scholarly journals The Orphan Response Regulator HP1021 of Helicobacter pylori Regulates Transcription of a Gene Cluster Presumably Involved in Acetone Metabolism

2007 ◽  
Vol 189 (6) ◽  
pp. 2339-2349 ◽  
Author(s):  
Michael Pflock ◽  
Melanie Bathon ◽  
Jennifer Schär ◽  
Stefanie Müller ◽  
Hans Mollenkopf ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Gene Cluster ◽  
Response Regulator ◽  
Transcriptional Profiling ◽  
Acid Resistance ◽  
Housekeeping Genes ◽  
Open Reading Frames ◽  
Considerable Impact ◽  
Two Component Systems ◽  
H Pylori

ABSTRACT Helicobacter pylori is a gastric pathogen for which no nonhuman reservoir is known. In accordance with the tight adaptation to its unique habitat, the human stomach, H. pylori is endowed with a very restricted repertoire of regulatory proteins. Nevertheless, the three complete two-component systems of H. pylori were shown to be involved in the regulation of important virulence traits like motility and acid resistance and in the control of metal homeostasis. HP1021 is an orphan response regulator with an atypical receiver domain whose inactivation has a considerable impact on the growth of H. pylori. Here we report the identification of HP1021-regulated genes by whole-genome transcriptional profiling. We show that the transcription of the essential housekeeping genes nifS and nifU, which are required for the assembly of Fe-S clusters, is activated by HP1021. Furthermore, we demonstrate that the expression of a gene cluster comprising open reading frames hp0690 to hp0693 and hp0695 to hp0697 which is probably involved in acetone metabolism is strongly upregulated by HP1021. Evidence is provided for a direct regulation of the hp0695-to-hp0697 operon by the binding of HP1021 to its promoter region.

scholarly journals Characterization of the ArsRS Regulon of Helicobacter pylori, Involved in Acid Adaptation

2006 ◽  
Vol 188 (10) ◽  
pp. 3449-3462 ◽  
Author(s):  
Michael Pflock ◽  
Nadja Finsterer ◽  
Biju Joseph ◽  
Hans Mollenkopf ◽  
Thomas F. Meyer ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Response Regulator ◽  
Transcriptional Profiling ◽  
Acid Resistance ◽  
Two Component System ◽  
Gene Encoding ◽  
Acid Adaptation ◽  
Genes Encoding ◽  
H Pylori ◽  
A Site

ABSTRACT The human gastric pathogen Helicobacter pylori is extremely well adapted to the highly acidic conditions encountered in the stomach. The pronounced acid resistance of H. pylori relies mainly on the ammonia-producing enzyme urease; however, urease-independent mechanisms are likely to contribute to acid adaptation. Acid-responsive gene regulation is mediated at least in part by the ArsRS two-component system consisting of the essential OmpR-like response regulator ArsR and the nonessential cognate histidine kinase ArsS, whose autophosphorylation is triggered in response to low pH. In this study, by global transcriptional profiling of an ArsS-deficient H. pylori mutant grown at pH 5.0, we define the ArsR∼P-dependent regulon consisting of 109 genes, including the urease gene cluster, the genes encoding the aliphatic amidases AmiE and AmiF, and the rocF gene encoding arginase. We show that ArsR∼P controls the acid-induced transcription of amiE and amiF by binding to extended regions located upstream of the −10 box of the respective promoters. In contrast, transcription of rocF is repressed by ArsR∼P at neutral, acidic, and mildly alkaline pH via high-affinity binding of the response regulator to a site overlapping the promoter of the rocF gene.

scholarly journals Molecular Characterization of a Flagellar Export Locus of Helicobacter pylori

