This project will determine how nitric oxide (NO) modulates important aspects of plant growth, development and seed yield, processes critical to food security and plant biomass production. NO acts as a signaling molecule in all higher organisms, and the discoverers of the role of NO in cells were awarded the Nobel Prize. This project will use biochemical and genetic approaches in Arabidopsis thaliana to investigate how a specific protein enzyme, known as GSNOR, controls the amount and effects of NO in plants. Results will contribute to our overall understanding of the mechanism of NO action in all organisms. One postdoctoral researcher, a graduate student, two undergraduates and a middle school teacher will receive training in laboratory research, as well as mentoring in written and oral communication of scientific data for specialist and lay audiences. In addition, outreach programs will focus on engaging teachers and students in Amherst middle and high schools with the goals of developing hands on experiments for middle school students involving plant materials and exposing high school students to University of Massachusetts STEM undergraduate majors and their campus experiences. The engagement with Amherst schools will also serve the important goal of strengthening community ties with the university.
Despite the clear involvement of nitric oxide (NO) in multiple plant processes, including germination, root growth and fertility, a basic understanding of the mechanisms by which NO exerts its effects on systems critical for plant growth and development is lacking and will be addressed in this project. The cellular signaling and regulatory effects of NO are mediated by reversible, posttranslational protein redox modifications, including cysteine (Cys) nitrosation (Cys-SNO) and glutathionylation (Cys-SG). These reversible, covalent modifications result primarily from reaction with S-nitrosoglutathione (GSNO), a soluble NO-adduct of the abundant cellular redox buffer glutathione. They can have major effects on protein activity, indicating there must be mechanisms to control NO and GSNO concentrations in both time and space within cells. This project investigates how the enzyme S-nitrosoglutathione reductase (GSNOR) plays a major role in modulating GSNO and protein nitrosation levels. A model for regulation of GSNOR activity by nitrosation of specific Cys residues has been developed from biochemical studies of Arabidopsis, yeast and human GSNOR. This project will test this model both in vitro (Aim 1) and in vivo in Arabidopsis thaliana (Aim 2) using biochemistry and genetics, and possible regulation of GSNOR activity by other posttranslational modifications will be investigated. Experiments will also determine how GSNOR nitrosation is reversed by specific cytosolic redox proteins in the thioredoxin and thioredoxin reductase families (Aims 1 and 2). Results will develop a complete picture of the possible redox modifications controlling GSNOR activity and thereby cellular NO levels, and provide novel data on the activity and specificity of cytosolic thioredoxins. Finally, changes in protein nitrosation associated with the reduced fertility of Arabidopsis GSNOR null mutants will be determined by novel mass spectrometry methods (Aim 3). In total, experiments address how mechanisms controlling activity of a single enzyme (GSNOR) are propagated to result in phenotypic changes at the whole organism level.
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.
