Cell proliferation is regulated by the cell cycle, and in order to divide the cell must enter a mitotic state. Prior to mitosis the cell is required to pass through a number of checkpoints, including the critical G1/S restriction point governed by the successive phosphorylation of the retinoblastoma protein, pRb. The various proteins and regulatory factors governing pRb phosphorylation have been a major focus of study in recent years, given the central importance of G1/S transition deregulation in cancer development. This review summarises the molecular biology around the G1/S transition, focussing on the critical roles of the transcription factor family E2F and the cyclin-dependent kinase (CDK) and cyclin families involved in E2F release from pRb. Interestingly, E2F release from pRb is associated with cell proliferation; however, above a certain threshold E2F has the potential to trigger apoptosis. The review focuses on the following topics: (i) how E2F and other substrates bind to pRb at the molecular level; (ii) mechanisms by which pRb function is modulated within the cell; (iii) mechanisms that inhibit or enhance cell proliferation via the pRb/E2F pathway; (iv) how E2F can potentiate apoptotic pathways; and (v) what controls whether E2F mediates cell proliferation or apoptosis. The case for the development of agents that perturb pRb:E2F interactions will be made, as a strategy to further inform the molecular biology around this important target and as a therapeutic strategy against cancer.