Abstract

The primary goal of this grant is to conduct structure-function studies to gain a better understanding of the regulation of transcriptional elongation by RNA Polymerase II (RNAP II). The phosphorylation state of the C- terminal domain (CTD) of RNAP II plays a key role in regulation of transcriptional elongation. FCP1, the first known CTD-specific phosphatase was recently cloned. In addition to its phosphatase activity, human FCP1 has been shown to directly interact with RAP74 (the large subunit of TFIIF), TFIIB and HIV-1 Tat, three factors which are known to affect the transcriptional elongation rates of RNAP II.
The specific aims of this grant are to 1) determine a high-resolution NMR structure of the FCP1 binding domain of RAP74; 2) determine a high-resolution NMR structure of a complex between RAP74 and FCP1; 3) characterize the interaction between TFIIB and FCP1 using NMR spectroscopy; 4) conduct preliminary binding and NMR studies with FCPI, the Tat protein and the TAR RNA. This research will help us understand how regulation of FCP1 affects transcriptional elongation by RNAP II. In addition, our studies will provide new insights into how disruptions in the control of transcriptional regulation may occur in many forms of human cancer and viral infections.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM060298-02
Application #
6351315
Study Section
Biophysical Chemistry Study Section (BBCB)
Program Officer
Wehrle, Janna P
Project Start
2000-02-01
Project End
2005-01-31
Budget Start
2001-02-01
Budget End
2002-01-31
Support Year
2
Fiscal Year
2001
Total Cost
$205,794
Indirect Cost

  Institution

Name
University of Georgia
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
City
Athens
State
GA
Country
United States
Zip Code
30602

  Publications

Di Lello, Paola; Jenkins, Lisa M Miller; Jones, Tamara N et al. (2006) Structure of the Tfb1/p53 complex: Insights into the interaction between the p62/Tfb1 subunit of TFIIH and the activation domain of p53. Mol Cell 22:731-40
Jenkins, Lisa M Miller; Durell, Stewart R; Maynard, Andrew T et al. (2006) Comparison of the specificity of interaction of cellular and viral zinc-binding domains with 2-mercaptobenzamide thioesters. J Am Chem Soc 128:11964-76
Abbott, Karen L; Renfrow, Matthew B; Chalmers, Michael J et al. (2005) Enhanced binding of RNAP II CTD phosphatase FCP1 to RAP74 following CK2 phosphorylation. Biochemistry 44:2732-45
Di Lello, Paola; Nguyen, Bao D; Jones, Tamara N et al. (2005) NMR structure of the amino-terminal domain from the Tfb1 subunit of TFIIH and characterization of its phosphoinositide and VP16 binding sites. Biochemistry 44:7678-86
Abbott, Karen L; Archambault, Jacques; Xiao, Hua et al. (2005) Interactions of the HIV-1 Tat and RAP74 proteins with the RNA polymerase II CTD phosphatase FCP1. Biochemistry 44:2716-31
Poole, Allison; Poore, Tim; Bandhakavi, Sricharan et al. (2005) A global view of CK2 function and regulation. Mol Cell Biochem 274:163-70
Nguyen, Bao D; Di Lello, Paola; Legault, Pascale et al. (2005) 1H, 15N, and 13C resonance assignment of the amino-terminal domain of the Tfb1 subunit of yeast TFIIH. J Biomol NMR 31:173-4
Jenkins, Lisa M Miller; Byrd, J Calvin; Hara, Toshiaki et al. (2005) Studies on the mechanism of inactivation of the HIV-1 nucleocapsid protein NCp7 with 2-mercaptobenzamide thioesters. J Med Chem 48:2847-58
Nguyen, Bao D; Abbott, Karen L; Potempa, Krzysztof et al. (2003) NMR structure of a complex containing the TFIIF subunit RAP74 and the RNA polymerase II carboxyl-terminal domain phosphatase FCP1. Proc Natl Acad Sci U S A 100:5688-93
Nguyen, Bao D; Chen, Hung-Ta; Kobor, Michael S et al. (2003) Solution structure of the carboxyl-terminal domain of RAP74 and NMR characterization of the FCP1-binding sites of RAP74 and human TFIIB. Biochemistry 42:1460-9