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The rpoE gene encoding the sigma E (sigma 24) heat shock sigma factor of Escherichia coli.
EMBO J. 1995 Mar 01; 14(5):1043-55.EJ

Abstract

Previous work has established that the transcription factor sigma E (sigma 24) is necessary for maintaining the induction of the heat shock response of Escherichia coli at high temperatures. We have identified the gene encoding sigma E using a genetic screen designed to isolate trans-acting mutations that abolish expression from either htrA or rpoHP3, two promoters recognized uniquely by sigma E-containing RNA polymerase. Such a screen was achieved by transducing strains carrying a single copy of either phtrA-lacZ or rpoHP3-lacZ fusions with mutagenized bacteriophage P1 lysates and screening for Lac- mutant colonies at 22 degrees C. Lac- mutants were subsequently tested for inability to grow at 43 degrees C (Ts- phenotype). Only those Lac- Ts- mutants that were unable to accumulate heat shock proteins at 50 degrees C were retained for further characterization. In a complementary approach, those genes which when cloned on a multicopy plasmid led to higher constitutive expression of the sigma E regulon were characterized and mapped. Both approaches identified the same gene, rpoE, mapping at 55.5 min on the E.coli genetic map and encoding a polypeptide of 191 amino acid residues. The wild-type and a mutant rpoE gene products were over-expressed and purified. It was found that the purified wild-type sigma E protein, when used in in vitro run-off transcription assays in combination with core RNA polymerase, was able to direct transcription from the htrA and rpoHP3 promoters, but not from known sigma 70-dependent promoters. In vivo and in vitro analyses of rpoE transcriptional regulation showed that the rpoE gene is transcribed from two major promoters, one of which is positively regulated by sigma E itself.

Authors+Show Affiliations

Département de Biochimie Médicale, Centre Médical Universitaire, Genève, Switzerland.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

7889935

Citation

Raina, S, et al. "The rpoE Gene Encoding the Sigma E (sigma 24) Heat Shock Sigma Factor of Escherichia Coli." The EMBO Journal, vol. 14, no. 5, 1995, pp. 1043-55.
Raina S, Missiakas D, Georgopoulos C. The rpoE gene encoding the sigma E (sigma 24) heat shock sigma factor of Escherichia coli. EMBO J. 1995;14(5):1043-55.
Raina, S., Missiakas, D., & Georgopoulos, C. (1995). The rpoE gene encoding the sigma E (sigma 24) heat shock sigma factor of Escherichia coli. The EMBO Journal, 14(5), 1043-55.
Raina S, Missiakas D, Georgopoulos C. The rpoE Gene Encoding the Sigma E (sigma 24) Heat Shock Sigma Factor of Escherichia Coli. EMBO J. 1995 Mar 1;14(5):1043-55. PubMed PMID: 7889935.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - The rpoE gene encoding the sigma E (sigma 24) heat shock sigma factor of Escherichia coli. AU - Raina,S, AU - Missiakas,D, AU - Georgopoulos,C, PY - 1995/3/1/pubmed PY - 1995/3/1/medline PY - 1995/3/1/entrez SP - 1043 EP - 55 JF - The EMBO journal JO - EMBO J VL - 14 IS - 5 N2 - Previous work has established that the transcription factor sigma E (sigma 24) is necessary for maintaining the induction of the heat shock response of Escherichia coli at high temperatures. We have identified the gene encoding sigma E using a genetic screen designed to isolate trans-acting mutations that abolish expression from either htrA or rpoHP3, two promoters recognized uniquely by sigma E-containing RNA polymerase. Such a screen was achieved by transducing strains carrying a single copy of either phtrA-lacZ or rpoHP3-lacZ fusions with mutagenized bacteriophage P1 lysates and screening for Lac- mutant colonies at 22 degrees C. Lac- mutants were subsequently tested for inability to grow at 43 degrees C (Ts- phenotype). Only those Lac- Ts- mutants that were unable to accumulate heat shock proteins at 50 degrees C were retained for further characterization. In a complementary approach, those genes which when cloned on a multicopy plasmid led to higher constitutive expression of the sigma E regulon were characterized and mapped. Both approaches identified the same gene, rpoE, mapping at 55.5 min on the E.coli genetic map and encoding a polypeptide of 191 amino acid residues. The wild-type and a mutant rpoE gene products were over-expressed and purified. It was found that the purified wild-type sigma E protein, when used in in vitro run-off transcription assays in combination with core RNA polymerase, was able to direct transcription from the htrA and rpoHP3 promoters, but not from known sigma 70-dependent promoters. In vivo and in vitro analyses of rpoE transcriptional regulation showed that the rpoE gene is transcribed from two major promoters, one of which is positively regulated by sigma E itself. SN - 0261-4189 UR - https://www.unboundmedicine.com/medline/citation/7889935/The_rpoE_gene_encoding_the_sigma_E__sigma_24__heat_shock_sigma_factor_of_Escherichia_coli_ L2 - https://onlinelibrary.wiley.com/resolve/openurl?genre=article&sid=nlm:pubmed&issn=0261-4189&date=1995&volume=14&issue=5&spage=1043 DB - PRIME DP - Unbound Medicine ER -