1999 ◽  
Vol 67 (5) ◽  
pp. 2060-2070 ◽  
Author(s):  
Steffen Porwollik ◽  
Brian Noonan ◽  
Paul W. O’Toole
Keyword(s):  
Helicobacter Pylori ◽  
Virulence Factor ◽  
Housekeeping Genes ◽  
Reading Frame ◽  
Reverse Transcription Pcr ◽  
Mutant Strains ◽  
H Pylori ◽  
Export Protein ◽  
Successful Colonization

ABSTRACT Motility of Helicobacter species has been shown to be essential for successful colonization of the host. We have investigated the organization of a flagellar export locus in Helicobacter pylori. A 7-kb fragment of the H. pylori CCUG 17874 genome was cloned and sequenced, revealing an operon comprising an open reading frame of unknown function (ORF03), essential housekeeping genes (ileS and murB), flagellar export genes (fliI and fliQ), and a homolog to a gene implicated in virulence factor transport in other pathogens (virB11). A promoter for this operon, showing similarity to the Escherichia coli ς70 consensus, was identified by primer extension. Cotranscription of the genes in the operon was demonstrated by reverse transcription-PCR, and transcription of virB11, fliI, fliQ, andmurB was detected in human or mouse biopsies obtained from infected hosts. The genetic organization of this locus was conserved in a panel of H. pylori clinical isolates. EngineeredfliI and fliQ mutant strains were completely aflagellate and nonmotile, whereas a virB11 mutant still produced flagella. The fliI and fliQ mutant strains produced reduced levels of flagellin and the hook protein FlgE. Production of OMP4, a member of the outer membrane protein family identified in H. pylori 26695, was reduced in both thevirB11 mutant and the fliI mutant, suggesting related functions of the virulence factor export protein (VirB11) and the flagellar export component (FliI).

scholarly journals Small Molecule Inhibitors of the Response Regulator ArsR Exhibit Bactericidal Activity against Helicobacter pylori

2020 ◽  
Vol 8 (4) ◽  
pp. 503 ◽  
Author(s):  
Andrés González ◽  
Javier Casado ◽  
Eduardo Chueca ◽  
Sandra Salillas ◽  
Adrián Velázquez-Campoy ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Antibiotic Resistance ◽  
High Throughput Screening ◽  
Lithocholic Acid ◽  
Response Regulator ◽  
Bacterial Pathogen ◽  
Antimicrobial Effect ◽  
Chemical Library ◽  
Secondary Bile Acid ◽  
H Pylori

Helicobacter pylori is considered the most prevalent bacterial pathogen in humans. The increasing antibiotic resistance evolved by this microorganism has raised alarm bells worldwide due to the significant reduction in the eradication rates of traditional standard therapies. A major challenge in this antibiotic resistance crisis is the identification of novel microbial targets whose inhibitors can overcome the currently circulating resistome. In the present study, we have validated the use of the essential response regulator ArsR as a novel and promising therapeutic target against H. pylori infections. A high-throughput screening of a repurposing chemical library using a fluorescence-based thermal shift assay identified several ArsR binders. At least four of these low-molecular weight compounds noticeably inhibited the DNA binding activity of ArsR and showed bactericidal effects against antibiotic-resistant strains of H. pylori. Among the ArsR inhibitors, a human secondary bile acid, lithocholic acid, quickly destroyed H. pylori cells and exhibited partial synergistic action in combination with clarithromycin or levofloxacin, while the antimicrobial effect of this compound against representative members of the normal human microbiota such as Escherichia coli and Staphylococcus epidermidis appeared irrelevant. Our results enhance the battery of novel therapeutic tools against refractory infections caused by multidrug-resistant H. pylori strains.

scholarly journals Identification of novel bacterial urease inhibitors through molecular shape and structure based virtual screening approaches

RSC Advances ◽  
2020 ◽  
Vol 10 (27) ◽  
pp. 16061-16070
Author(s):  
Muhammad Imran ◽  
Saba Waqar ◽  
Koji Ogata ◽  
Mahmood Ahmed ◽  
Zobia Noreen ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Virtual Screening ◽  
Acid Resistance ◽  
Molecular Shape ◽  
Urease Inhibitors ◽  
H Pylori ◽  
Screening Approaches

