IL-6 induces MUC4 expression through gp130/STAT3 pathway in gastric cancer cell lines

The gastric mucosal levels of the pro-inflammatory cytokine Interleukin 6 (IL-6) have been reported to be increased in Helicobacter pylori-infected subjects and, in gastric adenocarcinomas, the up-regulation of intestinal mucin genes (MUC2 and MUC4) has been detected. To analyse the regulatory effects of IL-6 on the activation of intestinal mucins, six gastric cancer cell lines were treated for different times with several concentrations of IL-6, and the expression of MUC2 and MUC4 was evaluated. IL-6 induced MUC4 expression, detected by quantitative RT-PCR, Western blot and immunofluorescence, and MUC2 expression was not affected. MUC4 mRNA levels decreased after blocking the gp130/STAT3 pathway at the level of the receptor, and at the level of STAT3 activation using the AG490 specific inhibitor. MUC4 presents two putative binding sites for STAT factors that may regulate MUC4 transcription after a pro-inflammatory stimulus as IL-6. By EMSA, ChIP and site-directed mutagenesis we show that STAT3 binds to a cis-element at -123/-115, that conveys IL-6 mediated up-regulation of MUC4 transcriptional activity. We also demonstrated that p-STAT3 binds to MUC4 promoter and a three-fold increase in p-STAT3 binding was observed after treating GP220 cells with IL-6. In conclusion, IL-6 treatment induced MUC4 expression through the gp130/STAT3 pathway, indicating the direct role of IL-6 on the activation of the intestinal mucin gene MUC4 in gastric cancer cells.