The RNA polymerase II holoenzyme plays an important role in transcriptional regulatory mechanisms in the yeast Saccharomyces cerevisiae, as in other eukaryotes. The Srb/mediator complex, which is associated with the carboxy terminal domain (CTD), functions to integrate both positive and negative regulatory inputs to the polymerase. We recently showed that the holoenzyme is a direct target of the highly conserved Snfl protein kinase-signaling pathway, which responds to glucose limitation. This novel regulatory interaction provides an important new paradigm for transcriptional regulation. Genetic and biochemical analysis of the interactions of Snfl with the holoenzyme will provide insight into mechanisms. The effects of Snfl on genomic transcription patterns will be examined by DNA microarray analysis, and the interaction of Snfl with appropriate promoter sequences will be assayed by chromatin immunoprecipitation. We will examine the role of Snfl in effecting the changes in CTD phosphorylation pattern and SrblO/Srbl 1 kinase levels that occur in response to glucose depletion. The holoenzyme protein(s) that directly interact with Snfl will be identified. Msn3 (Std 1), a protein implicated in both the glucose response and the function of the transcription apparatus, will be characterized with respect to its role in the physical and functional interaction of Snfl with the holoenzyme. The functional relationship of Snfl and the CTD kinase CTDK-l will also be examined. Finally, we will address the role of the Srb/mediator complex in transcriptional repression by Sfl 1, a DNA-binding protein that interacts with the complex. The yeast system offers unique advantages for genetic analysis of transcriptional regulatory mechanisms that are conserved in mammals. The proposed studies will further our understanding of the functional interactions of RNA polymerase II holoenzymes with regulatory proteins and signal transduction pathways.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM047259-11
Application #
6636047
Study Section
Genetics Study Section (GEN)
Program Officer
Anderson, James J
Project Start
1992-05-01
Project End
2005-04-30
Budget Start
2003-05-01
Budget End
2004-04-30
Support Year
11
Fiscal Year
2003
Total Cost
$301,247
Indirect Cost
Name
Columbia University (N.Y.)
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
621889815
City
New York
State
NY
Country
United States
Zip Code
10032
Vyas, Valmik K; Berkey, Cristin D; Miyao, Takenori et al. (2005) Repressors Nrg1 and Nrg2 regulate a set of stress-responsive genes in Saccharomyces cerevisiae. Eukaryot Cell 4:1882-91
Berkey, Cristin D; Vyas, Valmik K; Carlson, Marian (2004) Nrg1 and nrg2 transcriptional repressors are differently regulated in response to carbon source. Eukaryot Cell 3:311-7
Kuchin, Sergei; Vyas, Valmik K; Kanter, Ellen et al. (2003) Std1p (Msn3p) positively regulates the Snf1 kinase in Saccharomyces cerevisiae. Genetics 163:507-14
Vincent, O; Kuchin, S; Hong, S P et al. (2001) Interaction of the Srb10 kinase with Sip4, a transcriptional activator of gluconeogenic genes in Saccharomyces cerevisiae. Mol Cell Biol 21:5790-6
Song, W; Carlson, M (1998) Srb/mediator proteins interact functionally and physically with transcriptional repressor Sfl1. EMBO J 17:5757-65
Treich, I; Ho, L; Carlson, M (1998) Direct interaction between Rsc6 and Rsc8/Swh3,two proteins that are conserved in SWI/SNF-related complexes. Nucleic Acids Res 26:3739-45
Kuchin, S; Carlson, M (1998) Functional relationships of Srb10-Srb11 kinase, carboxy-terminal domain kinase CTDK-I, and transcriptional corepressor Ssn6-Tup1. Mol Cell Biol 18:1163-71
Carlson, M (1997) Genetics of transcriptional regulation in yeast: connections to the RNA polymerase II CTD. Annu Rev Cell Dev Biol 13:1-23
Treich, I; Carlson, M (1997) Interaction of a Swi3 homolog with Sth1 provides evidence for a Swi/Snf-related complex with an essential function in Saccharomyces cerevisiae. Mol Cell Biol 17:1768-75
Song, W; Treich, I; Qian, N et al. (1996) SSN genes that affect transcriptional repression in Saccharomyces cerevisiae encode SIN4, ROX3, and SRB proteins associated with RNA polymerase II. Mol Cell Biol 16:115-20

Showing the most recent 10 out of 17 publications