Cloning and DNA sequence of<i>sigB</i>gene of<i>Stigmatella aurantiaca</i>

Skladny, Heyko; Heidelbach, M; Schairer, Hans Ulrich

doi:10.1093/nar/20.23.6416

Cited by 9 publications

(2 citation statements)

References 4 publications

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“…A positive clone, pAR1, was sequenced and characterized. The 1.8-kb insert in pAR1 contained an open reading frame encoding a protein that is a member of the 32 protein family, which includes homologs from E. coli (8), Citrobacter freundii (18), Haemophilus influenzae (17), Myxococcus xanthus (1, 2), Pseudomonas aeruginosa (5), and Stigmatella aurantiaca (43). The 32 family includes SigB and SigC, proteins reportedly involved in spore maturation and fruiting body formation in M. xanthus (1,2).…”

Section: Resultsmentioning

confidence: 99%

Regulation of a heat shock sigma32 homolog in Caulobacter crescentus

1996

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High temperature and other environmental stresses induce the expression of several heat shock proteins in Caulobacter crescentus, including the molecular chaperones DnaJ, DnaK, GrpE, and GroEL and the Lon protease. We report here the isolation of the rpoH gene encoding a homolog of the Escherichia coli RNA polymerase 32 subunit, the sigma factor responsible for the transcription of heat shock promoters. The C. crescentus 32 homolog, predicted to be a 33.7-kDa protein, is 42% identical to E. coli 32 and cross-reacts with a monoclonal antibody to E. coli 32. Functional homology was demonstrated by complementing the temperature-sensitive growth defect of an E. coli rpoH deletion mutant with the C. crescentus rpoH gene. Immunoblot analysis showed a transient rise in 32 levels after a temperature shift from 30 to 42؇C similar to that described for E. coli. In addition, increasing the cellular content of 32 by introducing a plasmid-encoded copy of rpoH induced DnaK expression in C. crescentus cultures grown at 30؇C. The C. crescentus rpoH gene was transcribed from either of two heat shock consensus promoters. rpoH transcription and 32 levels increased coordinately following heat shock, indicating that transcriptional regulation contributes to 32 expression in this organism. Both the rpoH gene and 32 protein were expressed constitutively throughout the cell cycle at 30؇C. The isolation of rpoH provides an important tool for future studies of the role of 32 in the normal physiology of C. crescentus.Heat shock proteins (HSPs) are generally either molecular chaperones that act to maintain the proper folding of cell proteins, such as DnaK or Hsp70 and GroEL or Hsp60, or proteases, such as Clp and Lon. This group of proteins, found in prokaryotes and eukaryotes, functions both at physiological temperature and during cellular responses to high temperature and other environmental stresses. Distinct modes of transcriptional induction of the HSPs have been described for Escherichia coli and Saccharomyces cerevisiae (reviewed in reference 27). In E. coli, heat shock enhances the level of the 32 subunit of RNA polymerase which then recognizes the promoters of the heat shock genes and increases their levels of transcription (10,22,24). By contrast, in S. cerevisiae a transcriptional activator, HSF, binds constitutively to a highly conserved DNA repeat, or heat shock element, in the promoter region. Elevated temperature induces a conformational change in HSF which enables it to activate the transcription of heat shock genes (27).Although the general heat shock response in bacteria is highly conserved, its regulation by 32 has previously been documented in bacteria that are members of the ␥ subdivision of proteobacteria. Only recently have 32 homologs been reported in ␣ proteobacteria (32). The role of 32 in the heat shock response is best understood for E. coli (reviewed in references 7, 19, 21, and 50). In this organism, 32 levels increase 15-to 20-fold in response to a sudden rise in temperature (25,44 promoters and a E promoter ...

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Section: Resultsmentioning

confidence: 99%

Regulation of a heat shock sigma32 homolog in Caulobacter crescentus

1996

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“…Therefore, it is reasonable to speculate that the RpoH-specific sequences may be involved in the chaperon-mediated negative control of the synthesis and degradation of 32 proteins. The 32 proteins are also closely related to the SigB (2) and SigC (3) proteins of M. xanthus and to the SigB protein of S. aurantiaca (41). The M. xanthus SigC protein and the S. aurantiaca SigB protein have not been demonstrated to be involved in heat shock response.…”

Section: Discussionmentioning

confidence: 99%

Isolation, identification, and transcriptional specificity of the heat shock sigma factor sigma32 from Caulobacter crescentus

Newton

1996

J Bacteriol

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We report the identification of the Caulobacter crescentus heat shock factor 32 as a 34-kDa protein that copurifies with the RNA polymerase holoenzyme. The N-terminal amino acid sequence of this protein was determined and used to design a degenerate oligonucleotide as a probe to identify the corresponding gene, rpoH, which encodes a predicted protein with a molecular mass of 33,659 Da. The amino acid sequence of this protein is similar to those of known bacterial heat shock sigma factors of Escherichia coli (41% identity), Pseudomonas aeruginosa (40% identity), and Citrobacter freundii (38% identity). The isolated C. crescentus gene complements the growth defect of an E. coli rpoH deletion strain at 37؇C, and Western blot (immunoblot) analysis confirmed that the gene product is related to the E. coli 32 protein. The purified RpoH protein in the presence of RNA polymerase core enzyme specifically recognizes the heat shock-regulated promoter P1 of the C. crescentus dnaK gene, and base pair substitutions in either the ؊10 or ؊35 region of this promoter abolish transcription. S1 nuclease mapping indicates that rpoH transcripts originate from two promoters, P1 and P2, under the normal growth conditions. The P2 promoter is similar to the 32 promoter consensus, and the P2-specific transcript increases dramatically during heat shock, while the P1-specific transcript remains relatively constant. These results suggest that although the structure and function of C. crescentus 32 appear to be very similar to those of its E. coli counterpart, the C. crescentus rpoH gene contains a novel promoter structure and may be positively autoregulated in response to environmental stress.The Caulobacter crescentus cell cycle generates two different daughter cells: a motile swarmer cell which possesses a single polar flagellum, and a nonmotile stalked cell. The progeny swarmer and stalked cell types differ not only in morphology and motility but also in their programs of gene expression and capacities to reinitiate DNA replication. Formation of the new swarmer cell results from a series of discrete morphogenic events during the cell cycle that occur at one pole of the dividing cell (reviewed in references 8 and 38). This developmental sequence depends on completion of successive cell cycle checkpoints, and there is now evidence that the regulation of cell division and some developmental events is mediated by two-component signal transduction pathways (reviewed in reference 27).Many developmental events in C. crescentus are regulated at the level of transcription initiation. The specificity of promoter recognition is conferred by the sigma subunit of RNA polymerase (RNAP), which binds to the core RNAP (E) containing ␤, ␤Ј, and ␣ 2 . In eubacterial systems examined to date, multiple sigma factors have been identified, each of which programs RNAP to recognize a distinct promoter sequence. Although the predominant sigma subunit ( 70 in gram-negative bacteria) directs expression of the bulk of the genome, highly specialized genes involved in ...

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