EAGER: Biochemical and Genetic Tools for Analysis of Paralogous Polymerase Subunits in Grasses

点击翻译按钮获取中文摘要

One of the most important questions in biology is how every cell in an organism can contain the same set of instructions (the DNA the comprises the genome) yet produce diverse cell types that respond differently to various environmental signals. This diversity of cells and responses occurs when different parts of the genome are transcribed by RNA Polymerases (Pols). Understanding the function of various Pols might help us engineer organisms in beneficial ways, and will therefore have impacts on medicine, agriculture, and environmental conservation. This project is focused on new Pol subunits discovered in the grass family, which includes key crops such as rice, wheat, and corn. This project will test the function of these novel Pol proteins and create genetic and biochemical tools for subsequent investigations. Additionally, this project will train a graduate student in cutting-edge biological approaches such as genome editing and analysis of high throughput sequencing data. This project also contains experiments suitable for undergraduate students to participate in research and these students will be recruited from groups that are underrepresented in STEM fields.Plants have evolved two specialized RNA polymerases, Pol IV and Pol V, which are responsible for transcriptional gene silencing of transposons and adjacent genes. Although they share a number of smaller subunits, these polymerases have divergent biological activities due to unique subunits that arose through gene duplication and functional divergence. Grasses have additional duplications of Pol IV and Pol V subunits. Phylogenetic analysis indicates that these duplicates have neofunctionalized with respect to both function and holoenzyme assembly. The goal of this project is to genetically and biochemically test whether the duplicate (paralogous) Pol subunits have diverged in function and evolved into a sixth RNA Polymerase in grasses. These questions will be answered through the creation and analysis of Pol subunit mutants and tagged transgenic lines in rice. Comparison of mutant phenotypes and assessment of subunit interactions will determine whether the paralogous subunits have distinct phenotypes and binding partners, or whether they function redundantly.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.