Quantitative Analysis of Transcription Elongation Perturbation Networks

Indiana University, Bloomington IN

Investigators

Abstract

The main goal of this project is to identify how cells respond and adapt to genetic changes that alter the process of RNA synthesis. Using baker's yeast as the study system, experimental approaches will include state-of-the-art laboratory technologies and computational methods used to interpret the resulting quantitative data. As part of the project, a summer internship program will be established with DePauw University, an undergraduate institution, to give undergraduate students hands-on research experience at a research-intensive institution. These experiences will provide early exposure to cutting-edge research technologies that are broadly applicable in both academia and industry. The project will also expose students at all levels of training to the importance of quantitative methods to interrogate biology. The specific focus of the research will be to understand the consequences on RNA polymerase II-catalyzed RNA synthesis (transcription) of genetic perturbations that knock out or reduce expression of non-essential and essential transcription elongation factors. Preliminary data suggest that such genetic perturbations lead to significant adaptations within the RNA polymerase II protein-protein interaction network. These adaptations may include (but are not limited to) disruption of interactions with RNA polymerase II, compensation leading to novel protein recruitment, and activation of rescue pathways to dispose of arrested RNA polymerase II. Perturbation-induced changes will be measured using quantitative mass spectrometry, followed by functional genomics characterization by chromatin immunoprecipitation and DNA sequencing. The research will have two major outcomes: development of generalizable, quantitative approaches for detecting new protein-protein interactions that mediate cellular responses to genetic perturbations, and identification of novel functions for proteins that regulate transcriptional elongation by RNA polymerase II.