MEMBRANE PHOSPHORYLATION PROTECTS THE CARDIAC SARCOPLASMIC-RETICULUM CA2-ATPASE AGAINST CHLORINATED OXIDANTS IN-VITRO()

Objective: The calcium (Ca) pump of cardiac sarcoplasmic reticulum (SR ) membranes is vulnerable to oxidation and hence likely to be damaged by chlorinated compounds, specifically hypochlorite (NaOCl) and monoch loramine (NH2Cl), the most potent oxidants produced upon neutrophil ac tivation. This could occur during prolonged ischemia or myocardial inf arction when tissue levels of catecholamines are high. Phospholamban ( PLN), the phosphorylatable regulator of the Ca pump, plays a central r ole in the effects of beta-adrenergic agonists on the heart. The purpo se of this study was to investigate a possible role of PLN in determin ing the pump's sensitivity to NaOCl and NH2Cl. Methods: Ca-uptake and Ca2+-ATPase activities in purified phosphorylated and control canine c ardiac microsomes, incubated at increasing concentrations of NaOCl or NH2Cl, were related to the extent of PLN phosphorylation by protein ki nase A, which was quantitated by PhosphorImager analysis. Results and Conclusions: Our data indicate that microsomal phosphorylation protect s the Ca pump fully against 10 mu M NaOCl or NH2Cl. which inhibit Ca-u ptake by 21-41% when assayed at 25 or 37 degrees C and saturating Ca2 in unphosphorylated microsomes, and protects partially at higher oxid ant concentrations. The protective effect of protein kinase A on Ca-up take is proportional to the amount of phosphorylated PLN. No comparabl e protection against similar oxidative damage of the Ca pump is observ ed when light fast skeletal muscle microsomes, which lack PLN. are inc ubated under conditions favorable for phosphorylation nor when PLN's i nhibition of the cardiac Ca pump is relieved by proteolytic cleavage o f its cytoplasmic domain. Our findings contribute toward an understand ing of possible endogenous protective mechanisms that may promote calc ium homeostasis in myocardial cells in inflammatory states associated with neutrophil activation and may suggest an approach toward developm ent of protective strategies against oxidative damage in the heart. (C ) 1997 Elsevier Science B.V.