Synaptotagmins (Syts) play a key role in the regulation of Ca(2+)-triggered exocytosis and membrane fusion events, two crucial events associated to the phagocytic process. In the present study, we investigated the role of Syt V, a regulator of focal exocytosis, in phagocytosis. In macrophages, Syt V is localized on recycling endosomes and on filopodia-like structures and is recruited to the nascent phagosomes independently of the phagocytic receptor engaged. Silencing of Syt V by RNA interference revealed a role for this protein for phagocytosis, particularly under conditions of high membrane demand. In contrast, silencing of Syt V had no effect on the recruitment of the lysosomal marker LAMP1 to phagosomes, indicating that phagosome maturation is not regulated by Syt V. Collectively, these results illustrate the importance of Syt V in the regulation of an important innate function of macrophages. Furthermore, our results are consistent with the concept that focal exocytosis of endocytic organelles is a key event in phagocytosis and suggest that Syt V regulates this process.