Transcriptional Analysis of Major Heat Shock Genes of
            <i>Helicobacter pylori</i>

Homuth, Georg; Domm, Stephanie; Kleiner, Diethelm; Schumann, Wolfgang

doi:10.1128/jb.182.15.4257-4263.2000

Cited by 38 publications

(35 citation statements)

References 45 publications

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“…1. P gro transcribes one bicistronic mRNA encoding the chaperonin machinery GroESL; P hrc transcribes a tricistronic mRNA encoding the putative repressor HrcA, the chaperone DnaK, and its cochaperone GrpE (8,17); and P cbp is likely to transcribe a tricistronic mRNA encoding the DnaJ homologue CbpA, the HspR repressor, and a putative DNA helicase (17). To investigate the transcriptional response of these promoters to a sudden temperature increase, total RNA was isolated from H. pylori strain G27 grown at 37°C and at different time points after upshift of the culture to 42°C.…”

Section: Resultsmentioning

confidence: 99%

“…The control of transcription by HspR depends on the binding of this protein to large operators mapping to different positions surrounding the promoters (17). In addition to this regulation, the possibility that another transcriptional repressor, a homolog of the B. subtilis HrcA protein (16) encoded by the hrcA-grpE-dnaK locus, might also be involved in controlling stress response has been postulated (8,17). Evidences that chaperone gene transcription in this system responds to an increase in salinity and temperature elevation to 42°C, but not to 45°C, have been recently reported (8,17).…”

Section: Discussionmentioning

confidence: 99%

“…While the P gro and P cbp promoters were found to be strongly inducible by treatment with 300 mM NaCl, no induction could be observed on either promoter by incubation of cultures at 45°C, indicating that heat shock does not induce transcription from HspRdependent promoters. Recently, however, Homuth and coworkers (8) have shown that the P gro and P hrc promoters are strongly inducible by a mild heat shock at 42°C, suggesting that HspR can indeed mediate the transcriptional response to a sudden temperature increase. In the present work we determine in detail the transcriptional response of the three HspRdependent promoters to different environmental stresses.…”

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confidence: 99%

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Characterization of the HspR-Mediated Stress Response inHelicobacter pylori

Spohn¹,

Delany²,

Rappuoli³

et al. 2002

J Bacteriol

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The major heat shock genes of Helicobacter pylori are regulated by the HspR repressor. In the present study we characterize the transcriptional response of the three known HspR-dependent promoters P cbp , P gro , and P hrc to different environmental stresses. A temperature shift from 37 to 42°C causes a typical heat shock response at all three promoters characterized by an immediate and strong induction phase of transcription and a subsequent adaptation phase, which is specific for each promoter and whose onset is determined partially by the half-lives of the respective mRNAs. Exposure to high osmolarity induces a similar response on the P gro and P cbp promoters while no such response is detectable at the P hrc promoter. Puromycin treatment induces transcription from all three HspR-dependent promoters, indicating that different environmental stresses are intracellularly sensed by the regulatory machinery through the accumulation of nonnative proteins. The implications of these data for the regulatory network controlling the heat shock response in H. pylori are discussed.The heat shock proteins of the gastric pathogen Helicobacter pylori have been studied in some detail both because of their general role in the protection of the bacteria from the hostile environment represented by the human stomach and because of their involvement in specific pathogenic processes. The H. pylori GroEL (Hsp60) homologue (12, 20) has been proposed to play a role in regulating the activity of the nickel-dependent urease enzyme, which generates ammonia ions from the hydrolysis of urea and therefore protects the bacteria from the low pH of the stomach lumen (4, 6). Its cochaperone GroES (Hsp10) is thought to contribute to this regulation by controlling the availability of nickel ions by means of its intrinsic metal-binding activity (10,20). GroEL as well as another major heat shock protein, DnaK (Hsp70), can furthermore be found in association with the outer membrane, and this surface localization has been suggested to modify the glycolipid binding specificity of H. pylori cells at a low pH (5, 9, 15).Because of their multiple functions in both the general stress response and in specific disease mechanisms, the H. pylori heat shock proteins are expected to be tightly regulated in their expression levels. It was recently demonstrated that transcription of the groESL, hrcA-grpE-dnaK, and cbpA-hspR-orf operons encoding the major chaperones of H. pylori is negatively regulated by HspR (17). HspR is a homologue of the repressor of the dnaK operon of Streptomyces coelicolor (2, 7). By RNA analyses and footprint experiments with purified protein preparations, we have shown that HspR represses transcription by binding to regions of 75 bp on the three chaperone genetranscribing promoters P gro , P hrc , and P cbp . In an attempt to identify the environmental stresses, which induce transcription from these promoters, we subjected H. pylori cultures to both osmotic shock and thermal shock at 45°C. While the P gro and P cbp promoters were found t...

