Caulobacter crescentus is an asymmetrically dividing, Gram negative bacterium that has proved to be an excellent model for the study of cell differentiation. The most striking feature of the stalked cell cycle of this organism is the stage specific sequence of events that leads to the generation of a new motile swarmer cell at division. Our long term goal is to understand the developmental program that controls the precise sequence of these events and coordinates their timing and localization of cell surface structures. Our studies have focused on flagellum formation which requires over 50 genes that are organized in a regulatory hierarchy. Periodic expression of the flagellar genes in the cell cycle is mediated by a cascade of trans-acting regulatory factors. The goal of work proposed in this application is to characterize the trans-acting factors and the DNA sequence elements with which they interact to determine the observed temporal regulation of transcriptional specificity.
Our specific aims are to i) Determine the location and mechanism of FlbD binding to DNA sequence elements required for regulation of sigma 54 promoters; ii) Determine if FlbD, which is a functional homologue of response regulatory protein NtrC, is modified by phosphorylation and the role of modification for its function as a transcriptional factor; iii) Identify genes in the flaO and flbF operons that are required along with flbD for activation of sigma 54 promoters at the bottom of the hierarchy; iv) Purify RNA polymerases from Caulobacter to characterize specialized sigma factors in order to asses their role in determining promoter specificity; v) Investigate the role of DNA synthesis as a cell cycle signal that triggers the flagellar gene cascade; and vi) Examine the hook protein for sequences that target it to the cell pole for assembly using translation fusions to reporter genes.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM022299-17
Application #
3271066
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Project Start
1978-09-01
Project End
1995-08-31
Budget Start
1991-09-01
Budget End
1992-08-31
Support Year
17
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Princeton University
Department
Type
Schools of Arts and Sciences
DUNS #
002484665
City
Princeton
State
NJ
Country
United States
Zip Code
08544
Mullin, D A; Ohta, N; Mullin, A H et al. (2001) Organization, expression, and function of Caulobacter crescentus genes needed for assembly and function of the flagellar hook. Mol Genet Genomics 265:445-54
Wu, J; Ohta, N; Zhao, J L et al. (1999) A novel bacterial tyrosine kinase essential for cell division and differentiation. Proc Natl Acad Sci U S A 96:13068-73
Ward, D V; Newton, A (1999) Cell cycle expression and transcriptional regulation of DNA topoisomerase IV genes in caulobacter. J Bacteriol 181:3321-9
Wu, J; Ohta, N; Newton, A (1998) An essential, multicomponent signal transduction pathway required for cell cycle regulation in Caulobacter. Proc Natl Acad Sci U S A 95:1443-8
Wu, J; Newton, A (1997) The Caulobacter heat shock sigma factor gene rpoH is positively autoregulated from a sigma32-dependent promoter. J Bacteriol 179:514-21
Anderson, D K; Newton, A (1997) Posttranscriptional regulation of Caulobacter flagellin genes by a late flagellum assembly checkpoint. J Bacteriol 179:2281-8
Wu, J; Ohta, N; Benson, A K et al. (1997) Purification, characterization, and reconstitution of DNA-dependent RNA polymerases from Caulobacter crescentus. J Biol Chem 272:21558-64
Wu, J; Newton, A (1997) Regulation of the Caulobacter flagellar gene hierarchy; not just for motility. Mol Microbiol 24:233-9
Wu, J; Newton, A (1996) Isolation, identification, and transcriptional specificity of the heat shock sigma factor sigma32 from Caulobacter crescentus. J Bacteriol 178:2094-101
Anderson, D K; Ohta, N; Wu, J et al. (1995) Regulation of the Caulobacter crescentus rpoN gene and function of the purified sigma 54 in flagellar gene transcription. Mol Gen Genet 246:697-706

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