Functional effects of protein kinases and peroxynitrite on cardiac carnitine palmitoyltransferase-1 in isolated mitochondria.
We have previously shown that metoprolol can inhibit carnitine palmitoyltransferase-1 catalytic activity and decrease its malonyl CoA sensitivity within 30 min, suggesting the importance of a covalent modification. The aim of this study was to characterize the effects of PTMs on CPT-1 in the heart. Mitochondria were isolated from the hearts of male Wistar rats and incubated with kinases of interest (protein kinase A, CAMK-II, p38 MAPK, Akt) or with peroxynitrite and sodium nitroprusside. PKA decreased CPT-1 malonyl CoA sensitivity, associated with phosphorylation of CPT-1A, whereas CAMK-II increased malonyl CoA sensitivity by phosphorylating CPT-1B. p38 bound to CPT-1B and stimulated CPT-1 activity. The association of CPT-1 with these kinases and their scaffolding proteins was confirmed in co-localization studies. Peroxynitrite and sodium nitroprusside reversibly stimulated CPT-1 activity, and the change in CPT-1B activity was most consistently associated with glutathiolation of CPT-1B. These studies have identified a new regulatory system of kinases, scaffolding proteins and thiol redox chemistry which can control cardiac CPT-1 in vitro.
Division of Pharmacology and Toxicology, Faculty of Pharmaceutical Sciences, The University of British Columbia, Vancouver, Canada.
Vijay
V
Sharma
Thomas
T
Abraham
Amie
A
So
Michael F
MF
Allard
John H
JH
McNeill
D000818:Animals; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology; D002334:Carnitine O-Palmitoyltransferase / Q000378:metabolism*; D002458:Cell Fractionation; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D066298:In Vitro Techniques; D008297:Male; D008316:Malonyl Coenzyme A / Q000378:metabolism; D008929:Mitochondria, Heart / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism*; D009599:Nitroprusside / Q000494:pharmacology; D051058:Oncogene Protein v-akt / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology; D030421:Peroxynitrous Acid / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D011494:Protein Kinases / Q000378:metabolism / Q000494:pharmacology* / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D014018:Tissue Distribution; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology
D030421:Peroxynitrous Acid; D009599:Nitroprusside; D008316:Malonyl Coenzyme A; D002334:Carnitine O-Palmitoyltransferase; D011494:Protein Kinases; D051058:Oncogene Protein v-akt; D017868:Cyclic AMP-Dependent Protein Kinases; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D048051:p38 Mitogen-Activated Protein Kinases