GKN1–miR-185–DNMT1 Axis Suppresses Gastric Carcinogenesis through Regulation of Epigenetic Alteration and Cell Cycle

Abstract

Gastrokine 1 (GKN1) functions to protect the gastric antral mucosa and promotes healing by facilitating restoration and proliferation after injury. GKN1 is downregulated in Helicobacter pylori-infected gastric epithelial cells and loss of GKN1 expression is closely associated with gastric carcinogenesis, but underlying mechanisms of the tumor-suppressing effects of GKN1 remain largely unknown. AGS, MKN1, MKN28 gastric cancer cells and HFE-145 immortalized non-neoplastic gastric mucosal cells were transfected with GKN1 or shGKN1. We conducted molecular and functional studies of GKN1 and miR-185 and investigated the mechanisms of alteration. We also analyzed epigenetic alterations in 80 gastric cancer tissues. Restoration of GKN1 protein suppressed gastric cancer cell growth by inducing endogenous miR-185 that directly targets epigenetic effectors DNMT1 and EZH2 in gastric cancer cells. In addition, ectopic expression of GKN1 upregulated Tip60 and downregulated HDAC1 in an miR-185-independent manner, thereby inducing cell-cycle arrest by regulating cell-cycle proteins in gastric cancer cells. Notably, GKN1 expression was inversely correlated with DNMT1 and EZH2 expression in a subset of 80 gastric cancer tissues and various gastric cancer cell lines. Interestingly, it was found that GKN1 exerted a synergistic anti-cancerous effect with 5-fluorouracil on tumor cell growth, which suggests a possible therapeutic intervention method for gastric cancer. Our results show that GKN1 has an miR-185-dependent and -independent mechanism for chromatic and DNA epigenetic modification, thereby regulating the cell cycle. Thus, the loss of GKN1 function contributes to malignant transformation and proliferation of gastric epithelial cells in gastric carcinogenesis.