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Calcium‐independent phospholipase A 2 in isolated rabbit ventricular myocytes
Author(s) -
McHowat Jane,
Creer Michael H.
Publication year - 1998
Publication title -
lipids
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.601
H-Index - 120
eISSN - 1558-9307
pISSN - 0024-4201
DOI - 10.1007/s11745-998-0324-5
Subject(s) - myocyte , biochemistry , cytosol , phospholipase , phospholipase c , chinese hamster ovary cell , phospholipase a2 , phospholipase a , protein kinase c , biology , chemistry , endocrinology , receptor , kinase , enzyme
Abstract We characterized phospholipase A 2 (PLA 2 ) activity in isolated rabbit ventricular myocytes with respect to subcellular distribution, substrate specificity, and Ca 2+ dependency. Membrane‐associated PLA 2 was found to be an order of magnitude greater than cytosolic PLA 2 . Ventricular myocyte PLA 2 activity was enhanced following protease‐activated receptor stimulation with thrombin and was found to be largely Ca 2+ ‐independent and selective for phospholipid substrates containing arachidonic acid at the sn ‐2 position. Immunoblot analysis using an antibody to cytosolic Ca 2+ ‐independent PLA 2 from Chinese hamster ovary cells recognized a membrane‐associated protein with a molecular mass of approximately 80 kDa; however, differences in pH optima, response to inhibitors, and substrate selectivity of membrane‐associated and cytosolic PLA 2 activity suggest the presence of multiple Ca 2+ ‐independent PLA 2 . Pretreatment with bromoenol lactone, a specific inhibitor of Ca 2+ ‐independent PLA 2 , significantly attenuated membrane‐associated and cytosolic PLA 2 in unstimulated and thrombin‐stimulated myocytes. Pretreatment with methyl arachidonyl fluorophosphonate, mepacrine, or dibucaine had no significant effect on PLA 2 activity under all conditions tested. Ventricular myocyte PLA 2 activity was significantly inhibited by ATP, GTP, and their nonhydrolyzable analogs and was regulated by protein kinase C activity. These studies demonstrate the presence of one or more unique membrane‐associated Ca 2+ ‐independent PLA 2 in isolated ventricular myocytes that exhibit a preference for phospholipids with arachidonate at the sn ‐2 position and that are activated by thrombin stimulation.