Protein kinase C-epsilon coimmunoprecipitates with cytochrome oxidase subunit IV and is associated with improved cytochrome-c oxidase activity and cardioprotection.
We have utilized an in situ rat coronary ligation model to establish a PKC-epsilon cytochrome oxidase subunit IV (COIV) coimmunoprecipitation in myocardium exposed to ischemic preconditioning (PC). Ischemia-reperfusion (I/R) damage and PC protection were confirmed using tetrazolium-based staining methods and serum levels of cardiac troponin I. Homogenates prepared from the regions at risk (RAR) and not at risk (RNAR) for I/R injury were fractionated into cell-soluble (S), 600 g low-speed centrifugation (L), Percoll/Optiprep density gradient-purified mitochondrial (M), and 100,000 g particulate (P) fractions. COIV immunoreactivity and cytochrome-c oxidase activity measurements estimated the percentages of cellular mitochondria in S, L, M, and P fractions to be 0, 55, 29, and 16%, respectively. We observed 18, 3, and 3% of PKC-delta, -epsilon, and -zeta isozymes in the M fraction under basal conditions. Following PC, we observed a 61% increase in PKC-epsilon levels in the RAR M fraction compared with the RNAR M fraction. In RAR mitochondria, we also observed a 2.8-fold increase in PKC-epsilon serine 729 phosphoimmunoreactivity (autophosphorylation), indicating the presence of activated PKC-epsilon in mitochondria following PC. PC administered before prolonged I/R induced a 1.9-fold increase in the coimmunoprecipitation of COIV, with anti-PKC-epsilon antisera and a twofold enhancement of cytochrome-c oxidase activity. Our results suggest that PKC-epsilon may interact with COIV as a component of the cardioprotection in PC. Induction of this interaction may provide a novel therapeutic target for protecting the heart from I/R damage.
Department of Pharmacology & Toxicology, School of Medicine, Medical College of Georgia, Augusta, Georgia 30912-2300, USA.
Dehuang
D
Guo
Tiffany
T
Nguyen
Mourad
M
Ogbi
Huda
H
Tawfik
Guochun
G
Ma
Qilin
Q
Yu
Robert W
RW
Caldwell
John A
JA
Johnson
D000818:Animals; D003331:Coronary Vessels / Q000601:surgery; D004195:Disease Models, Animal; D003576:Electron Transport Complex IV / Q000378:metabolism*; D004789:Enzyme Activation; D047468:Immunoprecipitation; D019194:Ischemic Preconditioning; D007527:Isoenzymes / Q000378:metabolism; D008026:Ligation; D008929:Mitochondria, Heart / Q000201:enzymology*; D046912:Multiprotein Complexes / Q000378:metabolism; D009203:Myocardial Infarction / Q000150:complications / Q000201:enzymology / Q000473:pathology / Q000628:therapy*; D015428:Myocardial Reperfusion Injury / Q000201:enzymology / Q000209:etiology / Q000473:pathology / Q000517:prevention & control*; D009206:Myocardium / Q000201:enzymology* / Q000473:pathology; D010085:Oxidative Phosphorylation; D051744:Protein Kinase C-epsilon / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D019210:Troponin I / Q000097:blood
D007527:Isoenzymes; D046912:Multiprotein Complexes; D019210:Troponin I; D003576:Electron Transport Complex IV; D051744:Protein Kinase C-epsilon