RUNX3 is a tumor suppressor and belongs to a family of transcription factors RUNX that has been reported to be inactivated in more than 80% of gastric cancers.In this study, we demonstrated that Akt transcription can be suppressed by RUNX3 in that two RUNX3 binding sites on Akt promoter were identified by reporter and ChIP assay.Together with that up-regulation of both Akt mRNA and protein was shown in the clinical RUNX3-1oss gastric tumor tissues examined, these results suggested that loss of RUNX3 may cause AKT over-expression in gastric cancer progression.In addition, our results showed that the cell proliferation rate was significantly inhibited in a dose-dependent manner when reinforcing the RUNX3 expression in RUNX-3 loss AGS cells.The flow cytometric analysis further showed that reinforce of the RUNX3 expression in AGS cells can induce the cell cycle arrest at G1/G0 phase.Since the protein levels ofAkt,-catenin and cyclin D1 were down-regulated in RUNX3 overexpressing AGS cells, we then hypothesized that the tumor suppression activity of RUNX3 may through down-regulating Akt/β-catenir/cyclin D1 signaling pathway.Interestingly, the growth inhibition of AGS cells by RUNX3 was almost completely recovered by enforced expression of exogenous cycline D1 validated the cyclin D 1 indeed as a major target for RUNX3 in suppressing tumor cell growth.Restoration of-catenin expression could dramatically increase cyclin D1 transcription, and enforced expression of RUNX3 suppressed the effect of-catenin on cyclin D1 transcription.In conclusion, loss of RUNX3 expression in gastric cancer cells can promote cell cycle progression through up-regulating Akt/β-catenin/cyclin D 1 signaling pathway.