Dataset Information

The Middle Fragment of Helicobacter pylori CagA Induces Actin Rearrangement and Triggers Its Own Uptake into Gastric Epithelial Cells.

ABSTRACT: Cytotoxin-associated gene product A (CagA) is a major virulence factor secreted by Helicobacter pylori. CagA activity in the gastric epithelium is associated with higher risk of gastric cancer development. Bacterial type IV secretion system (T4SS)-mediated translocation of CagA into the cytosol of human epithelial cells occurs via a poorly understood mechanism that requires CagA interaction with the host membrane lipid phosphatidylserine (PS) and host cell receptor integrin ????. Here we have characterized the isolated recombinant middle fragment of CagA (CagA-M) that contains the positively-charged PS-binding region (aa 613-636) and a putative ?? integrin binding site, but lacks the EPIYA region, secretion signal peptide and the CagA multimerization motif. We show that CagA-M, when immobilized on latex beads, is capable of binding to, and triggering its own uptake into, gastric epithelial cells in the absence of infection with cagA-positive H. pylori. Using site-directed mutagenesis, fluorescent and electron microscopy, and highly-specific inhibitors, we demonstrate that the cell-binding and endocytosis-like internalization of CagA-M are dependent on (1) binding to PS; (2) ?? integrin activity; and (3) actin dynamics. Interaction of CagA-M with the host cells is accompanied by the development of long filopodia-like protrusions (macrospikes). This novel morphology is different from the hummingbird phenotype induced by the translocation of full-length CagA. The determinants within CagA-M and within the host that are important for endocytosis-like internalization into host cells are very similar to those observed for T4SS-mediated internalization of full-length CagA, suggesting that the latter may involve an endocytic pathway.

SUBMITTER: Tohidpour A

PROVIDER: S-EPMC5577571 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

The Middle Fragment of Helicobacter pylori CagA Induces Actin Rearrangement and Triggers Its Own Uptake into Gastric Epithelial Cells.

Tohidpour Abolghasem A Gorrell Rebecca J RJ Roujeinikova Anna A Kwok Terry T

Toxins 20170728 8

Cytotoxin-associated gene product A (CagA) is a major virulence factor secreted by <i>Helicobacter pylori</i>. CagA activity in the gastric epithelium is associated with higher risk of gastric cancer development. Bacterial type IV secretion system (T4SS)-mediated translocation of CagA into the cytosol of human epithelial cells occurs via a poorly understood mechanism that requires CagA interaction with the host membrane lipid phosphatidylserine (PS) and host cell receptor integrin α₅β₁. Here we ...[more]

PMID: 28788072

Similar Datasets

Project description:BACKGROUND: Most of what is known about the Helicobacter pylori (H. pylori) cytotoxin, CagA, pertains to a much-vaunted role as a determinant of gastric inflammation and cancer. Little attention has been devoted to potential roles of CagA in the majority of H. pylori infected individuals not showing oncogenic progression, particularly in relation to host tolerance. Regenerating islet-derived (REG)3? encodes a secreted C-type lectin that exerts direct bactericidal activity against Gram-positive bacteria in the intestine. Here, we extend this paradigm of lectin-mediated innate immunity, showing that REG3? expression is triggered by CagA in the H. pylori-infected stomach. METHODOLOGY/PRINCIPAL FINDINGS: In human gastric mucosal tissues, REG3? expression was significantly increased in CagA-positive, compared to CagA-negative H. pylori infected individuals. Using transfected CagA-inducible gastric MKN28 cells, we recapitulated REG3? induction in vitro, also showing that tyrosine phosphorylated, not unphosphorylated CagA triggers REG3? transcription. In concert with induced REG3?, pro-inflammatory signalling downstream of the gp130 cytokine co-receptor via the signal transducer and activator of transcription (STAT)3 and transcription of two cognate ligands, interleukin(IL)-11 and IL-6, were significantly increased. Exogenous IL-11, but not IL-6, directly stimulated STAT3 activation and REG3? transcription. STAT3 siRNA knockdown or IL-11 receptor blockade respectively abrogated or subdued CagA-dependent REG3? mRNA induction, thus demonstrating a requirement for uncompromised signalling via the IL-11/STAT3 pathway. Inhibition of the gp130-related SHP2-(Ras)-ERK pathway did not affect CagA-dependent REG3? induction, but strengthened STAT3 activation as well as augmenting transcription of mucosal innate immune regulators, IL-6, IL-8 and interferon-response factor (IRF)1. CONCLUSIONS/SIGNIFICANCE: Our results support a model of CagA-directed REG3? expression in gastric epithelial cells via activation of the IL-11/gp130/STAT3 pathway. This response might allow Gram-negative H. pylori to manipulate host immunity to favour its own survival, by reducing the fitness of co-habiting Gram-positive bacteria with which it competes for resources in the gastric mucosal niche.

Project description:BackgroundAs a marker of Helicobacter pylori, Cytotoxin-associated gene A (cagA) has been revealed to be the major virulence factor causing gastroduodenal diseases. However, the molecular mechanisms that underlie the development of different gastroduodenal diseases caused by cagA-positive H. pylori infection remain unknown. Current studies are limited to the evaluation of the correlation between diseases and the number of Glu-Pro-Ile-Tyr-Ala (EPIYA) motifs in the CagA strain. To further understand the relationship between CagA sequence and its virulence to gastric cancer, we proposed a systematic entropy-based approach to identify the cancer-related residues in the intervening regions of CagA and employed a supervised machine learning method for cancer and non-cancer cases classification.MethodologyAn entropy-based calculation was used to detect key residues of CagA intervening sequences as the gastric cancer biomarker. For each residue, both combinatorial entropy and background entropy were calculated, and the entropy difference was used as the criterion for feature residue selection. The feature values were then fed into Support Vector Machines (SVM) with the Radial Basis Function (RBF) kernel, and two parameters were tuned to obtain the optimal F value by using grid search. Two other popular sequence classification methods, the BLAST and HMMER, were also applied to the same data for comparison.ConclusionOur method achieved 76% and 71% classification accuracy for Western and East Asian subtypes, respectively, which performed significantly better than BLAST and HMMER. This research indicates that small variations of amino acids in those important residues might lead to the virulence variance of CagA strains resulting in different gastroduodenal diseases. This study provides not only a useful tool to predict the correlation between the novel CagA strain and diseases, but also a general new framework for detecting biological sequence biomarkers in population studies.

Dataset Information

The Middle Fragment of Helicobacter pylori CagA Induces Actin Rearrangement and Triggers Its Own Uptake into Gastric Epithelial Cells.

Publications

The Middle Fragment of Helicobacter pylori CagA Induces Actin Rearrangement and Triggers Its Own Uptake into Gastric Epithelial Cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets