Nucleotide sequence of the heat shock regulatory gene of E. coli suggests its protein product may be a transcription factor

Landick, Robert; Vaughn, Vicki; Lau, Elizabeth T.; VanBogelen, Ruth A.; Erickson, James; Neidhardt, Frederick C.

doi:10.1016/0092-8674(84)90538-5

Cited by 198 publications

(129 citation statements)

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“…Induction of the heat shock response is regulated at the level of transcription. In E. coli, the heat shock genes are positively regulated by the rpoH gene, whose product, a 32-kDa sigma factor, directs core RNA polymerase to recognize the promoters for heat shock genes (12,25,34). In B. subtilis, only one transcription start site is preceded by a promoter sequence recognized by sigma-43, the vegetative sigma factor.…”

Section: Discussionmentioning

confidence: 99%

Cloning, sequencing, and molecular analysis of the dnaK locus from Bacillus subtilis

et al. 1992

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By using an internal part of the dnaK gene from BaciUlus megaterium as a probe, a 5.2-kb Hindlll fragment of chromosomal DNA of BaciUlus subtilis was cloned. Downstream sequences were isolated by in vivo chromosome walking. Sequencing of 5,085 bp revealed four open reading frames in the order orf9-grpEdnaK-dnaJ. orJ39 encodes a 39-kDa polypeptide of unknown biological function with no noticeable homology to any other protein within the data bases. Alignment of the GrpE protein with those of three other bacterial species revealed a low overall homology, but a higher homology restricted to two regions which might be involved in interactions with other proteins. Alignment of the DnaK protein with six bacterial DnaK polypeptides revealed that a contiguous region of 24 amino acids is absent from the DnaK proteins of all known gram-positive species. Primer extension studies revealed three potential transcription start sites, two preceding orJ39 (Si and S2) and a third one in front of grpE (S3). S2 and S3 were activated at a high temperature. Northern (RNA) analysis led to the detection of three mRNA species of 4.9, 2.6, and 1.5 kb. RNA dot blot experiments revealed an at-least-fivefold increase in the amount of specific mRNA from 0 to 5 min postinduction and then a rapid decrease. A transcriptional fusion between dnaK and the amyL reporter gene exhibited a slight increase in a-amylase activity after heat induction. A 9-bp inverted repeat was detected in front of the coding region of orJ39. This inverted repeat is present in a number of other heat shock operons in other microorganisms ranging from cyanobacteria to mycobacteria. The biological property of this inverted repeat as a putative key element in the induction of heat shock genes is discussed. The dnaK locus was mapped at about 2230 on the B. subtilis genetic map.The heat shock response is a homeostatic mechanism that enables cells to survive a variety of environmental stresses. It is characterized by the increased synthesis of a group of evolutionarily conserved proteins, heat shock proteins (HSPs), and is a universal feature of both prokaryotic and eukaryotic cells (35). When Eschertichia coli cells are shifted to a high temperature, the synthesis of a set of about 20 HSPs transiently increases and then declines rapidly to steady-state levels characteristic of the new ambient temperature (44).The highly conserved HSPs perform similar functions in all organisms. One of these functions is the well-established regulation of protein-protein interactions by the chaperonins (69). One of the most abundant HSPs, HSP70, is highly conserved in evolution. It is found in such diverse organisms as E. coli, Saccharomyces cerevisiae, Drosophila melanogaster, and Homo sapiens (14, 35).The dnaK gene of E. coli was originally discovered because mutations in it blocked bacteriophage lambda DNA replication at all temperatures (22,23). Subsequently, the dnaK gene product was shown to be essential for E. coli viability at high and low temperatures (22,30,46,51,52), and genetic evid...

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Section: Discussionmentioning

confidence: 99%

Cloning, sequencing, and molecular analysis of the dnaK locus from Bacillus subtilis

et al. 1992

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“…It has been demonstrated that Streptomyces, like E. coli (Grossman et al, 1984;Landick et al, 1984), Bacillus (Losick & Pero, 1981) and recently the mntrogen-regulated genes of enteric bacteria (Hirschman et al, 1985) possess multiple sigma (C) factors (Westpheling et al, 1985;Buttner & Brown, 1985) which are each believed to initiate transcription from a specific class of promoter. In S. coelicolor one of the a-factors identified, probably the o.40 found by Buttner & Brown (1985) or o45 of Westpheling et al (1985), may be analogous to the E. coli a`70 factor that recognizes the consensus promoter of TTGACA (-35 region) and TATAAT (-10 region), and may be partly responsible for the capacity of Streptomyces to recognize and initiate transcription from promoters derived from a diverse source of bacteria including E. coli (Bibb & Cohen, 1982;Jaurin & Cohen, 1984;Westpheling et al, 1985;Herbert et al, 1986).…”

Section: Streptomyces Lividansmentioning

confidence: 99%

Transformation of Arthrobacter and studies on the transcription of the Arthrobacter ermA gene in Streptomyces lividans and Escherichia coli

Roberts¹,

Barnett²,

Brenner³

1987

Biochemical Journal

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We report the development of a plasmid-mediated transformation system for Arthrobacter sp. NRRLB3381, using the Streptomyces cloning vector pIJ702. Our procedure gives a transformation frequency of 10(3)/micrograms of plasmid DNA. In addition we have explored the expression of the Arthrobacter ermA gene in Streptomyces lividans and Escherichia coli, and shown that the ermA promoter is recognized in S. lividans not E. coli. The relationship between Arthrobacter, Streptomyces and E. coli promoters is discussed.

