The control of gene expression through regulated transcription by RNA polymerase I (pol II) is fundamental to the cell viability and is often corrupted in disease. The transcription cycle comprises initiation, elongation and termination phases. This proposal focuses on the latter two steps because the mechanisms that govern them are less well elucidated than initiation. Our lab uses strategies that combine genetics to make mutations in pol II and elongation factors with biochemical assays of elongation and termination in vivo using budding yeast and mammalian cells. Termination is required to recycle the RNA polymerase and to prevent interference between adjacent genes but the mechanism that triggers disassembly of the extremely stable transcription elongation complex at the end of genes is quite incompletely understood. We will investigate this mechanism by asking how it is affected by pol II phosphorylation and transcription elongation rate. We will also look for new players in termination by using two approaches: 1) a candidate approach based on our identification of protein factors that interact with the termination factor Xrn2, an RNA exonuclease and 2) an unbiased genetic screen for shRNA that inhibit termination. We will also test a model we recently proposed for control of elongation by premature termination that we call the """"""""decap and torpedo"""""""" model. This model suggests that the flux of pol II from the pause site at the beginning of most human genes into the gene body is limited by decapping of the transcript and subsequent premature termination of transcription by Xrn2 and associated factors. We will test the details of this model and ask whether decapping and termination factors function in regulation of the elongation phase of the transcription cycle. We will focus on the potential role of decapping and termination factors in control of transcription by the activators HIV Tat, c-myc, heat shock factor and hypoxia inducible factors.

Public Health Relevance

Cancer and many inherited syndromes are diseases caused by the misexpression of genes which is often caused by corruption of the transcription step that makes disposable RNA copies of the genetic information. Our work will help clarify the mechanisms responsible for the elongation and termination of RNA transcripts. The importance of this topic is illustrated by our finding that a block to elongation which limits expression of te c-myc oncogene is lost in Burkitt's lymphoma cells where the gene is translocated and overexpressed.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM063873-09
Application #
8467297
Study Section
Special Emphasis Panel (ZRG1-GGG-T (02))
Program Officer
Sledjeski, Darren D
Project Start
2003-07-01
Project End
2017-02-28
Budget Start
2013-03-01
Budget End
2014-02-28
Support Year
9
Fiscal Year
2013
Total Cost
$308,894
Indirect Cost
$106,394
Name
University of Colorado Denver
Department
Biochemistry
Type
Schools of Medicine
DUNS #
041096314
City
Aurora
State
CO
Country
United States
Zip Code
80045
Sheridan, Ryan M; Bentley, David L (2016) Selectable one-step PCR-mediated integration of a degron for rapid depletion of endogenous human proteins. Biotechniques 60:69-74
Saldi, Tassa; Cortazar, Michael A; Sheridan, Ryan M et al. (2016) Coupling of RNA Polymerase II Transcription Elongation with Pre-mRNA Splicing. J Mol Biol 428:2623-35
Bentley, David L (2015) The union of transcription and mRNA processing: 20 years of coupling. RNA 21:569-70
Fong, Nova; Brannan, Kristopher; Erickson, Benjamin et al. (2015) Effects of Transcription Elongation Rate and Xrn2 Exonuclease Activity on RNA Polymerase II Termination Suggest Widespread Kinetic Competition. Mol Cell 60:256-67
Klein, Brianna J; Piao, Lianhua; Xi, Yuanxin et al. (2014) The histone-H3K4-specific demethylase KDM5B binds to its substrate and product through distinct PHD fingers. Cell Rep 6:325-35
Bentley, David L (2014) Coupling mRNA processing with transcription in time and space. Nat Rev Genet 15:163-75
Perales, Roberto; Erickson, Benjamin; Zhang, Lian et al. (2013) Gene promoters dictate histone occupancy within genes. EMBO J 32:2645-56
Brannan, Kris; Bentley, David L (2012) Control of Transcriptional Elongation by RNA Polymerase II: A Retrospective. Genet Res Int 2012:170173
Larochelle, Stéphane; Amat, Ramon; Glover-Cutter, Kira et al. (2012) Cyclin-dependent kinase control of the initiation-to-elongation switch of RNA polymerase II. Nat Struct Mol Biol 19:1108-15
Brannan, Kris; Kim, Hyunmin; Erickson, Benjamin et al. (2012) mRNA decapping factors and the exonuclease Xrn2 function in widespread premature termination of RNA polymerase II transcription. Mol Cell 46:311-24

Showing the most recent 10 out of 23 publications