Monocytes are central components of the innate immune response and normally circulate for a short period of time before undergoing spontaneous apoptosis. During inflammation, differentiation, or oncogenic transformation, the life span of monocytes is prolonged, by preventing the activation of the apoptotic program. Here, we show that caspase-3, a cysteine protease required for monocyte apoptosis, is a phosphoprotein. We identified PKCdelta as a member of the protein kinase C family that associates with and phosphorylates caspase-3. The PKCdelta-dependent phosphorylation of caspase-3 results in an increase in the activity of caspase-3. This effect of PKCdelta is specific to caspase-3, evidenced by the absence of similar effects on caspase-9. The activity of PKCdelta precedes the activation of caspase-3 during spontaneous monocyte apoptosis and in monocyte-induced-apoptosis. We found that the over-expression of PKCdelta results in an increase of apoptosis, while its inhibition blocks caspase-3 activity and decreases apoptosis. Our results provide evidence that the PKCdelta-dependent phosphorylation of caspase-3 provides a novel pro-apoptotic mechanism involved in the regulation of monocyte life span.