The Complete Genome Sequence of<i>Helicobacter pylori</i>Strain G27

Baltrus, David A.; Amieva, Manuel R.; Covacci, Antonello; Lowe, Todd M.; Merrell, D. Scott; Ottemann, Karen M.; Stein, Markus; Salama, Nina R.; Guillemin, Karen

doi:10.1128/jb.01416-08

Cited by 182 publications

(169 citation statements)

References 14 publications

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“…We utilized the numerous previous transcriptional and proteomic studies regarding H. pylori Fur regulation (7,8,23,(32)(33)(34) to identify potential uncharacterized apo-Fur-repressed gene targets. Analysis of those studies showed considerable variability; thus, we imposed the following criteria to narrow the list of genes for study: (I) changes in expression (mRNA or protein) were consistent with apo Fur regulation (i.e., a Fur-dependent decrease in expression in the WT background upon iron chelation); (ii) the genes were present in strains G27 (35), 26695 (13), J99 (36), and HPAG1 (37), which are each well-characterized H. pylori strains; (iii) the genes did not appear to be essential (38); and (iv) the changes in expression exhibited in response to iron chelation and/or fur deletion were substantial (defined as a minimum 2-fold change). Based on these criteria, the cytochrome c 553 gene (HP1227) (23), hydA (HP0631) (7,23,32,34), and serB (HP0652) (23,32,33) were chosen for further study.…”

Section: Resultsmentioning

confidence: 99%

Identification and Characterization of Novel Helicobacter pylori apo-Fur-Regulated Target Genes

et al. 2013

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cIn Helicobacter pylori, the ferric uptake regulator (Fur) has evolved additional regulatory functions not seen in other bacteria; it can repress and activate different groups of genes in both its iron-bound and apo forms. Because little is understood about the process of apo-Fur repression and because only two apo-Fur-repressed genes (pfr and sodB) have previously been identified, we sought to expand our understanding of this type of regulation. Utilizing published genomic studies, we selected three potential new apo-Fur-regulated gene targets: serB, hydA, and the cytochrome c 553 gene. Transcriptional analyses confirmed Fur-dependent repression of these genes in the absence of iron, as well as derepression in the absence of Fur. Binding studies showed that apo-Fur directly interacted with the suspected hydA and cytochrome c 553 promoters but not that of serB, which was subsequently shown to be cotranscribed with pfr; apo-Fur-dependent regulation occurred at the pfr promoter. Alignments of apo-regulated promoter regions revealed a conserved, 6-bp consensus sequence (AAATGA). DNase I footprinting showed that this sequence lies within the protected regions of the pfr and hydA promoters. Moreover, mutation of the sequence in the pfr promoter abrogated Fur binding and DNase protection. Likewise, fluorescence anisotropy studies and binding studies with mutated consensus sequences showed that the sequence was important for apo-Fur binding to the pfr promoter. Together these studies expand the known apo-Fur regulon in H. pylori and characterize the first reported apo-Fur box sequence.

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Section: Resultsmentioning

confidence: 99%

Identification and Characterization of Novel Helicobacter pylori apo-Fur-Regulated Target Genes

et al. 2013

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“…In the final step, the upstream Kan r PCR product was fused the 5= end of the arsR construct by SOE PCR using Up1F and Dn4R. The resulting SOE PCR product was used to transform G27 (DSM1) (51). Transformants were selected on 25 g/ml Kan. Dn1F and Dn4R primers were used to PCR amplify Kan-resistant colonies, and the resulting amplicons were screened by restriction digestion with Apo1; the G¡A transition introduces an Apo1 site.…”