The enzyme urease is an essential colonizing factor of the notorious carcinogenic pathogen Helicobacter pylori (H. pylori), conferring acid resistance to the bacterium.

scholarly journals Colonization and Inflammation Deficiencies in Mongolian Gerbils Infected by Helicobacter pylori Chemotaxis Mutants

2005 ◽  
Vol 73 (3) ◽  
pp. 1820-1827 ◽  
Author(s):  
David J. McGee ◽  
Melanie L. Langford ◽  
Emily L. Watson ◽  
J. Elliot Carter ◽  
Yu-Ting Chen ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Immune Cell ◽  
Response Regulator ◽  
Critical Role ◽  
Mongolian Gerbils ◽  
Wild Type ◽  
In The Wild ◽  
H Pylori

ABSTRACT Helicobacter pylori causes disease in the human stomach and in mouse and gerbil stomach models. Previous results have shown that motility is critical for H. pylori to colonize mice, gerbils, and other animal models. The role of chemotaxis, however, in colonization and disease is less well understood. Two genes in the H. pylori chemotaxis pathway, cheY and tlpB, which encode the chemotaxis response regulator and a methyl-accepting chemoreceptor, respectively, were disrupted. The cheY mutation was complemented with a wild-type copy of cheY inserted into the chromosomal rdxA gene. The cheY mutant lost chemotaxis but retained motility, while all other strains were motile and chemotactic in vitro. These strains were inoculated into gerbils either alone or in combination with the wild-type strain, and colonization and inflammation were assessed. While the cheY mutant completely failed to colonize gerbil stomachs, the tlpB mutant colonized at levels similar to those of the wild type. With the tlpB mutant, there was a substantial decrease in inflammation in the gerbil stomach compared to that with the wild type. Furthermore, there were differences in the numbers of each immune cell in the tlpB-mutant-infected stomach: the ratio of lymphocytes to neutrophils was about 8 to 1 in the wild type but only about 1 to 1 in the mutant. These results suggest that the TlpB chemoreceptor plays an important role in the inflammatory response while the CheY chemotaxis regulator plays a critical role in initial colonization. Chemotaxis mutants may provide new insights into the steps involved in H. pylori pathogenesis.

scholarly journals Molecular Characterization of Two-Component Systems of Helicobacter pylori

2000 ◽  
Vol 182 (8) ◽  
pp. 2068-2076 ◽  
Author(s):  
Dagmar Beier ◽  
Rainer Frank
Keyword(s):  
Helicobacter Pylori ◽  
Histidine Kinase ◽  
Response Regulator ◽  
Response Regulators ◽  
Reading Frame ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

ABSTRACT Two-component systems are frequently involved in the adaptation of bacteria to changing environmental conditions at the level of transcriptional regulation. Here we report the characterization of members of the two-component systems of the gastric pathogenHelicobacter pylori deduced from the genome sequence of strain 26695. We demonstrate that the response regulators HP166, HP1043, and HP1021 have essential functions, as disruption of the corresponding genes is lethal for the bacteria, irrespective of the fact that HP1043 and HP1021 have nonconserved substitutions in crucial amino acids of their receiver domains. An analysis of the in vitro phosphorylation properties of the two-component proteins demonstrates that HP244-HP703 and HP165-HP166 are cognate histidine kinase-response regulator pairs. Furthermore, we provide evidence that the variability of the histidine kinase HP165 caused by a poly(C) tract of variable length close to the 3′ end of open reading frame 165/164 does not interfere with the kinase activity of the transmitter domain of HP165.

scholarly journals Helicobacter pylori Containing Only Cytoplasmic Urease Is Susceptible to Acid

