Death-associated protein kinase (DAP-kinase) is a calcium/calmodulin-dependent serine/threonine kinase, and participates in various apoptosis systems. However, its apoptosis-promoting mechanism is poorly understood. Here, we demonstrate that DAP-kinase suppresses integrin-mediated cell adhesion and signal transduction, whereas dominant-negative interference of this kinase promotes adhesion. This effect of DAP-kinase is neither a consequence of apoptosis nor a result of decreased expression of integrins. Rather, DAP-kinase downregulates integrin activity through an inside-out mechanism. We present evidence indicating that this adhesion-inhibitory effect accounts for a major mechanism of the apoptosis induced by DAP-kinase. First, in growth-arrested fibroblasts, DAP-kinase triggers apoptosis in cells plated on fibronectin, but does not affect the death of cells on poly-l-lysine. Second, in epithelial cells, DAP-kinase induces apoptosis in the anoikis-sensitive MCF10A cells, but not in the anoikis-resistant BT474 cells. Most importantly, the apoptosis-promoting effect of DAP-kinase is completely abolished by enforced activation of integrin-mediated signaling pathways from either integrin itself or its downstream effector, FAK. Finally, we show that integrin or FAK activation blocks the ability of DAP-kinase to upregulate p53. Our results indicate that DAP-kinase exerts apoptotic effects by suppressing integrin functions and integrin-mediated survival signals, thereby activating a p53-dependent apoptotic pathway.