Helicobacter pylori induced transactivation of SRE and AP-1 through the ERK signalling pathway in gastric cancer cells

Background and aims-Helicobacter pylori infection induces expression of pro inflammatory cytokines such as interleukin (IL)-8 and tumour necrosis facto r a (TNF-alpha) in gastric mucosa, and their genes have AP-1 binding sites in the promoter region. c-Fos is important for transactivation of AP-1 whic h has SRE in the promoter region. We conducted this study to confirm H pylo ri induced transactivation of these binding sites. Methods-Transactivation of SRE and AP-1 was evaluated in human gastric canc er cells TMK1 and MKN45 by luciferase reporter assay in transient transfect ion. We compared the effects of coculture with four H pylori strains, a cag pathogenicity island (PAI) positive strain TN2, its isogenic vacA negative (TN2-Delta vacA) or cagE negative (TN2-Delta cagE) mutants, and a cag PAI negative clinical isolate T68. Phosphorylation of ERK1/2, JNK, and c-Jun wa s measured by immunoblot, induction of IL-8 secretion by ELISA, and the eff ects of MEK by inhibitor U0126. Results-Both SRE and AP-1 were transactivated by coculture with TN2. Althou gh TN2-Delta vacA induced comparable transactivation, TN2-Delta cagE and T6 8 showed decreased transactivation of SRE (65% and 51%) and AP-1 (71% and 5 4%, respectively, of TN2). Heat killed TN2 or indirect contact using a perm eable membrane inhibited transactivation. Levels of phosphorylated ERK1/2, JNK, and c-Jun were increased by coculture with TN2. MEK inhibitor U0126 re duced TN2 induced transactivation of SRE and AP1, as well as secretion of I L-8, by 83%, 87%, and 53%, respectively, of TN2. Conclusions-Transactivation of SRE and AP-1, through ERK/MAPK and JNK/SAPK cascades, respectively, was found in gastric cancer cells cocultured with P i pylori. Direct contact with viable bacteria possessing intact cag PAI is a prerequisite for the onset of intracellular signalling leading to AP-1 tr ansactivation.