Phospholipase A2-activating protein (PLAA) enhances cisplatin-induced apoptosis in HeLa cells

Phospholipase A2 (PLA2)-activating protein (PLAA) is a novel signaling molecule that regulates eicosanoid production and participates in inflammatory responses. In our current study, we revealed that PLAA production was induced by the chemotherapeutic drug cisplatin in HeLa cervical carcinoma cells. To determine the potential pro-apoptotic effects of PLAA induction by cisplatin, we utilized HeLa (Tet-off) cells overexpressing the plaa gene (plaahigh) and compared them with control (plaalow) cells, which produce endogenous plaa from the chromosome. Cisplatin-stimulated plaahigh cells contained significantly higher levels of DNA fragmentation, caspase 3, 8 and 9 activities, PLA2 enzyme activity, and cytochrome c leakage from mitochondria than did the cisplatin-stimulated plaalow cells. Importantly, siRNA against PLAA (siRNA–PLAA) reduced the levels of cisplatin-induced PLAA, DNA fragmentation, and PLA2 activation, while promoting cell viability in both plaahigh and plaalow cells. Cisplatin-induced-cytochrome c leakage in plaahigh cells was reduced by siRNA–PLAA and restored by the addition of exogenous arachidonic acid (AA), suggesting to us that PLAA induction by cisplatin promoted cytochrome c leakage/mitochondrial damage partially by accumulating AA. In addition, cisplatin-stimulated plaahigh cells produced less cytoprotective clusterin than did the cisplatin-stimulated plaalow cells, and siRNA–PLAA promoted clusterin production from both plaahigh and plaalow cells. We showed that clusterin reduced DNA fragmentation in cisplatin-stimulated plaahigh and plaalow cells, which is consistent with the notion that clusterin promotes cancer chemoresistance. Furthermore, cisplatin-stimulated plaahigh cells produced more IL-32 (a pro-apoptotic protein) than did cisplatin-stimulated plaalow cells, and siRNA–PLAA reduced IL-32 production from both plaahigh and plaalow cells. Finally, our proteomic analysis revealed that cisplatin-stimulated plaahigh cells contained higher levels of phosphorylated JNK/c-Jun and FasL than did plaalow cells treated the same way. In summary, our data indicated that PLAA induction enhanced cisplatin-induced-apoptosis through four pathways, namely by: 1) accumulation of AA and mitochondrial damage, 2) downregulation of the cytoprotective clusterin, 3) upregulation of the pro-apoptotic IL-32, and 4) induction of JNK/c-Jun signaling and FasL expression.