Oxygen radicals are responsible for a large fraction of the toxic and mutagenic effects of ionizing radiation. Such radicals are also produced by a variety of environmental chemicals and as side products of normal aerobic metabolism. Multilayered cellular defenses have evolved in response to this threat, consisting in part of some enzymes that prevent damage by scavenging reactive oxygen species and others that repair free radical damage to the critical molecular DNA. These defenses are coordinated as inducible systems in Escherichia coli that respond to different kinds of oxidative stress. One group of proteins (the soxR regulon) is induced specifically by the intracellular generation of superoxide radical and increases the levels of several enzymes that destroy superoxide, produce reducing equivalents in the form of NADPH, and repair oxidative damage to DNA. Another set of responses is triggered by hydrogen peroxide, and includes inducible enzymes that destroy organic or hydrogen peroxides, along with a large number of unknown inducible functions. We will dissect the biological functions of these adaptive responses to oxidative stress by a detailed characterization of the soxR gene and its regulon, and by the identification of H2O2-inducible genes through the use of gene fusions. We will determine the DNA damages formed in vivo that require the soxR-controlled enzyme endonuclease IV for their repair. This integrated molecular genetic and biochemical analysis of the biological responses to oxidative damage in a facile bacterial system will provide important information about general biological mechanisms for dealing with the toxicity of intrinsic and radiation-induced oxygen radical damage.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA037831-06
Application #
3175695
Study Section
Chemical Pathology Study Section (CPA)
Project Start
1984-08-01
Project End
1994-06-30
Budget Start
1990-07-01
Budget End
1991-06-30
Support Year
6
Fiscal Year
1990
Total Cost
Indirect Cost
Name
Harvard University
Department
Type
Schools of Arts and Sciences
DUNS #
071723621
City
Cambridge
State
MA
Country
United States
Zip Code
02138
Liu, Chunhua; Kim, Eunsuk; Demple, Bruce et al. (2012) A DNA-based nanomechanical device used to characterize the distortion of DNA by Apo-SoxR protein. Biochemistry 51:937-43
Lee, Paul E; Demple, Bruce; Barton, Jacqueline K (2009) DNA-mediated redox signaling for transcriptional activation of SoxR. Proc Natl Acad Sci U S A 106:13164-8
Lee, Yunho; Pena-Llopis, Samuel; Kang, Yoon-Suk et al. (2006) Expression analysis of the fpr (ferredoxin-NADP+ reductase) gene in Pseudomonas putida KT2440. Biochem Biophys Res Commun 339:1246-54
Park, Woojun; Pena-Llopis, Samuel; Lee, Yunho et al. (2006) Regulation of superoxide stress in Pseudomonas putida KT2440 is different from the SoxR paradigm in Escherichia coli. Biochem Biophys Res Commun 341:51-6
Koutsolioutsou, Anastasia; Pena-Llopis, Samuel; Demple, Bruce (2005) Constitutive soxR mutations contribute to multiple-antibiotic resistance in clinical Escherichia coli isolates. Antimicrob Agents Chemother 49:2746-52
Chander, Monica; Demple, Bruce (2004) Functional analysis of SoxR residues affecting transduction of oxidative stress signals into gene expression. J Biol Chem 279:41603-10
Pomposiello, Pablo J; Koutsolioutsou, Anastasia; Carrasco, Daniel et al. (2003) SoxRS-regulated expression and genetic analysis of the yggX gene of Escherichia coli. J Bacteriol 185:6624-32
Chander, Monica; Raducha-Grace, Laura; Demple, Bruce (2003) Transcription-defective soxR mutants of Escherichia coli: isolation and in vivo characterization. J Bacteriol 185:2441-50
Koutsolioutsou, A; Martins, E A; White, D G et al. (2001) A soxRS-constitutive mutation contributing to antibiotic resistance in a clinical isolate of Salmonella enterica (Serovar typhimurium). Antimicrob Agents Chemother 45:38-43
Pomposiello, P J; Bennik, M H; Demple, B (2001) Genome-wide transcriptional profiling of the Escherichia coli responses to superoxide stress and sodium salicylate. J Bacteriol 183:3890-902

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