The goal of this proposal is to understand the physiology of the heat shock (hs) response in Escherichia coli. We will investigate the mechanism of induction and regression of the response and in addition, the exact roles of the hs proteins both in normal bacterial growth and in protection of the cells at high temperature. Because of the tremendous conservation of the hs response throughout the living world, it is expected that many of the conclusions reached by the proposed studies on E. coli will contribute to increasing our understanding of the regulation and the significance of the hs response in all organisms. We will address several specific areas of research. (a) We will characterize the genes coding for the unidentified hs proteins by cloning, mapping and isolating mutations in them. The roles of these genes in E. coli physiology will be studied first by analyzing the phenotypes of mutants in both normal and hs conditions. The gene products will be purified and analyzed biochemically. (b) We will study the regulation of the hs response at the molecular level by investigating the roles of the htpR, dnaK, and other hs proteins in the modulation of the hs response. We will do this by the isolation of many mutants altered in the genes of interest, examination of their various phenotypes, and isolation of intergenic suppressors of the original mutations. Our biochemical approach will be to purify the wild-type and mutant proteins and also to study hs gene expression in vitro using standard techniques for transcription or coupled transcription and translation. (c) Bacteriophage lambda infection results in the induction of the hs response of E. coli in the absence of a temperature shift. We will identify the bacteriophage gene(s) responsible for this effect by analysis of mutants, purify their products, analyze the effects of the proteins in the in vitro system described above. This system may provide insight into the mechanisms by which animal viruses, such as adenovirus, polyoma, and SV40, induce the synthesis of certain host hs proteins during infection.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Microbial Physiology and Genetics Subcommittee 2 (MBC)
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University of Utah
Schools of Medicine
Salt Lake City
United States
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Wu, B; Ang, D; Snavely, M et al. (1994) Isolation and characterization of point mutations in the Escherichia coli grpE heat shock gene. J Bacteriol 176:6965-73
Zeilstra-Ryalls, J; Fayet, O; Georgopoulos, C (1994) Two classes of extragenic suppressor mutations identify functionally distinct regions of the GroEL chaperone of Escherichia coli. J Bacteriol 176:6558-65
Zeilstra-Ryalls, J; Fayet, O; Baird, L et al. (1993) Sequence analysis and phenotypic characterization of groEL mutations that block lambda and T4 bacteriophage growth. J Bacteriol 175:1134-43
Karow, M; Georgopoulos, C (1993) The essential Escherichia coli msbA gene, a multicopy suppressor of null mutations in the htrB gene, is related to the universally conserved family of ATP-dependent translocators. Mol Microbiol 7:69-79
Osipiuk, J; Georgopoulos, C; Zylicz, M (1993) Initiation of lambda DNA replication. The Escherichia coli small heat shock proteins, DnaJ and GrpE, increase DnaK's affinity for the lambda P protein. J Biol Chem 268:4821-7
Delaney, J M; Wall, D; Georgopoulos, C (1993) Molecular characterization of the Escherichia coli htrD gene: cloning, sequence, regulation, and involvement with cytochrome d oxidase. J Bacteriol 175:166-75
Delaney, J M; Ang, D; Georgopoulos, C (1992) Isolation and characterization of the Escherichia coli htrD gene, whose product is required for growth at high temperatures. J Bacteriol 174:1240-7
Karow, M; Georgopoulos, C (1992) Isolation and characterization of the Escherichia coli msbB gene, a multicopy suppressor of null mutations in the high-temperature requirement gene htrB. J Bacteriol 174:702-10
Delaney, J M; Georgopoulos, C (1992) Physical map locations of the trxB, htrD, cydC, and cydD genes of Escherichia coli. J Bacteriol 174:3824-5
Liberek, K; Galitski, T P; Zylicz, M et al. (1992) The DnaK chaperone modulates the heat shock response of Escherichia coli by binding to the sigma 32 transcription factor. Proc Natl Acad Sci U S A 89:3516-20

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