PRDM14 (PRDI-BF1 and RIZ domain-containing 14), a transcription factor, plays important roles in primordial germ cell specification and embryonic stem cell pluripotency, and supports the maintenance of self-renewal by promoting the expression of stem cell markers while also repressing the expression of differentiation factors. As a proto-oncogene, the ectopic expression of PRDM14 can enhance breast cell growth and reduce breast cell sensitivity to chemotherapeutic drugs. Conversely, knockdown of PRDM14 expression induces apoptosis in breast cancer cells and restores their sensitivity to chemotherapeutic drugs. Here, we sought to identify the role of PRDM14 in 293T cells. PRDM14-infected 293T cells exhibited an abnormal morphology, and we found that ectopic expression of PRDM14 inhibits colony formation, cell proliferation and metastasis. In addition, our data indicated that PRDM14 influences the G1/S phase transition of 293T cells by inducing the expression of cell cycle regulators. In conclusion, these results showed that PRDM14 inhibits 293T cell proliferation by influencing the G1/S phase transition and impacts cell migration by regulating the level of MMP/TIMP expression, thus mediating extracellular matrix degradation. •Exogenous PRDM14 was specifically localized to the nucleus in 293T cells.•Ectopic expression of PRDM14 inhibited 293T cell proliferation, changed clonal formation ability and cell migration ability.•PRDM14 overexpression influenced the G1/S phase transition of 293T cells through multiple parallel mechanisms.