DISSOCIATION OF PHOSPHOLAMBAN REGULATION OF CARDIAC SARCOPLASMIC-RETICULUM CA2+ ATPASE BY QUERCETIN

Quercetin had a biphasic effect on Ca2+ uptake and calcium-stimulated ATP hydrolysis in isolated cardiac sarcoplasmic reticulum (SR). Stimul ation of Ca(2+)ATPase was observed at low quercetin concentrations (<2 5 mu M) followed by inhibition at higher concentrations. The effects w ere dependent upon the SR protein concentration, the MgATP concentrati on, and intact phospholamban regulation of cardiac Ca(2+)ATPase. Only the inhibitory effects at higher quercetin concentrations were observe d in skeletal muscle SR which lacks phospholamban and in cardiac SR tr eated to remove phospholamban regulation, Stimulation was additive wit h monoclonal antibody 1D11 (directed against phospholamban) at submaxi mal antibody concentrations; however, the maximal antibody and quercet in stimulation were identical. Quercetin increased the calcium sensiti vity of the Ca(2+)ATPase like that observed with phosphorylation of ph ospholamban or treatment with monoclonal antibody 1D11. In addition, l ow concentrations of quercetin increased the steady-state formation of phosphoenzyme from ATP or P-i, but higher quercetin decreased phospho enzyme levels. Quercetin, even under stimulatory conditions, was a com petitive inhibitor of ATP, but ap pears to relieve the Ca(2+)ATPase fr om phospholamban inhibition, thereby, producing an activation. The sub sequent inhibitory action of higher quercetin concentrations results f rom competition of quercetin with the nucleotide binding site of the C a(2+)ATPase. The data suggest that quercetin interacts with the nucleo tide binding site to mask phospholamban's inhibition of the SR Ca(2+)A TPase and suggests that phospholamban may interact at or near the nucl eotide binding site.