1998 ◽  
Vol 66 (11) ◽  
pp. 5060-5066 ◽  
Author(s):  
Partha Krishnamurthy ◽  
Mary Parlow ◽  
Jason B. Zitzer ◽  
Nimish B. Vakil ◽  
Harry L. T. Mobley ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Urease Activity ◽  
Acid Resistance ◽  
Etiologic Agent ◽  
Stationary Phase Culture ◽  
Colonization Factor ◽  
Acid Environment ◽  
H Pylori

ABSTRACT Helicobacter pylori, an important etiologic agent in a variety of gastroduodenal diseases, produces large amounts of urease as an essential colonization factor. We have demonstrated previously that urease is located within the cytoplasm and on the surface of H. pylori both in vivo and in stationary-phase culture. The purpose of the present study was to assess the relative contributions of cytoplasmic and surface-localized urease to the ability of H. pylori to survive exposure to acid in the presence of urea. Toward this end, we compared the acid resistance in vitro of H. pylori cells which possessed only cytoplasmic urease to that of bacteria which possessed both cytoplasmic and surface-localized or extracellular urease. Bacteria with only cytoplasmic urease activity were generated by using freshly subcultured bacteria or by treating repeatedly subcultured H. pylori with flurofamide (1 μM), a potent, but poorly diffusible urease inhibitor. H. pyloriwith cytoplasmic and surface-located urease activity survived in an acid environment when 5 mM urea was present. In contrast, H. pylori with only cytoplasmic urease shows significantly reduced survival when exposed to acid in the presence of 5 mM urea. Similarly,Escherichia coli SE5000 expressing H. pyloriurease and the Ni2+ transport protein NixA, which expresses cytoplasmic urease activity at levels similar to those in wild-typeH. pylori, survived minimally when exposed to acid in the presence of 5 to 50 mM urea. We conclude that cytoplasmic urease activity alone is not sufficient (although cytoplasmic urease activity is likely to be necessary) to allow survival of H. pyloriin acid; the activity of surface-localized urease is essential for resistance of H. pylori to acid under the assay conditions used. Therefore, the mechanism whereby urease becomes associated with the surface of H. pylori, which involves release of the enzyme from bacteria due to autolysis followed by adsorption of the enzyme to the surface of intact bacteria (“altruistic autolysis”), is essential for survival of H. pylori in an acid environment. The ability of H. pylori to survive exposure to low pH is likely to depend on a combination of both cytoplasmic and surface-associated urease activities.

scholarly journals Systematic Identification of Selective Essential Genes in Helicobacter pylori by Genome Prioritization and Allelic Replacement Mutagenesis

2001 ◽  
Vol 183 (4) ◽  
pp. 1259-1268 ◽  
Author(s):  
Alison F. Chalker ◽  
Heather W. Minehart ◽  
Nicky J. Hughes ◽  
Kristin K. Koretke ◽  
Michael A. Lonetto ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Essential Genes ◽  
Open Reading Frames ◽  
Comparative Genomic ◽  
Allelic Replacement ◽  
Genomic Approach ◽  
H Pylori ◽  
Systematic Identification ◽  
Reading Frames

ABSTRACT A comparative genomic approach was used to identifyHelicobacter pylori 26695 open reading frames (ORFs) which are conserved in H. pylori J99 but highly diverged in other eubacteria. A survey of selected pathways of central intermediary metabolism was also carried out, and genes with a potentially selective role in H. pylori were identified. Forty-five ORFs identified in these two analyses were screened using a rapid vector-free allelic replacement mutagenesis technique, and 33 were shown to be essential in vitro. Notably, 13 ORFs gave essentiality results which are unexpected in view of their known or proposed functions, and phylogenetic analysis was used to investigate the annotation of 7 such ORFs which are highly diverged. We propose that the products of a number of these H. pylori-specific essential genes may be suitable targets for novel anti-H. pylori therapies.

scholarly journals Transcriptional profiling of Helicobacter pylori Fur- and iron-regulated gene expression