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Characterization of the HspR-Mediated Stress Response inHelicobacter pylori

Spohn¹,

Delany²,

Rappuoli³

et al. 2002

J Bacteriol

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“…Initial studies indicated that while transcription of the groESL and cbpA-hspR-orf operons was strongly inducible by treatment with 300 mM NaCl, no induction was observed when cultures were incubated at 45°C (31). Subsequently, Homouth and coworkers (15) showed that transcription of these operons is strongly inducible by a mild heat shock at 42°C, suggesting that HspR can indeed mediate the transcriptional response to a sudden temperature increase, characterized by fast and strong induction of transcription and a subsequent shutoff phase, whose onset is determined largely by the stability of the respective mRNAs (29). Until recently, the study of HrcAdependent regulation was hampered by difficulties in purifying a recombinant protein to map the HrcA binding sites on the promoters (25,31).…”

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Transcriptional Regulation of Stress Response and Motility Functions inHelicobacter pyloriIs Mediated by HspR and HrcA

et al. 2007

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The hrcA and hspR genes of Helicobacter pylori encode two transcriptional repressor proteins that negatively regulate expression of the groES-groEL and hrcA-grpE-dnaK operons. While HspR was previously shown to bind far upstream of the promoters transcribing these operons, the binding sites of HrcA were not identified. Here, we demonstrate by footprinting analysis that HrcA binds to operator elements similar to the so-called CIRCE sequences overlapping both promoters. Binding of HspR and HrcA to their respective operators occurs in an independent manner, but the DNA binding activity of HrcA is increased in the presence of GroESL, suggesting that the GroE chaperonin system corepresses transcription together with HrcA. Comparative transcriptome analysis of the wild-type strain and hspR and hrcA singly and doubly deficient strains revealed that a set of 14 genes is negatively regulated by the action of one or both regulators, while a set of 29 genes is positively regulated. While both positive and negative regulation of transcription by HspR and/or HrcA could be confirmed by RNA primer extension analyses for two representative genes, binding of either regulator to the promoters could not be detected, indicating that transcriptional regulation at these promoters involves indirect mechanisms. Strikingly, 14 of the 29 genes which were found to be positively regulated by HspR or HrcA code for proteins involved in flagellar biosynthesis. Accordingly, loss of motility functions was observed for HspR and HrcA single or double mutants. The possible regulatory intersections of the heat shock response and flagellar assembly are discussed.

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“…Interestingly, the molecular chaperone genes dnaJK, the heat shock protein gene grpE, and the heat shock response transcriptional regulator gene hrcA, and the chaperone genes groESL, the ATP-dependent clp protease genes clpEPC, and the transcriptional regulator gene ctsR, show higher FPKM values and are mainly known for their induction under heat stress. The chaperones encoded by dnaK and dnaJ are involved in the degradation of misfolded proteins in E. coli (26) and other bacteria after application of higher temperatures (27)(28)(29)(30). Together with hrcA and grpE, the operon structure hrcA-grpE-dnaK-dnaJ is already described in B. subtilis (31), Enterococcus faecalis (32), Streptococcus mutans (33), and others.…”

Section: Discussionmentioning

confidence: 99%

The Transcriptional Response of Lactobacillus sanfranciscensis DSM 20451 ^T and Its tcyB Mutant Lacking a Functional Cystine Transporter to Diamide Stress

