Recipient Organization
LOUISIANA STATE UNIVERSITY
202 HIMES HALL
BATON ROUGE,LA 70803-0100
Performing Department
Plant Pathology & Crop Physiol
Non Technical Summary
Photosynthesis is the process in which pants and some microorganisms convert solar energy and CO2 in to sugars to use for food. Photosynthesis requires the addition of carbon (carboxylation) to ribulose 1,5, bisphosphate (RuBP) by the enzyme Rubisco. Under current atmospheric conditions approximately 20% of the time Rubisco will attach an oxygen (oxygenation) to RuBP creating a compound called 2-phosphoglycolate (2-PG) that can inhibit photosynthesis. To recycle the 2-PG plants have evolved a complex multi-compartment process called photorespiration. Overall due to the energetic demand of photorespiration the process can reduce the efficiency of photosynthesis by 15-50% depending on plant growing temperatures, reducing yields. The enzymatic steps in photorespiration have been studied for the past several decades but to date only 3 of the potential 28 transport steps necessary for photorespiration to function are known at the genetic level. With photorespiration integral to plant primary metabolism a better understanding of the transport steps (Objective 1) in the pathway would provide insight into plant productivity and for food crops targets for improved metabolic efficiency. Photorespiration metabolism has been implicated in abiotic stress response but how gene expression is controlled by these signals has yet to be fully understood. In objective two, we will elucidate promoter response elements and epigenetic factors related to the regulation of photorespiration gene expression by identifying changes in photorespiration gene expression and epigenetic states under acute and chronic abiotic stress conditions.Plant growth throughout the growing season is highly regulated and is linked to rates of photosynthesis. What is still unknown is how photorespiration influences plant growth throughout the day (diurnally) and throughout the growing season. The dynamics of photosynthesis and photorespiration are usually non-linear meaning we need higher resolution data that can better integrate over these scales to understand how the process of photorespiration influences crop yields at the field, canopy, leaf and cellular level. In objective 3 we will measure diurnal and seasonal changes to photosynthesis, photorespiration, leaf sugar content, and changes in gene expression in crop species important to agriculture productivity in the southeast (i.e., Louisiana, Mississippi, Alabama, and Georgia) such as sweet potato and soybean. By having a better understanding to these underlying mechanisms in control of photorespiration it will be possible to improve crop productivity through manipulation reducing the energy cost of photorespiration.
Animal Health Component
(N/A)
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Goals / Objectives
The regulation of photorespiration response at the genetic and epigenetic level is poorly understood including acute and acclimated responses to changes in environmental conditions. There has been little investigation into the regulation of the expression of photorespiration genes. Co-expression analysis has been used to identify potential genes important for the photorespiration process, but the elements and transcriptional control of photorespiration gene expression need further study.With photorespiration integral to plant primary metabolism a better understanding of the transport steps in the pathway would provide insight into plant productivity and for food crops targets for improved metabolic efficiency. In objective one we will characterize additional putative membrane proteins we have identified as having photorespiration dependent phenotypes.Photorespiration metabolism has been implicated in abiotic and biotic stress response but how gene expression is controlled by these signals has yet to be fully understood. In objective two, we will elucidate promoter response elements and epigenetic factors related to the regulation of photorespiration gene expression by identifying changes in photorespiration gene expression and epigenetic states under acute and chronic abiotic stress conditions.The dynamics of photosynthesis and photorespiration are usually non-linear meaning we need higher resolution data that can better integrate over these scales to understand how the process of photorespiration influences crop yields at the field, canopy, leaf and cellular level. In objective three we will measure diurnal and seasonal changes to photosynthesis, photorespiration, leaf sugar content, and changes in gene expression in crop species important to agriculture productivity in the southeast (i.e. Louisiana, Mississippi, Alabama, and Georgia) such as sweet potato, and rice.
