Identification of the RNA Pyrophosphohydrolase RppH of Helicobacter pylori and Global Analysis of Its RNA Targets
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ABSTRACT:
ORGANISM(S): Helicobacter pylori 26695
PROVIDER: GSE86943 | GEO | 2017/01/12
SECONDARY ACCESSION(S): PRJNA343039
REPOSITORIES: GEO
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Project description:Purpose: Next-generation sequencing (NGS) was used to analyze pH-responsive gene expression in H. pylori. The goals of this study are to compare H. pylori pH-responsive gene expression in H. pylori strain 26695 and 26695 dervatives containing mutations to the ArsRS two component system.
2021-01-19 | GSE165055 | GEO
Project description:Purpose: Next-generation sequencing (NGS) was used to analyze pH-responsive gene expression in H. pylori. The goals of this study are to compare H. pylori pH-responsive gene expression in H. pylori strain 26695 and 26695 dervatives containing mutations to the ArsS, CrdS and FlgS sensor kinases.
2021-01-19 | GSE165056 | GEO
Project description:RNA degradation is crucial for regulating gene expression in all organisms. Like the decapping of eukaryotic mRNAs, the conversion of the 5'-terminal triphosphate of bacterial transcripts to a monophosphate can trigger RNA decay by exposing the transcript to attack by 5'-monophosphate-dependent ribonucleases. In both biological realms, this deprotection step is catalyzed by members of the Nudix hydrolase family. The genome of the gastric pathogen Helicobacter pylori, a Gram-negative epsilonproteobacterium, encodes two proteins resembling Nudix enzymes. Here we present evidence that one of them, HP1228 (renamed HpRppH), is an RNA pyrophosphohydrolase that triggers RNA degradation in H. pylori, whereas the other, HP0507, lacks such activity. In vitro, HpRppH converts RNA 5'-triphosphates and diphosphates to monophosphates. It requires at least two unpaired nucleotides at the 5' end of its substrates and prefers three or more but has only modest sequence preferences. The influence of HpRppH on RNA degradation in vivo was examined by using RNA-seq to search the H. pylori transcriptome for RNAs whose 5'-phosphorylation state and cellular concentration are governed by this enzyme. Analysis of cDNA libraries specific for transcripts bearing a 5'-triphosphate and/or monophosphate revealed at least 63 potential HpRppH targets. These included mRNAs and sRNAs, several of which were validated individually by half-life measurements and quantification of their 5'-terminal phosphorylation state in wild-type and mutant cells. These findings demonstrate an important role for RppH in post-transcriptional gene regulation in pathogenic Epsilonproteobacteria and suggest a possible basis for the phenotypes of H. pylori mutants lacking this enzyme.
| S-EPMC5290964 | biostudies-literature
Project description:In this study transcriptional start sites (TSS) for H. pylori 26695 were determined
2015-07-01 | GSE67564 | GEO
Project description:Identification of the RNA Pyrophosphohydrolase RppH of Helicobacter pylori and Global Analysis of Its RNA Targets
| PRJNA343039 | ENA
Project description:In this study, we tested the hypothesis that high salt concentrations might alter gene expression in H. pylori. Analysis here provides into the role that salt may play in H. pylori pathogenesis. . H. pylori strain 26695 (SC#7) was grown for 15 h in BB-FBS medium containing 0.5% sodium chloride (i.e. BB-FBS-0.5%) The bacterial cells were then grown for 10 h, in fresh brucella broth at either 0.25% or 1.5% salt, harvested by centrifugation, and frozen at -80oC. RNA isolated was reverse transcribed and used to probe H. pylori Panorama arrays
2010-06-25 | E-GEOD-7183 | biostudies-arrayexpress
Project description:The genome of the gastric pathogen Helicobacter pylori harbors a remarkably low number of regulatory genes, including three and five open reading frames encoding two-component histidine kinases and response regulators, respectively, which are putatively involved in transcriptional regulation. Inactivation of the response regulator gene hp1021 resulted in a severe growth defect, as indicated by a small-colony phenotype. Recently we found that phosphorylation of the receiver domain HP1021 is not needed for its response regulator function and may not occur at all. No target genes have been identified so far. In this study we define the HP1021-dependent regulon consisting of 79 genes (51 activated, 28 repressed) by global transcriptional profiling of an HP1021-deficient H. pylori mutant. Transcriptome analyses were performed using a whole-genome microarray containing 1649 PCR products generated with specific primer pairs derived from the genome sequences of H. pylori 26695 (Tomb et al., 1997. The complete genome sequence of the gastric pathogen