The retinoblastoma protein, pRb, plays important roles in many processes implicated in cell fate decisions, including cell cycle, differentiation and apoptosis. In cell cycle regulation, pRb interacts principally with the E2F transcription factor family members to inhibit the transcription of many genes controlling cell cycle progression. In this study, we focused on the role of pRb in apoptosis, which is much less clear than its role in cell cycle regulation. Indeed, pRb has been found to be either pro- or anti-apoptotic. To clarify how the proliferative status of the cells impacts the role of pRb in apoptosis, we used Drosophila to induce RBF (the pRb fly homologue) expression in different cellular and developmental contexts. We found that RBF expression induces apoptosis in different proliferative tissues in a caspase-dependent manner, whereas this effect was not observed in differentiated post-mitotic cells. Furthermore, RBF-induced apoptosis in proliferating cells was inhibited by co-expression of dE2F1, an antagonistic partner of RBF in cell cycle regulation. These results are in agreement with the view that the apoptotic properties of pRb are tightly linked to, and are probably a consequence of, an effect on cell cycle progression. Moreover, we show for the first time that RBF has a direct anti-apoptotic effect on Dmp53-induced cell death in post-mitotic cells only. Taken together, these data clearly show that RBF can exert a dual role in the control of apoptotic processes, and that its properties depend on the proliferative status of the cells.