The death of insulin-secreting β-cells that causes type I diabetes mellitus (DM) occurs in part by apoptosis, and apoptosis also contributes to progressive β-cell dysfunction in type II DM. Recent reports indicate that ER stress-induced apoptosis contributes to β-cell loss in diabetes. Agents that deplete ER calcium levels induce β-cell apoptosis by a process that is independent of increases in [Ca2+]i. Here we report that the SERCA inhibitor thapsigargin induces apoptosis in INS-1 insulinoma cells and that this is inhibited by a bromoenol lactone (BEL) inhibitor of group VIA calcium-independent phospholipase A2 (iPLA 2β). Overexpression of iPLA2β amplifies thapsigargin-induced apoptosis of INS-1 cells, and this is also suppressed by BEL. The magnitude of thapsigargin-induced INS-1 cell apoptosis correlates with the level of iPLA2β expression in various cell lines, and apoptosis is associated with stimulation of iPLA2β activity, perinuclear accumulation of iPLA2β protein and activity, and caspase-3-catalyzed cleavage of full-length 84 kDa iPLA2β to a 62 kDa product that associates with nuclei. Thapsigargin also induces ceramide accumulation in INS-1 cells, and this response is amplified in cells that overexpress iPLA2β. These findings indicate that iPLA 2β participates in ER stress-induced apoptosis, a pathway that promotes β-cell death in diabetes.