The production of messenger RNA (mRNA) is the primary event in gene expression where genetic information is transferred from the gene's DNA into a disposable RNA copy. Corruption of this process is a hallmark of many diseases including cancer. mRNA synthesis requires not only making an RNA transcript but maturation of that transcript by 5' capping, excision of introns and splicing of exons and 3' end formation by cleavage/polyadenylation. The mRNA is also packaged with RNA binding proteins that facilitate its maturation and ultimate export to the cytoplasm as a messenger ribonucleoprotein particle (mRNP). The mRNA processing and packaging steps that radically transform the primary transcript occur largely co-transcriptionally; that is to say the substrate of mRNA processing and packaging factors is the growing nascent RNA that is extruded by an RNA polymerase II (pol II) molecule at rates of 0.5-4.5 kilobases/min. The goal of this proposal is to understand mRNP biosynthesis in its co- transcriptional context by focusing not on the mature mRNA products but on the nascent transcripts and how their transformation is affected by the process of transcription elongation that grows RNA chains. Our working model is that synthesis and processing of a mRNA precursor are carried out in an integrated fashion within a dynamic 'mRNA factory' complex that includes both RNA polymerase and processing factors some of which make direct contacts with the pol II C-terminal domain (CTD). We use genetic and genomic approaches to ask how ongoing transcription and mRNA maturation are coupled with one another in space through recruitment of factors to the `mRNA factory' and in time through kinetic coupling mechanisms to achieve efficient and accurate production of fully formed mRNP's. In this proposal we will address these questions: 1) How does pol II transcription elongation affect processing of the nascent transcript? 2) How are nascent transcripts folded, and how is folding affected by pol II transcription elongation? 3) How does transcription elongation affect RNA binding protein association with nascent transcripts? 4) How does elongation rate affect phosphorylation of the RNA pol II CTD and recruitment of processing factors to the transcription elongation complex? 5) How does co-transcriptional RNA processing affect elongation and chromatin structure?

Public Health Relevance

Many diseases including cancers are directly caused by the misexpression of genetic information that can occur by corruption of multiple steps in the pathway that makes messenger RNAs. A major point of vulnerability in this pathway is at essential steps in mRNA maturation by splicing and cleavage/polyadenylation; steps that must be properly coordinated with synthesis of these RNAs. The work in this proposal will help uncover how mistakes in the intricate coordination between synthesis and maturation of mRNAs can lead to errors in mRNA formation in diseased cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Unknown (R35)
Project #
5R35GM118051-02
Application #
9267500
Study Section
Special Emphasis Panel (ZGM1)
Program Officer
Sledjeski, Darren D
Project Start
2016-05-01
Project End
2021-04-30
Budget Start
2017-05-01
Budget End
2018-04-30
Support Year
2
Fiscal Year
2017
Total Cost
Indirect Cost
Name
University of Colorado Denver
Department
Biochemistry
Type
Schools of Medicine
DUNS #
041096314
City
Aurora
State
CO
Country
United States
Zip Code
80045
Ebmeier, Christopher C; Erickson, Benjamin; Allen, Benjamin L et al. (2017) Human TFIIH Kinase CDK7 Regulates Transcription-Associated Chromatin Modifications. Cell Rep 20:1173-1186
Fong, Nova; Saldi, Tassa; Sheridan, Ryan M et al. (2017) RNA Pol II Dynamics Modulate Co-transcriptional Chromatin Modification, CTD Phosphorylation, and Transcriptional Direction. Mol Cell 66:546-557.e3