The PI has uncovered an essential Caulobacter gene, cgtA, that encodes a small monomeric GTP-binding protein and is a member of a recently identified subfamily of GTPases (the Obg family) conserved from humans to archea to bacteria. This evolutionary conservation between distantly related species suggests that this family of GTP-binding proteins shares an important physiological role. Obg proteins play a crucial role in cell growth, and in some cases, cell differentiation. Recently, it has been determined that the human homolog, DRG, functions in a pathway that is directly involved in oncogenesis in humans. Despite these recent advances, little is known about the mechanism by which these GTP-binding proteins function. Because of the ease with which cell cycle events in Caulobacter can be analyzed combined with the molecular and genetic advantages available when using a prokaryotic model system, they are in a unique position to uncover the cellular role of the Caulobacter GTPase, CgtA. Their long term goal is to elucidate the role of the CgtA mediated GTPase signal transduction pathway in Caulobacter. Towards this end, the laboratory has completed a molecular characterization of cgtA, has demonstrated that this gene is essential for viability, is a bona fide GTP-binding protein and is autophosphorylated. They have purified the protein and have raised CgtA-specific antibodies. They are now in a position to determine the mode of CgtA function, as well as the interplay between CgtA and as yet unknown interacting proteins involved in the same signaling pathway. During this funding period, they propose to: (1) isolate and examine both the in vitro GTP- binding characteristics and in vivo phenotype of specific cgtA mutants, (2) identify proteins that functionally or physically interact with CgtA, (3) determine the in vivo guanine nucleotide bond state of CgtA, (4) investigate the role of autophosphorylation, and (5) determine the intercellular location of CgtA by immunoelectron microscopy using antibodies they have generated to CgtA. From these studies, they will be able to suggest a role for this subfamily of GTPases and will develop tools which may be helpful for the identification of regulators and downstream effector proteins in other organisms.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM055133-04
Application #
6342925
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Program Officer
Anderson, Richard A
Project Start
1998-01-01
Project End
2002-12-31
Budget Start
2001-01-01
Budget End
2001-12-31
Support Year
4
Fiscal Year
2001
Total Cost
$200,080
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
791277940
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Fuentes, Jennifer L; Datta, Kaustuv; Sullivan, Susan M et al. (2007) In vivo functional characterization of the Saccharomyces cerevisiae 60S biogenesis GTPase Nog1. Mol Genet Genomics 278:105-23
Datta, Kaustuv; Fuentes, Jennifer L; Maddock, Janine R (2005) The yeast GTPase Mtg2p is required for mitochondrial translation and partially suppresses an rRNA methyltransferase mutant, mrm2. Mol Biol Cell 16:954-63
Datta, Kaustuv; Skidmore, Jennifer M; Pu, Kun et al. (2004) The Caulobacter crescentus GTPase CgtAC is required for progression through the cell cycle and for maintaining 50S ribosomal subunit levels. Mol Microbiol 54:1379-92
Wout, P; Pu, K; Sullivan, S M et al. (2004) The Escherichia coli GTPase CgtAE cofractionates with the 50S ribosomal subunit and interacts with SpoT, a ppGpp synthetase/hydrolase. J Bacteriol 186:5249-57
Lin, Bin; Thayer, Desiree A; Maddock, Janine R (2004) The Caulobacter crescentus CgtAC protein cosediments with the free 50S ribosomal subunit. J Bacteriol 186:481-9
Kroos, Lee; Maddock, Janine R (2003) Prokaryotic development: emerging insights. J Bacteriol 185:1128-46
Phadke, N D; Molloy, M P; Steinhoff, S A et al. (2001) Analysis of the outer membrane proteome of Caulobacter crescentus by two-dimensional electrophoresis and mass spectrometry. Proteomics 1:705-20
Lin, B; Skidmore, J M; Bhatt, A et al. (2001) Alanine scan mutagenesis of the switch I domain of the Caulobacter crescentus CgtA protein reveals critical amino acids required for in vivo function. Mol Microbiol 39:924-34
Molloy, M P; Phadke, N D; Maddock, J R et al. (2001) Two-dimensional electrophoresis and peptide mass fingerprinting of bacterial outer membrane proteins. Electrophoresis 22:1686-96
Skidmore, J M; Ellefson, D D; McNamara, B P et al. (2000) Polar clustering of the chemoreceptor complex in Escherichia coli occurs in the absence of complete CheA function. J Bacteriol 182:967-73

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