Aim: Although the tumor suppressive role of EPB41L3 has been discovered in some research, our knowledge towards its molecular biology function and modulatory mechanisms remains indistinct.Moreover, the clinical significances of EPB41L3 expression have never been reported in esophageal squamous carcinoma (ESCC).We aim to find out whether EPB41L3 is a tumor suppressor in ESCC.Methods : We used MSP and pyrosequencing to determine the methylation status of EPB41L3 in ESCC, and then we used tissue microarray and a series of molecular biology experiments to explore the tumor suppresive role of EPB41L3.Results: In this study, we confirmed the reduced expression status of EPB41L3 in ESCC tissues and cell lines which was due to partial methylation.The ectopic expression of EPB41L3 in ESCC cells inhibited cell proliferation in vivo and in vitro by activating MAPK signaling.Moreover, EPB41L3 could induce apoptosis by activating caspase3/8/9 and generating G2/M arrest by activating Cdc2/Cyclin B 1 signalling.In clinical samples,ESCC tissues exhibited lower expression level of EPB41L3 compared with normal esophagous tissues.Patients with higher EPB41L3 expression had a markedly higher overall survival than patients with low EPB41L3 expression.Conclusion: Above all, our results indicate EPB41L3 as a functional tumor suppressive gene in ESCC development, representing a potential therapy target and prognosis indicator for clinical practice.