Section: Methodsmentioning

confidence: 99%

Characterization of Key Helicobacter pylori Regulators Identifies a Role for ArsRS in Biofilm Formation

et al. 2016

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Helicobacter pylori must be able to rapidly respond to fluctuating conditions within the stomach. Despite this need for constant adaptation, H. pylori encodes few regulatory proteins. Of the identified regulators, the ferric uptake regulator (Fur), the nickel response regulator (NikR), and the two-component acid response system (ArsRS) are each paramount to the success of this pathogen. While numerous studies have individually examined these regulatory proteins, little is known about their combined effect. Therefore, we constructed a series of isogenic mutant strains that contained all possible single, double, and triple regulatory mutations in Fur, NikR, and ArsS. A growth curve analysis revealed minor variation in growth kinetics across the strains; these were most pronounced in the triple mutant and in strains lacking ArsS. Visual analysis showed that strains lacking ArsS formed large aggregates and a biofilm-like matrix at the air-liquid interface. Biofilm quantification using crystal violet assays and visualization via scanning electron microscopy (SEM) showed that all strains lacking ArsS or containing a nonphosphorylatable form of ArsR (ArsR-D52N mutant) formed significantly more biofilm than the wild-type strain. Molecular characterization of biofilm formation showed that strains containing mutations in the ArsRS pathway displayed increased levels of cell aggregation and adherence, both of which are key to biofilm development. Furthermore, SEM analysis revealed prevalent coccoid cells and extracellular matrix formation in the ArsR-D52N, ⌬nikR ⌬arsS, and ⌬fur ⌬nikR ⌬arsS mutant strains, suggesting that these strains may have an exacerbated stress response that further contributes to biofilm formation. Thus, H. pylori ArsRS has a previously unrecognized role in biofilm formation. IMPORTANCEDespite a paucity of regulatory proteins, adaptation is key to the survival of H. pylori within the stomach. While prior studies have focused on individual regulatory proteins, such as Fur, NikR, and ArsRS, few studies have examined the combined effect of these factors. Analysis of isogenic mutant strains that contained all possible single, double, and triple regulatory mutations in Fur, NikR, and ArsS revealed a previously unrecognized role for the acid-responsive two-component system ArsRS in biofilm formation.

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“…The strains of H. pylori used in this study included the sequenced strains 26695 (26), J99 (27), and G27 (28), as well as a babA knockout mutant of G27, strain G27⌬babA (a gift from Steffen Backert). H. pylori was routinely cultured at 37°C on Columbia blood agar base (Oxoid) containing 7% (vol/vol) defibrinated horse blood under microaerophilic conditions generated using CampyGen gas packs (Oxoid).…”

Section: Methodsmentioning

confidence: 99%

Divergent Mechanisms of Interaction of Helicobacter pylori and Campylobacter jejuni with Mucus and Mucins

et al. 2013

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f Helicobacter pylori and Campylobacter jejuni colonize the stomach and intestinal mucus, respectively. Using a combination of mucus-secreting cells, purified mucins, and a novel mucin microarray platform, we examined the interactions of these two organisms with mucus and mucins. H. pylori and C. jejuni bound to distinctly different mucins. C. jejuni displayed a striking tropism for chicken gastrointestinal mucins compared to mucins from other animals and preferentially bound mucins from specific avian intestinal sites (in order of descending preference: the large intestine, proximal small intestine, and cecum). H. pylori bound to a number of animal mucins, including porcine stomach mucin, but with less avidity than that of C. jejuni for chicken mucin. The strengths of interaction of various wild-type strains of H. pylori with different animal mucins were comparable, even though they did not all express the same adhesins. The production of mucus by HT29-MTX-E12 cells promoted higher levels of infection by C. jejuni and H. pylori than those for the non-mucus-producing parental cell lines. Both C. jejuni and H. pylori bound to HT29-MTX-E12 mucus, and while both organisms bound to glycosylated epitopes in the glycolipid fraction of the mucus, only C. jejuni bound to purified mucin. This study highlights the role of mucus in promoting bacterial infection and emphasizes the potential for even closely related bacteria to interact with mucus in different ways to establish successful infections.

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The Complete Genome Sequence ofHelicobacter pyloriStrain G27

Cited by 182 publications

References 14 publications

Identification and Characterization of Novel Helicobacter pylori apo-Fur-Regulated Target Genes

Identification and Characterization of Novel Helicobacter pylori apo-Fur-Regulated Target Genes

Characterization of Key Helicobacter pylori Regulators Identifies a Role for ArsRS in Biofilm Formation

Divergent Mechanisms of Interaction of Helicobacter pylori and Campylobacter jejuni with Mucus and Mucins

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