Apoptosis is involved in the development and progression of atherosclerotic lesions. Protein kinase C (PKC) signalling is of importance in atherosclerosis as well as apoptosis. Therefore, we tested the involvement of PKC in lipid-induced apoptosis of human coronary artery endothelial cells (HCAEC). Protein expression of PKC isoforms alpha, beta I, delta, epsilon, and iota was detected, whereas no relevant protein amounts of PKC isoforms beta II, gamma, eta, theta, and zeta were found. Inhibition of classical and novel PKC isoforms by treatment with bisindolylmaleimide or PKC down-regulation by long-term treatment with 12-O-tetradecanoyl phorbol-13-acetate (TPA) could not prevent apoptosis induced by palmitate or stearate. In contrast, a specific myristoylated, cell-permeable PKC zeta/iota pseudosubstrate prevented lipid-induced apoptosis in HCAEC. Furthermore, saturated fatty acids activated PKC iota as evidenced by PKC iota down-regulation upon long-term treatment with stearate. Our data provide evidence that PKC iota is activated by saturated fatty acids and mediates lipid-induced apoptosis of HCAEC.