p63 is a member of the p53 protein family and plays a crucial role in epithelial development. p63 is expressed in many types of tumors including esophageal cancer; however, its function in cancer is controversial and its role in esophageal cancer has not been clearly established. In the present study, we aimed to identify the mechanisms by which p63 promotes proliferation of esophageal squamous carcinoma cells. Four human esophageal cancer cell lines (TE-8, TE-12, BE3 and OE33) were used in this study. We found that ΔNp63 was the predominantly expressed p63 isoform in esophageal squamous cancer cells. Silencing of p63 mRNA in the esopha-geal cancer cell lines TE-8 and TE-12 resulted in significant inhibition of cell proliferation in a dose-dependent manner. A colony formation assay also showed that colony formation by TE-8 and TE-12 cells was significantly inhibited by p63 siRNA. Furthermore, p63 siRNA significantly suppressed p-Akt and induced Akt expression in esophageal squamous carcinoma cell lines. On the other hand, overexpression of p63 in the esophageal cell lines BE3 and OE33 increased p-Akt expression. Silencing of p63 in TE-8 and TE-12 cell lines induced p53 and p27 expression and suppressed cyclin D1 and cyclin E1 expression, whereas overexpression of p63 in BE3 and OE33 cell lines resulted in decreased levels of p53 and p27 and increased levels of cyclin D1 and cyclin E1. Taken together, our results suggest that p63 may play a pivotal role in the progression of esophageal squamous carcinoma cells through regulation of the cell cycle via the Akt signaling pathway.