Regulation of p14ARF by Siva1 in H. pylori-infected gastric epithelial cells.

Abstract

243 Background: Helicobacter pylori ( H. pylori) is the strongest known risk factor for gastric cancer. Bacterial degradation of tumor suppressor proteins affect the host microbe’s interactions and host cellular response, which contribute to tumorigenesis. p14ARF, a crucial tumor suppressor protein that activates p53 protein under oncogenic stress plays a major role in oncogenic stress response (OSR) regulation. However, little is known about the mechanism of ARF and OSR regulation in H. pylori-infected gastric epithelial cells. Methods: The expression of p14ARF and cytotoxin-associated gene A (CagA) were analyzed in gastric cells co-cultured with H. pylori strains isolated from high-gastric risk and low-gastric risk areas by immunoblotting. To investigate the potential role of CagA in regulation of p14ARF, we employed isogenic cagA− and cagE− H. pylori mutants in gastric epithelial cells, and C57BL/6 mice (n = 10). We also analyzed the expression of Siva1 in human individual infected with cagA-positive (n = 13) and cagA-negative (n = 13) bacteria as well as uninfected human subjects (n = 6). siRNA was used to inhibit activity of Siva1 protein. Results: In this study, H. pylori strains expressing high levels of CagA virulence factor and associated with a higher gastric cancer risk more strongly suppress p14ARF compared with low-risk strains in vivo and in vitro. We found that degradation of p14ARF induced by CagA is mediated by E3 ubiquitin ligase Siva1, which works in concert with another E3 ubiquitin ligase TRIP12. Decreased expression of Siva1 protein and consequent up-regulation of p14ARF was also found in gastric mucosa of H. pylori-infected mice and human individuals. Tumorigenic strain 7.13 was more potent in upregulation of Siva1 and downregulation of p14ARF than non-tumorigenic strain B128. Inhibition of p14ARF protein by H. pylori causes inhibition of autophagy in infected cells. Conclusions: Our results provide first evidence that carcinogenic H. pylori strains significantly alter the host tumor suppressor protein p14ARF, leading to suppression of host OSR and autophagy, which may affect host-bacteria interactions and tumorigenic alteration in the stomach.