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Gene Expression Profiling of Transcription Factors of Helicobacter pylori under Different Environmental Conditions.

ABSTRACT: Helicobacter pylori is a Gram-negative bacterium that colonizes the human gastric mucosa and causes peptic ulcers and gastric carcinoma. H. pylori strain 26695 has a small genome (1.67 Mb), which codes for few known transcriptional regulators that control bacterial metabolism and virulence. We analyzed by qRT-PCR the expression of 16 transcriptional regulators in H. pylori 26695, including the three sigma factors under different environmental conditions. When bacteria were exposed to acidic pH, urea, nickel, or iron, the sigma factors were differentially expressed with a particularly strong induction of fliA. The regulatory genes hrcA, hup, and crdR were highly induced in the presence of urea, nickel, and iron. In terms of biofilm formation fliA, flgR, hp1021, fur, nikR, and crdR were induced in sessile bacteria. Transcriptional expression levels of rpoD, flgR, hspR, hp1043, and cheY were increased in contact with AGS epithelial cells. Kanamycin, chloramphenicol, and tetracycline increased or decreased expression of regulatory genes, showing that these antibiotics affect the transcription of H. pylori. Our data indicate that environmental cues which may be present in the human stomach modulate H. pylori transcription.

SUBMITTER: De la Cruz MA

PROVIDER: S-EPMC5385360 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Gene Expression Profiling of Transcription Factors of Helicobacter pylori under Different Environmental Conditions.

De la Cruz Miguel A MA Ares Miguel A MA von Bargen Kristine K Panunzi Leonardo G LG Martínez-Cruz Jessica J Valdez-Salazar Hilda A HA Jiménez-Galicia César C Torres Javier J

Frontiers in microbiology 20170410

Helicobacter pylori is a Gram-negative bacterium that colonizes the human gastric mucosa and causes peptic ulcers and gastric carcinoma. H. pylori strain 26695 has a small genome (1.67 Mb), which codes for few known transcriptional regulators that control bacterial metabolism and virulence. We analyzed by qRT-PCR the expression of 16 transcriptional regulators in H. pylori 26695, including the three sigma factors under different environmental conditions. When bacteria were e ...[more]

PMID: 28443084

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Project description:BackgroundSerine protease HtrA exhibits both proteolytic and chaperone activities, which are involved in cellular protein quality control. Moreover, HtrA is an important virulence factor in many pathogens including Helicobacter pylori, for which the crucial stage of infection is the cleavage of E-cadherin and other cell-to-cell junction proteins.MethodsThe in vitro study of H. pylori HtrA (HtrAHp) chaperone activity was carried out using light scattering assays and investigation of lysozyme protein aggregates. We produced H. pylori ?htrA deletion and HtrAHp point mutants without proteolytic activity in strain N6 and investigated the survival of the bacteria under thermal, osmotic, acidic and general stress conditions as well as the presence of puromycin or metronidazole using serial dilution tests and disk diffusion method. The levels of cellular and secreted proteins were examined using biochemical fraction and Western blotting. We also studied the proteolytic activity of secreted HtrAHp using zymography and the enzymatic digestion of ?-casein. Finally, the consequences of E-cadherin cleavage were determined by immunofluorescence microscopy.ResultsWe demonstrate that HtrAHp displays chaperone activity that inhibits the aggregation of lysozyme and is stable under various pH and temperature conditions. Next, we could show that N6 expressing only HtrA chaperone activity grow well under thermal, pH and osmotic stress conditions, and in the presence of puromycin or metronidazole. In contrast, in the absence of the entire htrA gene the bacterium was more sensitive to a number of stresses. Analysing the level of cellular and secreted proteins, we noted that H. pylori lacking the proteolytic activity of HtrA display reduced levels of secreted HtrA. Moreover, we compared the amounts of secreted HtrA from several clinical H. pylori strains and digestion of ?-casein. We also demonstrated a significant effect of the HtrAHp variants during infection of human epithelial cells and for E-cadherin cleavage.ConclusionHere we identified the chaperone activity of the HtrAHp protein and have proven that this activity is important and sufficient for the survival of H. pylori under multiple stress conditions. We also pinpointed the importance of HtrAHp chaperone activity for E- cadherin degradation and therefore for the virulence of this eminent pathogen.

Dataset Information

Gene Expression Profiling of Transcription Factors of Helicobacter pylori under Different Environmental Conditions.

Publications

Gene Expression Profiling of Transcription Factors of <i>Helicobacter pylori</i> under Different Environmental Conditions.

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