresh weight. In vitro assays indicated that phosphatidylinositol monophosphate (PIP) kinase specific activity increased from 75 to 230 pmol min−1mg−1 protein between d 7 and 12. When G. sulphuraria cells were osmostimulated, transient increases of up to 4-fold could be observed in inositol-1,4,5-trisphosphate (IP3) levels within 90 s, regardless of the age of the cells. In d-12 cells, the increase in IP3 was preceded by a transient increase of up to 5-fold in specific PIP kinase activity, whereas no such increase was detected after osmostimulation of d-7 cells. The increase in PIP kinase activity before IP3signaling in d-12 cells indicates that there is an additional pathway for regulation of phosphoinositide metabolism after stimulation other than an initial activation of phospholipase C. Also, the rapid activation of PIP2 biosynthesis in cells with already-high PIP2 levels suggests that the PIP2 present was not available for signal transduction. By comparing the response of the cells at d 7 and 12, we have identified two potentially distinct pools of PIP2.}, number={4}, journal={PLANT PHYSIOLOGY}, author={Heilmann, I and Perera, IY and Gross, W and Boss, WF}, year={1999}, month={Apr}, pages={1331–1339} }
 @article{perera_heilmann_boss_1999, title={Transient and sustained increases in inositol 1,4,5-hisphosphate precede the differential growth response in gravistimulated maize pulvini}, volume={96}, ISSN={["1091-6490"]}, DOI={10.1073/pnas.96.10.5838}, abstractNote={
            The internodal maize pulvinus responds to gravistimulation with differential cell elongation on the lower side. As the site of both graviperception and response, the pulvinus is an ideal system to study how organisms sense changes in orientation. We observed a transient 5-fold increase in inositol 1,4,5-trisphosphate (IP
            3
            ) within 10 s of gravistimulation in the lower half of the pulvinus, indicating that the positional change was sensed immediately. Over the first 30 min, rapid IP
            3
            fluctuations were observed between the upper and lower halves. Maize plants require a presentation time of between 2 and 4 h before the cells on the lower side of the pulvinus are committed to elongation. After 2 h of gravistimulation, the lower half consistently had higher IP
            3,
            and IP
            3
            levels on the lower side continued to increase up to ≈5-fold over basal levels before visible growth. As bending became visible after 8–10 h, IP
            3
            levels returned to basal values. Additionally, phosphatidylinositol 4-phosphate 5-kinase activity in the lower pulvinus half increased transiently within 10 min of gravistimulation, suggesting that the increased IP
            3
            production was accompanied by an up-regulation of phosphatidylinositol 4,5-bisphosphate biosynthesis. Neither IP
            3
            levels nor phosphatidylinositol 4-phosphate 5-kinase activity changed in pulvini halves from vertical control plants. Our data indicate the involvement of IP
            3
            and inositol phospholipids in both short- and long-term responses to gravistimulation. As a diffusible second messenger, IP
            3
            provides a mechanism to transmit and amplify the signal from the perceiving to the responding cells in the pulvinus, coordinating a synchronized growth response.
