PMID:7751275
| Citation |
Koh, YS and Roe, JH (1995) Isolation of a novel paraquat-inducible (pqi) gene regulated by the soxRS locus in Escherichia coli. J. Bacteriol. 177:2673-8 |
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| Abstract |
We have isolated promoters inducible by paraquat, a superoxide radical-generating agent, from Escherichia coli, using promoter-probing plasmid pJAC4 (Y.S. Koh and J.H. Roe, Korean J. Microbiol. 31:267-273, 1993). One promoter clone pqi-5 (pqi denotes paraquat-inducible gene) was mapped at 21.8 min on the E. coli chromosome by using the Kohara phage library. We constructed an operon fusion of the lacZ gene with the pqi-5 promoter to monitor the expression of the gene in the single-copy state. LacZ expression was induced about 7- to 13-fold by 77 to 780 microM paraquat. Other known superoxide generators such as menadione, phenazine methosulfate, and plumbagin also induced the expression of beta-galactosidase in this fusion strain. On the other hand, no significant induction was observed with treatment with hydrogen peroxide, ethanol, and heat shock. Induction of beta-galactosidase was significantly reduced by introducing a delta sox-8::cat or soxS3::Tn10 mutation into the fusion strain, indicating that pqi-5 is a member of the soxRS regulon. A DNA fragment containing the pqi-5 promoter was cloned and sequenced from the Kohara phage E2E5. We identified two pqi-5 open reading frames (ORFs); ORF-A encodes a predicted protein of 342 amino acids, and ORF-B is truncated at the cloning site. The transcription start site from the pqi-5 promoter was determined by primer extension and S1 nuclease protection analyses. Northern (RNA) and S1 analyses indicated that there are two kinds of pqi-5 transcript; one covers ORF-A only and the other covers ORF-A and possibly also ORF-B. |
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| Keywords |
Amino Acid Sequence; Bacterial Proteins/biosynthesis; Bacterial Proteins/genetics; Base Sequence; Chromosome Mapping; Cloning, Molecular; DNA-Binding Proteins; Escherichia coli/drug effects; Escherichia coli/genetics; Escherichia coli Proteins; Gene Expression Regulation, Bacterial; Genes, Bacterial/genetics; Membrane Proteins/biosynthesis; Membrane Proteins/genetics; Molecular Sequence Data; Mutagenesis, Insertional; Open Reading Frames/genetics; Oxidation-Reduction; Oxidative Stress/genetics; Paraquat/pharmacology; Promoter Regions, Genetic/genetics; RNA, Messenger/genetics; Recombinant Fusion Proteins/biosynthesis; Regulon/genetics; Repressor Proteins; Sequence Analysis, DNA; Trans-Activators; Transcription Factors/genetics |
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