Mammalian group IIA phospholipase A2 (PLA2) is believed to play important roles in inflammation, cell injury, and tumor resistance. However, the cellular site of action has not been clearly defined as it has long been recognized that group IIA PLA2 is both a secretory and mitochondrial protein. The purpose of this study was to determine the subcellular target of the group IIA PLA2 and its role in apoptosis stimulated by growth factor withdrawal. Cloning of the rat liver group IIA PLA2 demonstrated a typical secretory signal and no alternative splicing of the primary transcript. When a sequence including the signal peptide and first 8 residues in the mature enzyme or the entire PLA2 (including the signal peptide) was fused to enhanced green fluorescent protein, the fusion protein was directed to the secretory pathway rather than mitochondria in baby hamster kidney (BHK) cells. To examine the role of group IIA PLA2 in cell injury, wild type (wt) rat group HA PLA2 and a mutant group IIA PLA2 containing a His-47 → Gln mutation (at the catalytic center) were transfected into BHK cells and cells stably expressing these constructs were isolated. After deprivation of growth factors, both normal BHK cells and BHK cells expressing mutant PLA2 underwent massive apoptosis, while BHK cells expressing wt PLA2 showed considerable resistance to growth factor withdrawal-induced apoptosis. The secretory PLA2 inhibitors 12-epi-scalaradial and aristolochic acid abrogated resistance to apoptosis in the wt PLA2 expressing cells. These two inhibitors did not induce cell death in the presence of fetal bovine serum, suggesting that they induce cell death by blocking PLA2 generated survival signals. This study demonstrates that group IIA PLA2 generates anti- apoptotic survival signals in BHK cells targeting the secretory pathway, and suggests that high levels of group IIA PLA2 accumulated at inflammatory sites may not only regulate inflammation, but also may protect cells from unnecessary death induced by pro-inflammatory agents.