          }, number={10}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, author={Perera, IY and Heilmann, I and Boss, WF}, year={1999}, month={May}, pages={5838–5843} }
 @article{stevenson_perera_boss_1998, title={A phosphatidylinositol 4-kinase pleckstrin homology domain that binds phosphatidylinositol 4-monophosphate}, volume={273}, ISSN={["0021-9258"]}, DOI={10.1074/jbc.273.35.22761}, abstractNote={Pleckstrin homology (PH) domains are found in many proteins involved in signal transduction, including the family of large molecular mass phosphatidylinositol (PI) 4-kinases. Although the exact function of these newly discovered domains is unknown, it is recognized that they may influence enzyme regulation by binding different ligands. In this study, the recombinant PI 4-kinase PH domain was explored for its ability to bind to different phospholipids. First, we isolated partial cDNAs of the >7-kilobase transcripts of PI 4-kinases from carrot (DcPI4Kα) andArabidopsis (AtPI4Kα). The deduced primary sequences were 41% identical and 68% similar to rat and human PI 4-kinases and contained the telltale lipid kinase unique domain, PH domain, and catalytic domain. Antibodies raised against the expressed lipid kinase unique, PH, and catalytic domains identified a polypeptide of 205 kDa in Arabidopsis microsomes and an F-actin-enriched fraction from carrot cells. The 205-kDa immunoaffinity-purified Arabidopsis protein had PI 4-kinase activity. We have used the expressed PH domain to characterize lipid binding properties. The recombinant PH domain selectively bound to phosphatidylinositol 4-monophosphate (PI-4-P), phosphatidylinositol 4,5-bisphosphate (PI-4,5-P2), and phosphatidic acid and did not bind to the 3-phosphoinositides. The PH domain had the highest affinity for PI-4-P, the product of the reaction. Consideration is given to the potential impact that this has on cytoskeletal organization and the PI signaling pathway in cells that have a high PI-4-P/PI-4,5-P2 ratio.}, number={35}, journal={JOURNAL OF BIOLOGICAL CHEMISTRY}, author={Stevenson, JM and Perera, IY and Boss, WF}, year={1998}, month={Aug}, pages={22761–22767} }
 @inbook{heilmann_perera_stevenson_ransom_gross_boss_1998, title={Inositol lipid signaling: what can we learn from plants?}, booktitle={Advances in lipids research}, publisher={Sevilla, Spain: University of Sevilla Press}, author={Heilmann, I. and Perera, I. Y. and Stevenson, J. M. and Ransom, W. D. and Gross, W. and Boss, W. F.}, editor={J. Sanchez, E. Cerda-Olmedo and Martinez-Force, E.Editors}, year={1998}, pages={394–397} }
 @inbook{perera_boss_1996, place={Rockville, MD}, series={Current Topics in Plant Physiology}, title={Signal transduction pathways: a plant perspective of inositol metabolism and growth}, booktitle={Regulation of plant growth and development by light development}, publisher={American Society of Plant Physiologists}, author={Perera, I.Y. and Boss, W.F.}, editor={Briggs, Winslow R. and Heath, Robert L. and Tobin, ElaineM.Editors}, year={1996}, pages={114–126}, collection={Current Topics in Plant Physiology} }
 @article{perera_li_sze_1995, title={Several distinct genes encode nearly identical 16 kDa proteolipids of the vacuolar H+-ATPase from Arabidopsis thaliana}, volume={29}, ISSN={0167-4412 1573-5028}, url={http://dx.doi.org/10.1007/bf00043648}, DOI={10.1007/bf00043648}, abstractNote={To understand the subcellular roles and the regulation of vacuolar H(+)-ATPases, we have begun to identify the genes encoding the major subunits and to determine their patterns of expression in Arabidopsis thaliana. Two distinct cDNAs (AVA-P1 and AVA-P2) and one genomic sequence (AVA-P3) encoding the 16 kDa subunit have been isolated. The 16 kDa proteolipid is a major component of the membrane integral sector that forms the proton conductance pathway and is required for assembly of the V-ATPase complex. Interestingly, the open reading frame of one full-length cDNA (AVA-P1) and a genomic sequence (AVA-P3) encoded an identical polypeptide of 164 amino acids with a molecular mass of 16,570. The deduced amino acid sequences of the two cDNAs were nearly identical (99%) and hydropathy plots suggested a molecule with four membrane-spanning domains characteristic of V-ATPase proteolipids. The three genes differed mainly in their codon usage and in their 3'-untranslated regions. The coding region of the genomic sequence, AVA-P3, was interrupted by two introns located at the codons for Cys-26 and Arg-121. The presence of additional 16 kDa proteolipid genes was suggested from several