The presence of protein kinase C (PKC) in mammalian sperm was demonstrated by enzymatic activity assay and immunohistochemistry at the light and electron microscopy levels. The sperm head PKC is localized in the acrosome, equatorial segment, and postacrosomal region. In the flagellum, PKC is associated with the segmented column of the neck and is distributed along the mid, principal, and end pieces. Immunoreactive sites are observed in patches along the axoneme and outer dense fibers and are evenly distributed between these regions. Functional studies suggest the involvement of PKC in flagellar motility and acrosome reaction. The cross-talk between the signaling cascades that operate during sperm activation is discussed.