〔Abstract〕 Objective To investigate the effects of knockdown of E2F transcription factor 1(E2F1) in human tongue squamous cell carcinoma cells on proliferation, migration, and invasion of tongue squamous cell carcinoma cells, and to explore its possible mechanisms. Methods The expression ofE2F1in human tongue squamous cell carcinoma was detected by RT-PCR, and the background expression ofE2F1gene in HOEC, SCC-9, CAL-27, TSCCa and SCC-25 cells was detected by RT-PCR, and the cell lines with significantly high expression of E2F1 were screened for subsequent experiments. CCK-8, EdU, colony formation, cell scratch, Transwell, apoptosis and cell cycle experiments were used to detect the effects of knockdown ofE2F1on proliferation, migration, invasion and apoptosis of human tongue squamous cell carcinoma cells after knockdown ofE2F1gene in tongue squamous cell carcinoma candidate cells. Western blot was used to detect the expression of epithelial mesenchymal transition markers E-cadherin, N-cadherin and snail family transcription inhibitor 1(Snail) protein and WNT signaling pathway markers WNT family members 3A(WNT3A), β-catenin and cyclin D1(CCND1). Results RT-PCR results showed that the expression ofE2F1in cancer tissues was significantly higher than that in adjacent tissues(P<0.001). At the same time, compared with HOEC cells, the expression ofE2F1in SCC-9, CAL-27, TSCCa and SCC-25 cells significantly increased(P<0.05), and the high expression in SCC-25 cells was the most obvious. SCC-25 would be used as an experimental cell line in subsequent experiments. After knocking downE2F1in SCC-25 cells, the viability of CCK-8 cells was significantly inhibited(P<0.001). The number of EdU positive cells decreased significantly(P<0.001). The number of cell clones was significantly reduced(P<0.001). The proportion of total apoptosis significantly increased(P<0.001). The proportion of cells in G1phase increased(P<0.01). The proportion of cells in G2phase decreased(P<0.01). The cell migration and invasion ability were significantly inhibited(P<0.001). Western blot showed that the expression of WNT signaling pathway proteins WNT3A, β-catenin and CCND1 decreased(P<0.001), while the expression of EMT-related proteins N-cadherin and Snail decreased(P<0.001), while the expression of E-cadherin increased(P<0.001). Conclusion Knockdown ofE2F1inhibits the proliferation, migration, invasion and EMT process of human tongue squamous cell carcinoma SCC-25 cells, and promotes apoptosis. This anti-tumor effect may be achieved by blocking the activation of WNT signaling pathway.