polymerase chain reaction (PCR)-amplified fragments that differed from one another in the size of the second intron. PCR 1 had an intron of ca. 800 bp and its identity as AVA-P4, a fourth member of the gene family, was confirmed from sequence analyses of an EST cDNA. The mRNAs of three genes (AVA-P1, AVA-P2 and AVA-P3) were detected in Arabidopsis leaf, root, flower and silique; yet expression of AVA-P1 and AVA-P2 was lower in roots. All three genes were expressed in light- or dark-grown seedlings; however mRNA levels of AVA-P2 were enhanced in etiolated plants. Arabidopsis thaliana, therefore, has at least four distinct genes encoding nearly identical 16 kDa proteolipids, and the enhanced expression of AVA-P2 transcript in etiolated seedlings suggests that an increase in V-ATPase could accompany cell expansion.}, number={2}, journal={Plant Molecular Biology}, publisher={Springer Science and Business Media LLC}, author={Perera, Imara Y. and Li, Xuhang and Sze, Heven}, year={1995}, month={Oct}, pages={227–244} }
 @article{gawienowski_szymanski_perera_zielinski_1993, title={Calmodulin isoforms in Arabidopsis encoded by multiple divergent mRNAs}, volume={22}, ISSN={0167-4412 1573-5028}, url={http://dx.doi.org/10.1007/bf00014930}, DOI={10.1007/bf00014930}, abstractNote={Three new, unique cDNA sequences encoding isoforms of calmodulin (CaM) were isolated from an Arabidopsis cDNA library cloned in lambda gt10. These sequences (ACaM-4, -5, and -6) represent members of the Arabidopsis CaM gene family distinct from the three DNA sequences previously reported. ACaM-4 and -6 encode full-length copies of CaM mRNAs of ca. 0.75 kb. The ACaM-5 sequence encodes a partial length copy of CaM mRNA that is lacking sequences encoding the amino-terminal 10 amino acids of mature CaM and the initiator methionine. The derived amino acid sequence of ACaM-5 is identical to the sequences encoded by two of the previously characterized ACaM cDNAs, and is identical to TCH-1 mRNA, whose accumulation was increased by touch stimulation. The polypeptides encoded by ACaM-4 and -6 differ from that encoded by ACaM-5 by six and two amino acid substitutions, respectively. Most of the deduced amino acid sequence substitutions in the Arabidopsis CaM isoforms occurred in the fourth Ca(2+)-binding domain. Polymerase chain reaction amplification assays of ACaM-4, -5 and -6 mRNA sequences indicated that each accumulated in Arabidopsis leaf RNA fractions, but only ACaM-4 and -5 mRNAs were detected in silique total RNA. The six different CaM cDNA sequences each hybridize with unique EcoRI restriction fragments in genomic Southern blots of Arabidopsis DNA, indicating that these sequences were derived from distinct structural genes. Our results suggest that CaM isoforms in Arabidopsis may have evolved to optimize the interaction of this Ca(2+)-receptor protein with specific subsets of response elements.}, number={2}, journal={Plant Molecular Biology}, publisher={Springer Science and Business Media LLC}, author={Gawienowski, Margaret C. and Szymanski, Daniel and Perera, Imara Y. and Zielinski, Raymond E.}, year={1993}, month={May}, pages={215–225} }
 @article{perera_zielinski_1992, title={Structure and expression of the Arabidopsis CaM-3 calmodulin gene}, volume={19}, ISSN={0167-4412 1573-5028}, url={http://dx.doi.org/10.1007/bf00026791}, DOI={10.1007/bf00026791}, abstractNote={Genomic and cDNA sequences encoding a calmodulin (CaM) gene from Arabidopsis (ACaM-3) have been isolated and characterized. ACaM-3 represents a sequence distinct from two previously isolated Arabidopsis CaM cDNA clones. A 2.3 kb Eco RI restriction fragment was sequenced and found to encode a complete CaM-coding sequence interrupted by a single 491 bp intron, together with 750 bp and 600 bp of 5' and 3' flanking sequences, respectively. The polypeptide encoded by ACaM-3 is identical to that encoded by ACaM-2 and it differs from the one encoded by ACaM-1 by four of 148 residues. The putative promoter of ACaM-3 was atypical of CaM genes previously isolated from animals in that it contained consensus TATA and CAAT box sequences and lacked GC-rich regions. Two DNA sequence elements closely resembling cyclic AMP regulatory elements, which have been identified in animal CaM genes, were located in the 5' flanking region of ACaM-3. Northern blot and polymerase chain reaction amplification assays confirmed that each of the three ACaM mRNAs were expressed in similar but distinct patterns in different organs. ACaM-1 mRNA was the only species detectable in root RNA fractions, and ACaM-3 mRNA could not be detected in floral stalks. Accumulation of the three CaM mRNAs in leaves was induced by a touch stimulus, but the kinetics and extent of the induction varied among the three mRNA species. Run-on transcription assays indicated that a portion of the differences in accumulation of ACaM-1, 2, and 3 mRNAs in leaves and siliques was attributable to differences in their net rates of transcription.}, number={4}, journal={Plant Molecular Biology}, publisher={Springer Science and Business Media LLC}, author={Perera, Imara Y. and Zielinski, Raymond E.}, year={1992}, month={Jul}, pages={649–664} }
