RUI: Regulation Of Pyrimidine Metabolism In Plants

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Pyrimidine nucleotides are precursors in the biosynthesis of DNA and RNA, molecules which contain the essential genetic information for growth and development in plants and all other living organisms. UDP-sugars are also activated intermediates in the synthesis of certain types of lipids, as well as a major carbohydrate storage product (sucrose) and cell wall polysaccharides (cellulose, hemicellulose, pectins) in plants, accounting for much of the earth's biomass and production of food and fiber. Biochemical studies have helped to define major enzymes and pathways comprising pyrimidine metabolism, but plants remain the only major group of organisms for which basic mechanisms controlling pyrimidine metabolism have not been elucidated.A functional genomics approach will be used to study pyrimidine metabolism in the model plant, Arabidopsis. Using bioinformatics tools to "mine" information from integrated genomics databases, 146 genes encoding functionally characterized or putative enzymes in each of the major areas of pyrimidine metabolism (de novo synthesis of nucleotides, nucleotide interconversions, salvaging of preformed nucleobases/nucleosides to nucleotides, catabolic pathways) and related pathways (UDP-glucose and arginine synthesis) have been identified. The goals of this research are to 1) use DNA microarrays to characterize coordination of pathway activities, at the level of gene expression, and to identify potentially-regulated genes 2) to functionally characterize a small number of genes/enzymes in vitro (complementation of defined mutants, biochemical studies) or in planta (T-DNA knockouts, RNAi-mediated gene silencing, 35S promoter-driven overexpression in transgenic Arabidopsis lines) 3) to study expression of each gene in response to changes in pyrimidine availability in plant tissues (N-(phosphonacetyl)-L-aspartate (PALA) inhibition of de novo synthesis, exogenous pyrimidines, etc.) The proposed collaboration with The Institute for Genomic Research ensures that high-quality array data will be generated, while availability of full-length cDNAs/ESTs and Arabidopsis T-DNA knockout mutants for the enzymes to be studied will facilitate the proposed research.Broader impacts: These studies will significantly advance our understanding of the regulation of pyrimidine metabolism in plants and will provide important insights into coordination of these activities with other areas of plant metabolism. The project will provide training opportunities for undergraduate students in bioinformatics and plant functional genomics. Gene expression data from array experiments will also be used in future student investigator-driven projects in a bioinformatics course at Goucher College, expanding the educational impact of this research. Students will present their research at scientific meetings and will co-author publications. If the past trends continue, most of them will pursue post-baccalaureate training at top-ranked graduate or professional schools.