Systematic analysis of context-specific functions of transcriptional CDKs in human cells

University Of Colorado At Denver, Aurora CO

Investigators

Abstract

This project investigates the process by which the genetic information stored in the DNA molecule is expressed in cells. The first part of this process, known as gene transcription, involves the copying of segments of DNA (genes) into RNA molecules. Despite the undisputed importance of proper control of gene transcription in biology, there are still major unanswered questions in this field. How do cells adjust their gene transcription in response to changes in their environment? How is gene transcription controlled during the development of multicellular organisms? What are the key enzymes and other proteins within the cell that regulate transcription in these different contexts? Within this framework, the research team will investigate a group of enzymes known as "transcriptional CDKs" that are well recognized regulators of gene transcription in organisms from yeast to humans. This research will provide new insights into how transcriptional CDKs enable cells to alter their transcriptional activity in different contexts. Therefore, the knowledge generated will advance the field of molecular biology and expand our collective understanding of how the genetic information within DNA is expressed to ultimately control cell behavior. The research will be carried out by a diverse teaching-research team with a proven track record of training scientists from undergraduate to post-doctoral levels, including promoting underrepresented minorities in academia, and increasing scientific literacy in society at large. This project will elucidate the functional specialization of the transcriptional CDKs using a chemical genetics approach. Using genome editing technology, researchers will create alleles of each kinase that can be inhibited by bulky ATP substrate analogs. This panel of matching isogenic human cell lines will then be used to: 1) decipher the contribution of each CDK to the transcription cycle via genome-wide measurements of RNA synthesis, processing, and steady-state levels; 2) identify unique and shared proteins targeted for phosphorylation by each CDK; and c) identify genetic interactors for each kinase using CRISPR-based genetic screening in human cells. Collectively, these efforts will provide an advanced mechanistic understanding of how transcriptional CDKs regulate gene transcription, as well as the evolutionary mechanisms leading to their functional specialization in vertebrate cells. 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.