H. pylori (Hp) infects half of the human population and can cause gastritis, gastroduodenal ulcers, and gastric cancer. The ability of Hp to alter epithelial cell-cell adhesion may influence the clinical outcome of infection. Disruption of adherens junction (AJ) proteins E-cadherin and β–catenin have been implicated in carcinogenesis. The effects of Hp on the AJ and whether such changes may involve a known regulator of epithelial barrier function, myosin-light chain kinase (MLCK), are unclear. Aim: The aim of this study was to characterize the effects of Hp on the AJ in non-tumorigenic epithelial cell (SCBN) monolayers, and to examine the role of MLCK in these effects. Results: Western blotting revealed increased levels of phosphorylated myosin light chain (MLC) in cells challenged with Hp strain SS1 while myosin light chain-2 levels remained unchanged. Immunocytochemistry demonstrated disruption of membranous E-cadherin and redistribution of β–catenin in Hp infected monolayers, but not in controls. Immunoblotting revealed a time-dependent, Hp-induced decrease in E-cadherin expression. Pre-treatment with a selective MLCK inhibitor (ML-9) prevented these effects. Conclusion: Hp-induced phosphorylation of epithelial MLC is associated with AJ disruptions in non-tumorigenic epithelial cells. These alterations of E-cadherin and β–catenin are regulated, at least in part, by MLCK