Project Methods
MethodsObjective 1: Identify and characterize transport proteins important in photorespiration flux.Genetic analysis of candidate chloroplast transporter protein mutants mfl1-1Growth and yield measurements of mfl1-1Infrared gas analysis measuring photosynthesis and photorespiration rates.Fluorescent microscopy identifying membrane localization of MFL1 protein.Phenotypic compensation for membrane transport in yeast.Objective 2: Characterize genetic response elements important for gene regulation in photorespiration.Bioinformatic analysis of photorespiration and photosynthesis related genes in tobacco, cowpea, soybean, and sweet potatoDNA synthesis and molecular cloning of gene promoter elements from bioinformatic analysisGene expression analysis of tobacco, soybean, cowpea, and sweet potato under short- and long-term heat stressChromatin immunoprecipitation to map epigenetic markers of target genes.Objective 3: Identify acute, acclimated, diurnal, and seasonal changes in photorespiration and related gene expression responses.Chlorophyll fluorescence and infrared gas analysis to measure photosynthesis and photorespiration in tobacco, cowpea, soybean, and sweet potato.Field experimentation measuring diurnal and seasonal photosynthetic rates in cowpea, soybean, and sweet potato.Field harvest of leaf material for sugar, metabolite, and gene expression analysis.Biological data sets will be in the form of written observations (recorded in digital notebooks) digital instrument readouts (gene expression, photosynthetic, chlorophyll fluorescence data), Images (gels, Western blots) and photographs of plants. Large datasets will also be generated from RNA and DNA sequencing data that will be first stored on core facility servers and transferred to LSU for storage.Statistical AnalysisType of statistical analysis will be dependent on the individual experiment. Generally, Gene expression analysis by RNA sequencing will be analyzed filtering the dataset and normalized using transformed trimmed Log2 mean values. Specific gene analysis will be determined by analysis of variance (ANOVA) with significance followed by a post hoc Tukey's test. Analysis of chlorophyll fluorescence parameters and infra-red gas analysis will be tested using ANOVA followed by a post hoc Tukey's test. Data generated for yield and nutritional quality of harvested fruit or grain will be analyzed by either a Student's T-test or ANOVA. All statistical analysis will be performed using statistical software programs such as R, Origin pro, or SAS. Consultation will be used through the department of experimental statistics at Louisiana State University for design and use of statistical models.Efforts to share scientific knowledge for described methods will include laboratory instruction in BIOL3060 Plant Physiology, Teaching graduate level course BIO7010 Plant Molecular Biology, training undergraduate and graduate students in methods to complete research tasks.Evaluation of each objective will include milestones where data is analyzed, and project commitments are amended or modified to meet project timelines. In addition, weekly lab meetings, quarterly research reports and annual evaluations of scientific progress will be held to determine objective progress.Key Milestones:Objective 1Obtain gene constructs from DNA synthesis to characterize transporter protein MFL1.Obtain data on growth analysis, photosynthetic rates, and protein localization for MFL1 protein.Obtain yeast strains and data from yeast complementation assays to determine MFL1 function.Analyze data, write and submit manuscript on the characterization of MFL1 protein.Objective 2:Obtain data from bioinformatic analysis of Tobacco, cowpea, soybean, and sweet potato genomes with lists of candidate photorespiration and photosynthetic genes with identified promoter elements.Obtain gene constructs for characterization of target plant promoter elements.Obtain data of gene expression analysis of candidate gene targets during abiotic stress treatments.Obtain data from chromatin immunoprecipitation experiments on candidate target genes with differential expression under abiotic stress for photorespiration genes.Analyze data, write and submit manuscript characterizing genetic regulation of photorespiration.Objective 3:Obtain infrared gas exchange analysis and chlorophyll fluorescence data on cowpea, sweet potato, soybean, and tobacco plants under greenhouse and growth chamber conditions.Design field experiments for cowpea, soybean, and sweet potato to measure photosynthesis and harvest leaf material for metabolic, gene expression and sugar analysis.Obtain data from field experiments measuring diurnal and seasonal changes in photosynthesis and gene expression.