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“…Understanding this mode of regulation is central to our understanding of protein quality control as well as cellular stress responses. Here, we report the determinants required for chaperone regulation of 32 , the heat shock transcription factor in Escherichia coli (10)(11)(12). 32 regulon members control both the activity and stability of 32 .…”

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confidence: 99%

Analysis of σ ³² mutants defective in chaperone-mediated feedback control reveals unexpected complexity of the heat shock response

Yura

Guisbert²,

Poritz

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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Protein quality control is accomplished by inducing chaperones and proteases in response to an altered cellular folding state. In Escherichia coli, expression of chaperones and proteases is positively regulated by 32 . Chaperone-mediated negative feedback control of 32 activity allows this transcription factor to sense the cellular folding state. We identified point mutations in 32 altered in feedback control. Surprisingly, such mutants are resistant to inhibition by both the DnaK/J and GroEL/S chaperones in vivo and also show dramatically increased stability. Further characterization of the most defective mutant revealed that it has almost normal binding to chaperones and RNA polymerase and is competent for chaperone-mediated inactivation in vitro. We suggest that the mutants identify a regulatory step downstream of chaperone binding that is required for both inactivation and degradation of 32 .heat shock transcription factor ͉ proteolysis ͉ GroEL ͉ DnaK ͉ stress response T he heat shock response is a major homeostatic mechanism for controlling the state of protein folding and degradation in all cells (1-3). Upon heat stress, a set of highly conserved heat shock proteins (hsps), including chaperones and proteases, is rapidly and transiently induced. Hsps maintain optimal states of protein folding and turnover during normal growth and also minimize cellular damage from stress-induced protein misfolding and aggregation (4, 5). The level of hsps is controlled primarily by heat shock transcription factors that sense the cellular folding environment through negative feedback control mediated by chaperones (6-9). Understanding this mode of regulation is central to our understanding of protein quality control as well as cellular stress responses. Here, we report the determinants required for chaperone regulation of 32 , the heat shock transcription factor in Escherichia coli (10-12).32 regulon members control both the activity and stability of 32 . The DnaK/J/GrpE and GroEL/S chaperone machines each constitute a negative feedback loop that couples 32 activity to cellular protein folding state: overexpression of either chaperone machine decreases 32 activity; conversely, chaperone depletion or overexpression of chaperone substrates increases 32 activity (13-16). The chaperones are likely to act directly on 32 because they bind to 32 and inhibit its activity in a purified in vitro transcription system (17-19). Regulated degradation of 32 is mediated by the FtsH protease and facilitated by DnaK/J/GrpE and GroEL/S in vivo, but this process has not been completely recapitulated in vitro, where degradation of 32 by FtsH is slow and not facilitated by chaperones (20,21).We selected and characterized feedback-resistant mutants of 32 . The residues altered by the mutations map to a small patch in 32 . Surprisingly, most mutants simultaneously diminished all three negative feedback loops operative in vivo; the most severe mutant essentially eliminated chaperone-mediated activity control and degradation by FtsH protease. In stri...

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Nucleotide sequence of the heat shock regulatory gene of E. coli suggests its protein product may be a transcription factor

Cited by 198 publications

References 25 publications

Cloning, sequencing, and molecular analysis of the dnaK locus from Bacillus subtilis

Cloning, sequencing, and molecular analysis of the dnaK locus from Bacillus subtilis

Transformation of Arthrobacter and studies on the transcription of the Arthrobacter ermA gene in Streptomyces lividans and Escherichia coli

Analysis of σ ³² mutants defective in chaperone-mediated feedback control reveals unexpected complexity of the heat shock response

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Nucleotide sequence of the heat shock regulatory gene of E. coli suggests its protein product may be a transcription factor

Cited by 198 publications

References 25 publications

Cloning, sequencing, and molecular analysis of the dnaK locus from Bacillus subtilis

Cloning, sequencing, and molecular analysis of the dnaK locus from Bacillus subtilis

Transformation of Arthrobacter and studies on the transcription of the Arthrobacter ermA gene in Streptomyces lividans and Escherichia coli

Analysis of σ 32 mutants defective in chaperone-mediated feedback control reveals unexpected complexity of the heat shock response

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Analysis of σ ³² mutants defective in chaperone-mediated feedback control reveals unexpected complexity of the heat shock response