 @article{perera_zielinski_1992, title={Synthesis and Accumulation of Calmodulin in Suspension Cultures of Carrot (Daucus carota L.)}, volume={100}, DOI={10.1104/pp.100.2.812}, abstractNote={The expression of calmodulin mRNA and protein were measured during a growth cycle of carrot (Daucus carota L.) cells grown in suspension culture. A full-length carrot calmodulin cDNA clone isolated from a lambdagt10 library was used to measure steady-state calmodulin mRNA levels. During the exponential phase of culture growth when mitotic activity and oxidative respiration rates were maximal, calmodulin mRNA levels were 4- to 5-fold higher than they were during the later stages of culture growth, when respiration rates were lower and growth was primarily by cell expansion. Net calmodulin polypeptide synthesis, as measured by pulse-labeling in vivo with [(35)S]methionine, paralleled the changes in calmodulin steady-state mRNA level during culture growth. As a consequence, net calmodulin polypeptide synthesis declined 5- to 10-fold during the later stages of culture growth. The qualitative spectrum of polypeptides synthesized and accumulated by the carrot cells during the course of a culture cycle, however, remained largely unchanged. Calmodulin polypeptide levels, in contrast to its net synthesis, remained relatively constant during the exponential phases of the culture growth cycle and increased during the later stages of culture growth. Our data are consistent with increased calmodulin polypeptide turnover associated with periods of rapid cell proliferation and high levels of respiration.}, number={2}, journal={Plant Physiology}, author={Perera, I. and Zielinski, R.}, year={1992}, pages={812–819} }
 @article{sze_ward_lai_perera_1992, title={Vacuolar-type H+-translocating ATPases in plant endomembranes: subunit organization and multigene families}, volume={172}, journal={Journal of Experimental Biology}, author={Sze, H. and Ward, J.M. and Lai, S. and Perera, I.}, year={1992}, pages={123–135} }
 @article{ling_perera_zielinski_1991, title={Primary Structures of Arabidopsis Calmodulin Isoforms Deduced from the Sequences of cDNA Clones}, volume={96}, ISSN={0032-0889 1532-2548}, url={http://dx.doi.org/10.1104/pp.96.4.1196}, DOI={10.1104/pp.96.4.1196}, abstractNote={Complementary DNA (cDNA) clones encoding calmodulin isoforms were isolated from an Arabidopsis leaf lambdagt10 library by screening with cloned barley calmodulin cDNA probes. Two cDNAs, one a 626-base pair partial-length clone (ACaM-1) and one a 1400-base pair full-length clone (ACaM-2), encode calmodulin polypeptides that differ by four conservative amino acid substitutions. None of the amino acid sequence differences occur within the four Ca(2+)-binding domains of the proteins. Whereas the deduced amino acid sequences of the two Arabidopsis calmodulin isoforms share 97% identity, the nucleotide sequences encoding the two isoforms share 87% sequence identity. Most of these nucleotide sequence differences (80%) occur in codon wobble positions. ACaM-1 and ACaM-2 both hybridize with a distinct set of restriction fragments of Arabidopsis total DNA, indicating that they were derived from transcripts of separate genes; these genes are single- or very low-copy in the Arabidopsis genome. Both cDNAs hybridize to messenger RNA (mRNA) species of 0.8 kilobases that are expressed to a greater extent in developing siliques compared with leaves, flowers, and stems. Northern blot and polymerase chain reaction assays both indicate that ACaM-1 mRNA is more highly expressed than ACaM-2 mRNA in developing siliques. The steady-state levels of both isoform mRNAs increase as a result of touch stimulation; the kinetics and extent of increase are comparable for the two mRNAs.}, number={4}, journal={Plant Physiology}, publisher={American Society of Plant Biologists (ASPB)}, author={Ling, Vincent and Perera, Imara and Zielinski, Raymond E.}, year={1991}, month={Aug}, pages={1196–1202} }