Atherosclerosis is readily observed in regions of blood vessels where disturbed blood flow (d-flow) is known to occur. A positive correlation between protein kinase C ζ (PKCζ) activation and d-flow has been reported, but the exact role of d-flow–mediated PKCζ activation in atherosclerosis remains unclear. We tested the hypothesis that PKCζ activation by d-flow induces endothelial cell (EC) apoptosis by regulating p53. We found that d-flow–mediated peroxynitrite (ONOO⁻) increased PKCζ activation, which subsequently induced p53 SUMOylation, p53–Bcl-2 binding, and EC apoptosis. Both d-flow and ONOO⁻ increased the association of PKCζ with protein inhibitor of activated STATy (PIASy) via the Siz/PIAS-RING domain (amino acids 301–410) of PIASy, and overexpression of this domain of PIASy disrupted the PKCζ–PIASy interaction and PKCζ-mediated p53 SUMOylation. En face confocal microscopy revealed increases in nonnuclear p53 expression, nitrotyrosine staining, and apoptosis in aortic EC located in d-flow areas in wild-type mice, but these effects were significantly decreased in p53^−/− mice. We propose a novel mechanism for p53 SUMOylation mediated by the PKCζ–PIASy interaction during d-flow–mediated EC apoptosis, which has potential relevance to early events of atherosclerosis.