The product of the retinoblastoma tumor-suppressor gene (RB) is a ubiquitously expressed, 105-kDa nuclear phosphoprotein (pRB). The pRB protein negatively regulates the cellular G1/S phase transition, and it is at this point in the cell cycle that it is thought to play its role as a tumor suppressor. The growth-inhibitory effects of pRB are exerted, at least in part, through the E2F family of transcription factors. This chapter reviews the insights into the mechanism of action of the E2F family members that have been obtained through overexpression studies. Studies in RB-/- SAOS-2 cells have provided evidence in support of the hypothesis that the E2F family members are negatively regulated by pRB and the related protein p130. In particular, the results obtained are consistent with the earlier biochemical data which suggested that E2F1 is regulated primarily by pRB, and E2F4 by p130. Results relating to p107 are also discussed. Consistent with the proposed role of pRB and E2F1 as coregulators of entry into S phase, experiments have demonstrated that overexpression of E2F1 is sufficient to override the cell cycle arrests caused by serum deprivation of fibroblasts or transforming growth factor-beta (TGF-beta) treatment of mink lung epithelial cells. However, at least in the case of the serum deprivation induced arrest, the ultimate result of E2F1 overexpression is death by p53-dependent apoptosis. In light of this and other data, a model is discussed as to how functional inactivation of pRB and p53 might cooperate to promote tumorigenesis. A number of studies have demonstrated the oncogenic potential of E2F family members, at least under certain conditions. This is, again, in keeping with the notion that these proteins play a critical role in controlling proliferation.