Molecular and Cellular Biochemistry 99: 83-88, 1990. © 1990 Kluwer Academic Publishers. Printed in the Netherlands. Invited Paper The role of phospholamban in the regulation of calcium transport by cardiac sarcoplasmic reticulum Bruce A. Davis 1, Istvan Edes, Ramesh C. Gupta, Ellen F. Young, Hae Won Kim, Nancy A.E. Steenaart, Grazyna Szymanska and Evangelia G. Kranias Department of Pharmacology and Cell Biophysics, University of Cincinnati, College of Medicine, 231 Bethesda Avenue, Cincinnati, Ohio 45267-0575 USA; 1 Present address: Department of Cellular and Molecular Physiology, Yale University, New Haven, CT, USA Accepted30 April 1990 Key words: cardiac, sarcoplasmic reticulum, phospholamban, calcium transport, protein kinase, phosphatase Abstract The calcium transport mechanism of cardiac sarcoplasmic reticulum (SR) is regulated by a phospho- regulatory mechanism involving the phosphorylation-dephosphorylation of an integral membrane compo- nent, termed phospholamban. Phospholamban, a 27,000 Da proteolipid, contains phosphorylation sites for three independent protein kinases: 1) cAMP-dependent, 2) Ca2+-calmodulin-dependent, and 3) Ca 2+- phospholipid-dependent. Phosphorylation of phospholamban by any one of these kinases is associated with stimulation of the calcium transport rates in isolated SR vesicles. Dephosphorylation of phosphorylated phospholamban results in the reversal of the stimulatory effects produced by the protein kinases. Studies conducted on perfused hearts have shown that during exposure to beta-adrenergic agents, a good correlation exists between the in situ phosphorylation of phospholamban and the relaxation of the left ventricle. Phosphorylation of phospholamban in situ is also associated with stimulation of calcium transport rates by cardiac SR, similar to in vitro findings. Removal of beta-adrenergic agents results in the reversal of the inotropic response and this is associated with dephosphorylation of phospholamban. These findings indicate that a phospho-regulatory mechanism involving phospholamban may provide at least one of the controls for regulation of the contractile properties of the myocardium. Introduction In cardiac sarcoplasmic reticulum (SR) there are several types of regulatory processes involved in the function of the Ca2+-ATPase. These include substrate level regulation (e.g., Ca z+, ATP, Mg2÷, etc.), intermolecular interaction, and covalent modification via phosphorylation-dephosphoryla- tion of phospholamban, an intrinsic protein of the SR. Phospholamban is a complex proteolipid com- posed of 5 identical subunits [1, 2], each consisting of 52 amino acids [3]. Several positively charged amino acids render the protein rather alkaline (pI 10) [4]. However, a strong acidic shift is observed upon phosphorylation (pI 6.4-5.2) [2]. The mole- cular details of the mechanism by which phospho- lamban phosphorylation affects Ca2+-ATPase ac- tivity are not known. Work from several laboratories has shown that at least two second messengers, cAMP and calci- um, are involved in the regulation of the Ca 2+- ATPase in cardiac SR. Regulation occurs through