The function of cellular prion protein (PrP(C)), the essential protein for the pathogenesis and transmission of prion diseases, is still largely unknown. The putative roles of PrP(C) are thought to be related to cell signaling, survival, and differentiation. In a previous study, we showed that PrP(C) was overexpressed in gastric cancer tissues. In the present report, we show that ectopic expression of PrP(C) could promote tumorigenesis, proliferation, and G1/S transition in gastric cancer cells. Furthermore, CyclinD1, a protein related to cell cycle, was shown to be significantly up-regulated by PrP(C) at both mRNA and protein levels. PI3K/Akt pathway mediated above PrP(C) signal since PrP(C) increased the expression of phosphorylated Akt, and the specific inhibitor of Akt, LY294002, could markedly suppress growth of SGC7901 and transactivation of CyclinD1 induced by PrP(C). Octapeptide repeat region played a vital role in this function, as deletion of this region abolished or reduced these effects. Collectively, this study demonstrates that overexpression of PrP(C) might promote the tumorigenesis and proliferation of gastric cancer cells at least partially through activation of PI3K/Akt pathway and subsequent transcriptional activation of CyclinD1 to regulate the G1/S phase transition, in which octapeptide repeat region might be an indispensable region.