Helicobacter pylori. Nature 388:539-547) and J99 (Alm et al., 1999. Genomic-sequence comparison of two unrelated isolates of the human gastric pathogen Helicobacter pylori. Nature 397:176-180) which were spotted in duplicate. Microarrays were produced as described by Gressmann et al. (Gressmann et al., 2005. Gain and loss of multiple genes during the evolution of Helicobacter pylori. PLoS Genet 1(4):e43). To determine genes which are differentially expressed in the HP1021-deficient mutant 26695/1021::km, cDNA was prepared from RNA extracted from H. pylori 26695 WT and 26695/1021::km. A total of eight RNA samples from two independent RNA preparations from strain 26695 WT and 26695/HP1021::km, respectively, was used for cDNA labelling und hybridisation. Dye reversal colour swaps were performed as follows: one cDNA sample was generated using Cy3-dCTP and the other using Cy5-dCTP resulting in four labelled cDNAs per colour swap. Cy5-dCTP and Cy3-dCTP labelled cDNAs were combined and hybridized to the H. pylori microarray. The slides were scanned using ScanArray HT and analysed by using the ScanArray express software (Perkin Elmer, version 3.0). Spots were flagged and eliminated from analysis when the signal to background ratio was less then three or in obvious instances of high background or stray fluorescent signals. Median intensities of spots were background corrected and differences in dye bias were normalized by using the LOWESS algorithm (Yang et al., 2002. Normalization for cDNA microarray data: a robuste composite method addressing single and multiple slide systematic variation. Nucleic Acid Res. 30:e15). The signal ratios as measure of differential expression between the red and green channels were obtained from processed signal intensities. Ratios were further analysed with Microsoft Excel (Microsoft) and SAM software for statistic significance (Tusher et al., 2001. Significance analysis of microarrays applied to the ionizing radiation response. Proc. Natl. Acad. Sci. USA 98:5116-5121). To determine the significance of differential expression RNA was isolated from the H. pylori 26695 WT grown in BHI broth, and 20 µg of this RNA were labelled either with Cy3-dCTP or with Cy5-dCTP. The two cDNA probes generated were hybridized onto the same slide, and the data were analysed as mentioned above. Signal ratios < 0.5 and > 2.0 were analyzed further.
2010-07-01 | E-GEOD-5971 | biostudies-arrayexpress
Project description:We engineered a derivative of H. pylori strain 26695 that expresses an N-terminal DDK-tagged version of LolF from the endogenous locus. As controls, we also engineered strains to express DDK-tagged versions of two unrelated inner membrane proteins ExbD2 and FrdC. The tagged proteins were affinity purified and co-purifying proteins were identified by LC/MS-MS. We then performed an unbiased analysis using SAINTexpress (Significance Analysis of INTeractome) that included data from affinity purifications of test (i.e., LolF-DDK) and control (i.e., ExbD2-DDK and FrdC-DDK) bait proteins. This led to identification of HP0179 as an interaction partner of LolF. We then engineered a derivative of H. pylori strain 26695 that expresses a C-terminal HA-tagged version of HP0179. As controls, additional strains were generated that express HA-tagged versions of two unrelated proteins HP0838 and CagF. he tagged proteins were affinity purified and co-purifying proteins were identified by LC/MS-MS. We then performed an unbiased analysis using SAINTexpress that included data from affinity purifications of test (i.e., HP0179-HA) and control (i.e., HP0838-HA and CagF-HA) bait proteins. This led to identification of LolF as an interaction partner of HP0179.
2023-01-31 | MSV000091176 | MassIVE
Project description:The present study was aimed at analyzing (i) the biological cost of RNA polymerase (rpoB) mutations conferring rifampin resistance on H.pylori, (ii) the relationship between the cost of rpoB mutations and the chromosomal mutaion, (iii) the relationship between the cost of rpoB mutations and the transcription profile of sensitive and resistantrif strains of H.pylori (iv) and rpoB mutations in view of the possible fitness burden associated with resistance to another antibiotics. H.pylori reference strain 26695 was routinely maintained on Columbia agar plates and H. pylori-selective antibiotic mix Dent. Liquid culture was grown in BHI broth. Both plates and broth cultures were incubated at 37C under atmosphere enriched with 5% CO2 for 2-3 days . Mutant strains were selected by culturing H. pylori 26695 on selective plates containing rifampicin. In 5 days resistant colonies were picked up and passed under rifampicin pressure. RNA isolated was reverse transcribed and used to probe H. pylori home-made arrays
2010-11-30 | E-GEOD-25333 | biostudies-arrayexpress