Stetina

Behr

Vogel

2014

Appl Environ Microbiol

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As a result of its strong adaptation to wheat and rye sourdoughs, Lactobacillus sanfranciscensis has the smallest genome within the genus Lactobacillus. The concomitant absence of some important antioxidative enzymes and the inability to synthesize glutathione suggest a role of cystine transport in maintenance of an intracellular thiol balance. Diamide [synonym 1,1=-azobis(N,Ndimethylformamide)] disturbs intracellular and membrane thiol levels in oxidizing protein thiols depending on its initial concentration. In this study, RNA sequencing was used to reveal the transcriptional response of L. sanfranciscensis DSM 20451 T (wild type [WT]) and its ⌬tcyB mutant with a nonfunctional cystine transporter after thiol stress caused by diamide. Along with the different expression of genes involved in amino acid starvation, pyrimidine synthesis, and energy production, our results show that thiol stress in the wild type can be compensated through activation of diverse chaperones and proteases whereas the ⌬tcyB mutant shifts its metabolism in the direction of survival. Only a small set of genes are significantly differentially expressed between the wild type and the mutant. In the WT, mainly genes which are associated with a heat shock response are upregulated whereas glutamine import and synthesis genes are downregulated. In the ⌬tcyB mutant, the whole opp operon was more highly expressed, as well as a protein which probably includes enzymes for methionine transport. The two proteins encoded by spxA and nrdH, which are involved in direct or indirect oxidative stress responses, are also upregulated in the mutant. This work emphasizes that even in the absence of definitive antioxidative enzymes, bacteria with a small genome and a high frequency of gene inactivation and elimination use small molecules such as the cysteine/cystine couple to overcome potential cell damage resulting from oxidative stress. Lactobacillus sanfranciscensis is a highly adapted species, which can dominate wheat and rye sourdough microbiota. As a result of this strong adaptation, it has lost many genes present in other lactobacilli and therefore has the smallest genome currently known within the genus Lactobacillus (1). Aerobic growth in this strain results in a higher final cell yield and growth rate than does an anaerobic environment. While many lactic acid bacteria (LAB) produce reactive oxygen species (ROS), e.g., via the NADH oxidase 1 reaction, L. sanfranciscensis expresses NADH oxidase 2, which directly produces water and thus minimizes oxidative stress from that reaction (2). Still, the published genome data of L. sanfranciscensis TMW 1.1304 revealed enzymes which provide the strain's ability to cope with the formation of radicals. Genes thus far related to oxidative stress response include those for NADH oxidase, glutathione (GSH) reductase, glutaredoxin-like protein, two thioredoxin reductases, putative thioredoxin peroxidase, three thioredoxin-like proteins, and a cyst(e)ine transport protein (1).Cysteine acts in the catalytic site of enzyme...

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Transcriptional Analysis of Major Heat Shock Genes of Helicobacter pylori

Cited by 38 publications

References 45 publications

Characterization of the HspR-Mediated Stress Response inHelicobacter pylori

Characterization of the HspR-Mediated Stress Response inHelicobacter pylori

Transcriptional Regulation of Stress Response and Motility Functions inHelicobacter pyloriIs Mediated by HspR and HrcA

The Transcriptional Response of Lactobacillus sanfranciscensis DSM 20451 ^T and Its tcyB Mutant Lacking a Functional Cystine Transporter to Diamide Stress

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Transcriptional Analysis of Major Heat Shock Genes of Helicobacter pylori

Cited by 38 publications

References 45 publications

Characterization of the HspR-Mediated Stress Response inHelicobacter pylori

Characterization of the HspR-Mediated Stress Response inHelicobacter pylori

Transcriptional Regulation of Stress Response and Motility Functions inHelicobacter pyloriIs Mediated by HspR and HrcA

The Transcriptional Response of Lactobacillus sanfranciscensis DSM 20451 T and Its tcyB Mutant Lacking a Functional Cystine Transporter to Diamide Stress

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The Transcriptional Response of Lactobacillus sanfranciscensis DSM 20451 ^T and Its tcyB Mutant Lacking a Functional Cystine Transporter to Diamide Stress