Mechanisms and Impacts of De-regulating Aromatic Amino Acid Biosynthesis in Plants

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Plants produce aromatic amino acids (AAAs) from simple precursors, and use them to produce proteins, numerous natural products, including alkaloids, phenolics and additional organic compounds that have medicinal, nutritional and other useful properties. Understanding and improving the biosynthesis of AAAs will enhance the nutritional quality of food, achieve cost-effective production of medicines and enable sustainable production of industrial materials. The aims of this research are to gain fundamental knowledge concerning how plants regulate the production of AAAs, in order to precisely control the production of plant natural products that are derived from AAAs. "Pigment Art+" outreach activities will be conducted with K-12 students and their parents as a means of increasing the awareness of the importance of plant natural products. Colorful plant pigments derived from AAAs are used in this project as tools to inform the general public about the nutritional and pharmacological values of plant natural products. The activities are targeted for parents from communities that are low-income and underrepresented in science. This project also trains students and postdocs of diverse backgrounds, in collaborative interdisciplinary research that integrates biochemistry, structural and quantitative biology and the chemistry of plants.Plants are particularly suited to produce AAAs and their derivatives since they naturally synthesize and accumulate a large quantity of AAA-derived compounds. How plants regulate AAA biosynthesis and coordinate the overall metabolic network to efficiently produce AAAs remains elusive. A recent project identified plant enzymes that de-regulate the entry and final reactions of AAA biosynthesis, catalyzed by 3-deoxy-D-arabinoheptulosonate 7-phosphate synthase (DHS) and arogenate dehydrogenase (TyrA), respectively. Such novel regulatory enzymes of AAA biosynthesis are used to modulate this key intermediary pathway that connects central carbon metabolism and numerous plant natural product metabolic pathways. The major goals of this proposal are to define molecular mechanisms underlying the de-regulation of plant AAA biosynthesis and to determine their quantitative impacts on overall plant metabolic networks. The project specifically investigates the feedback regulation of DHS and TyrA variants, and quantitatively test their in planta roles in controlling AAA precursor supply. Dynamic carbon-13 CO2 labeling will be used for metabolic flux analysis. Time-dependent carbon-13 enrichment data will be used to estimate fluxes based on optimization of fit to a metabolic network model. This integrated molecular to network level analyses will provide a series of de-regulated DHS and TyrA variants, which will be tested in planta. These experiments will generate prototype plant chemical platforms that will display enhanced AAA precursor supply.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.