Identification of calcium-independent phospholipase A2γ in mitochondria and its role in mitochondrial oxidative stress

Oxidant-induced lipid peroxidation and cell death mediate pathologies associated with ischemia-reperfusion and inflammation. Our previous work in rabbit renal proximal tubular cells (RPTC) demonstrated that inhibition of Ca2+-independent phospholipase A(2) ( iPLA(2)) potentiates oxidant-induced lipid peroxidation and necrosis, implicating iPLA2 in phospholipid repair. This study was conducted to identify a RPTC mitochondrial PLA(2) and determine the role of PLA(2) in oxidant-induced mitochondrial dysfunction. iPLA2 activity was detected in Percoll-purified rabbit renal cortex mitochondria (RCM) and in isolated mitochondrial inner membrane fractions from rabbit and human RCM. Immunoblot analysis and inhibitor sensitivity profiles revealed that iPLA(2)gamma is the RCM iPLA2 activity. RCM iPLA2 activity was enhanced in the presence of ATP and was blocked by the PKC epsilon V1-2 inhibitor. Oxidant-induced mitochondrial lipid peroxidation and swelling were accelerated by pretreatment with R-BEL, but not S-BEL. Furthermore, oxidant treatment of isolated RCM resulted in decreased iPLA2 gamma activity. These results reveal that RCM iPLA(2) is iPLA(2)gamma, RCM iPLA(2)gamma is regulated by phosphorylation by PKC epsilon, iPLA(2)gamma protects RCM from oxidant-induced lipid peroxidation and dysfunction, and that a strategy to preserve or enhance iPLA(2)gamma activity may be of therapeutic benefit.