The mitochondrial transcription termination factor (MTERF) family is a group of highly conserved DNA-binding proteins composed of four key members, MTERF1–4. To date, several studies have investigated the binding sites of MTERF1 on mitochondrial genome and the regulation of mitochondrial gene transcription, but the more intricate connection between mitochondrial genes transcription regulation, mitochondrial oxidative phosphorylation (OXPHOS), and cell proliferation is still poorly understood. In this study, we constructed over-expression and knockdown vectors of MTERF1 that were transfected into HeLa cells to investigate the functions of MTERF1. Results showed that although MTERF1 is a positive regulatory factor of mitochondrial genes transcription, it had no significant effect on the replication of mitochondrial DNA. Over-expression of MTERF1 increased mitochondrial oxidative phosphorylation activity and promoted ATP synthesis, cyclin D1 expression, and cell proliferation, while its knockdown inhibited ATP synthesis, decreased cyclin D1 expression, and slowed the cell growth. These results suggested that MTERF1 may promote cell proliferation by regulating oxidative phosphorylation activity in HeLa cells. Ultimately, these findings create a foundation for further and more conclusive studies on the physiological functions of MTERF family by providing novel insights into the potential mechanisms underlying cell proliferation regulation.