Microbiology ◽  
2005 ◽  
Vol 151 (2) ◽  
pp. 533-546 ◽  
Author(s):  
Florian D. Ernst ◽  
Stefan Bereswill ◽  
Barbara Waidner ◽  
Jeroen Stoof ◽  
Ulrike Mäder ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Iron Homeostasis ◽  
Transcriptional Profiling ◽  
Northern Hybridization ◽  
Intracellular Iron ◽  
Iron Storage ◽  
Gastric Colonization ◽  
Genes Encoding ◽  
H Pylori ◽  
Regulated Gene Expression

Intracellular iron homeostasis is a necessity for almost all living organisms, since both iron restriction and iron overload can result in cell death. The ferric uptake regulator protein, Fur, controls iron homeostasis in most Gram-negative bacteria. In the human gastric pathogen Helicobacter pylori, Fur is thought to have acquired extra functions to compensate for the relative paucity of regulatory genes. To identify H. pylori genes regulated by iron and Fur, we used DNA array-based transcriptional profiling with RNA isolated from H. pylori 26695 wild-type and fur mutant cells grown in iron-restricted and iron-replete conditions. Sixteen genes encoding proteins involved in metal metabolism, nitrogen metabolism, motility, cell wall synthesis and cofactor synthesis displayed iron-dependent Fur-repressed expression. Conversely, 16 genes encoding proteins involved in iron storage, respiration, energy metabolism, chemotaxis, and oxygen scavenging displayed iron-induced Fur-dependent expression. Several Fur-regulated genes have been previously shown to be essential for acid resistance or gastric colonization in animal models, such as those encoding the hydrogenase and superoxide dismutase enzymes. Overall, there was a partial overlap between the sets of genes regulated by Fur and those previously identified as growth-phase, iron or acid regulated. Regulatory patterns were confirmed for five selected genes using Northern hybridization. In conclusion, H. pylori Fur is a versatile regulator involved in many pathways essential for gastric colonization. These findings further delineate the central role of Fur in regulating the unique capacity of H. pylori to colonize the human stomach.

scholarly journals The Helicobacter pylori Homologue of the Ferric Uptake Regulator Is Involved in Acid Resistance

2002 ◽  
Vol 70 (2) ◽  
pp. 606-611 ◽  
Author(s):  
Jetta J. E. Bijlsma ◽  
Barbara Waidner ◽  
Arnoud H. M. van Vliet ◽  
Nicky J. Hughes ◽  
Stephanie Häg ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Iron Homeostasis ◽  
Regulatory Protein ◽  
Acid Resistance ◽  
Low Ph ◽  
Main Function ◽  
Ferric Uptake Regulator ◽  
Acidic Ph ◽  
Human Pathogen ◽  
H Pylori

ABSTRACT The only known niche of the human pathogen Helicobacter pylori is the gastric mucosa, where large fluctuations of pH occur, indicating that the bacterial response and resistance to acid are important for successful colonization. One of the few regulatory proteins in the H. pylori genome is a homologue of the ferric uptake regulator (Fur). In most bacteria, the main function of Fur is the regulation of iron homeostasis. However, in Salmonella enterica serovar Typhimurium, Fur also plays an important role in acid resistance. In this study, we determined the role of the H. pylori Fur homologue in acid resistance. Isogenic fur mutants were generated in three H. pylori strains (1061, 26695, and NCTC 11638). At pH 7 there was no difference between the growth rates of mutants and the parent strains. Under acidic conditions, growth of the fur mutants was severely impaired. No differences were observed between the survival of the fur mutant and parent strain 1061 after acid shock. Addition of extra iron or removal of iron from the growth medium did not improve the growth of the fur mutant at acidic pH. This indicates that the phenotype of the fur mutant at low pH was not due to increased iron sensitivity. Transcription of fur was repressed in response to low pH. From this we conclude that Fur is involved in the growth at acidic pH of H. pylori; as such, it is the first regulatory protein implicated in the acid resistance of this important human pathogen.

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