The structure of RNA polymerase is very similar from bacteria to man. This enzyme is responsible for the bulk of differential gene expression in all organisms. The properties of the large enzyme are modified by interactions with proteins and small molecule effectors. In this proposal a bacterial system is used to learn how the common core can have its properties changed. 3 sigma factors are used, the main housekeeping sigma70, sigma38 and sigma54, each of which causes the core to assume a different mechanism of action. Sigma38 causes the core to respond uniquely to several small molecule effectors. Sigma54 causes the core to respond to enhancer binding proteins. Sigma70 induces neither of these responses. A combination of genetic screens, band shift assays and footprinting analysis will be used to investigate this problem. Screens are devised to identify the parts of sigmas that are involved in these responses. The biochemical assays are used on the mutants obtained to learn why they are no longer able to alter the behavior of the core RNA polymerase. These mutants will be mapped on the known high resolution structures. The outcome is expected to be a description of the 3-dimensional domains on sigma that are critical for changing how the core polymerase works and new knowledge about how this happens. Because the core polymerase core is so conserved we expect that its manner of response will also be conserved, enlarging the possibilities for understanding the basis for human diseases of gene expression. ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM035754-21
Application #
7107146
Study Section
Special Emphasis Panel (ZRG1-MBC-2 (01))
Program Officer
Tompkins, Laurie
Project Start
1985-12-01
Project End
2008-08-31
Budget Start
2006-09-01
Budget End
2008-08-31
Support Year
21
Fiscal Year
2006
Total Cost
$265,530
Indirect Cost
Name
University of California Los Angeles
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
092530369
City
Los Angeles
State
CA
Country
United States
Zip Code
90095
Gralla, Jay D; Huo, Yi-Xin (2008) Remodeling and activation of Escherichia coli RNA polymerase by osmolytes. Biochemistry 47:13189-96
Huo, Yi-Xin; Rosenthal, Adam Z; Gralla, Jay D (2008) General stress response signalling: unwrapping transcription complexes by DNA relaxation via the sigma38 C-terminal domain. Mol Microbiol 70:369-78
Rosenthal, Adam Z; Kim, Youngbae; Gralla, Jay D (2008) Poising of Escherichia coli RNA polymerase and its release from the sigma 38 C-terminal tail for osmY transcription. J Mol Biol 376:938-49
Rosenthal, Adam Z; Kim, Youngbae; Gralla, Jay D (2008) Regulation of transcription by acetate in Escherichia coli: in vivo and in vitro comparisons. Mol Microbiol 68:907-17
Gralla, Jay D; Vargas, David R (2006) Potassium glutamate as a transcriptional inhibitor during bacterial osmoregulation. EMBO J 25:1515-21
Kim, Youngbae; Lew, Chih M; Gralla, Jay D (2006) Escherichia coli pfs transcription: regulation and proposed roles in autoinducer-2 synthesis and purine excretion. J Bacteriol 188:7457-63
Gralla, Jay D (2005) Escherichia coli ribosomal RNA transcription: regulatory roles for ppGpp, NTPs, architectural proteins and a polymerase-binding protein. Mol Microbiol 55:973-7
Lew, Chih M; Gralla, Jay D (2004) Nucleotide-dependent isomerization of Escherichia coli RNA polymerase. Biochemistry 43:12660-6
Lee, Shun Jin; Gralla, Jay D (2004) Osmo-regulation of bacterial transcription via poised RNA polymerase. Mol Cell 14:153-62
Lee, Shun Jin; Gralla, Jay D (2003) Open complex formation in vitro by sigma38 (rpoS) RNA polymerase: roles for region 2 amino acids. J Mol Biol 329:941-8

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