Activation of mitochondrial calcium-independent phospholipase A 2γ (iPLA2γ) by divalent cations mediating arachidonate release and production of downstream eicosanoids

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Calcium-independent phospholipase A2γ (iPLA 2γ) (PNPLA8) is the predominant phospholipase activity in mammalian mitochondria. However, the chemical mechanisms that regulate its activity are unknown. Here, we utilize iPLA2γ gain of function and loss of function genetic models to demonstrate the robust activation of iPLA2γ in murine myocardial mitochondria by Ca2+ or Mg2+ ions. Calcium ion stimulated the production of 2-arachidonoyl-lysophosphatidylcholine (2-AA-LPC) from 1-palmitoyl-2-[ 14C]arachidonoyl- sn-glycero-3-phosphocholine during incubations with wild-type heart mitochondrial homogenates. Furthermore, incubation of mitochondrial homogenates from transgenic myocardium expressing iPLA 2γ resulted in 13- and 25-fold increases in the initial rate of radiolabeled 2-AA-LPC and arachidonic acid (AA) production, respectively, in the presence of calcium ion. Mass spectrometric analysis of the products of calcium-activated hydrolysis of endogenous mitochondrial phospholipids in transgenic iPLA2γ mitochondria revealed the robust production of AA, 2-AA-LPC, and 2-docosahexaenoyl-LPC that was over 10-fold greater than wild-type mitochondria. The mechanism- based inhibitor (R)-(E)-6- (bromomethylene)-3-(1-naphthalenyl)- 2H-tetrahydropyran-2-one (BEL) (iPLA 2γ selective), but not its enantiomer, (S)-BEL (iPLA 2β selective) or pyrrolidine (cytosolic PLA2β selective), markedly attenuated Ca2+-dependent fatty acid release and polyunsaturated LPC production. Moreover, Ca2+-induced iPLA 2γ activation was accompanied by the production of downstream eicosanoid metabolites that were nearly completely ablated by (R)-BEL or by genetic ablation of iPLA2γ. Intriguingly, Ca 2+-induced iPLA2γ activation was completely inhibited by long-chain acyl-CoA (IC50 ∼20 μM) as well as by a nonhydrolyzable acyl-CoA thioether analog. Collectively, these results demonstrate that mitochondrial iPLA2γis activated by divalent cations and inhibited by acyl-CoA modulating the generation of biologically active metabolites that regulate mitochondrial bioenergetic and signaling functions.

Original languageEnglish
Pages (from-to)14880-14895
Number of pages16
JournalJournal of Biological Chemistry
Issue number18
StatePublished - Apr 27 2012


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