CREPT (cell-cycle related and expression-elevated protein in tumor) was reported to be associated with growth of several human cancers; however, its clinical significance and regulatory mechanism still remain unclear in human gastric cancer. In the present study, we found CREPT was significantly increased in gastric cancer tissues compared to the matched adjacent normal tissues. CREPT silence inhibited the proliferation of gastric cancer cells through inducing G0/G1 phase cell cycle arrest, which was linked to the reduction of Cyclin D1 and Cyclin D-dependent kinase 4 (CDK4), and the elevation of p53 and p21. In addition, CREPT knockdown (KD) decreased migration of gastric cancer cells through up-regulating E-cadherin and down-regulating vimentin, N-cadherin and matrix metalloproteinase 1 (MMP-1) expressions. Further, CREPT KD induced apoptosis in gastric cancer cells, as evidenced by the increase of cleaved Caspase-3 and poly (ADP-ribose) polymerase (PARP). Intriguingly, suppressing p53 expressions significantly abolished CREPT silence-induced apoptosis, and reduction of cell viability. Moreover, CREPT KD caused reactive oxygen species (ROS) generation using discounted cash flow (DCF) analysis, which was reversed by ROS scavenger, N-acetyl-l-cysteine (NAC), pretreatment. Of note, NAC pretreatment abrogated apoptotic cell death in CREPT KD gastric cancer cells. In vivo, suppressing CREPT reduced the gastric tumor growth in gastric cancer xenograft models. Altogether, our results provided a novel insight into CREPT in regulating gastric cancer progression through apoptosis regulated by ROS/p53 pathways.