Phospholemman overexpression inhibits Na+-K+-ATPase in adult rat cardiac myocytes: relevance to decreased Na+pump activity in postinfarction myocytes

Abstract

Messenger RNA levels of phospholemman (PLM), a member of the FXYD family of small single-span membrane proteins with putative ion-transport regulatory properties, were increased in postmyocardial infarction (MI) rat myocytes. We tested the hypothesis that the previously observed reduction in Na+-K+-ATPase activity in MI rat myocytes was due to PLM overexpression. In rat hearts harvested 3 and 7 days post-MI, PLM protein expression was increased by two- and fourfold, respectively. To simulate increased PLM expression post-MI, PLM was overexpressed in normal adult rat myocytes by adenovirus-mediated gene transfer. PLM overexpression did not affect the relative level of phosphorylation on serine68of PLM. Na+-K+-ATPase activity was measured as ouabain-sensitive Na+-K+pump current (Ip). Compared with control myocytes overexpressing green fluorescent protein alone, Ip measured in myocytes overexpressing PLM was significantly ( P < 0.0001) lower at similar membrane voltages, pipette Na+([Na+]pip) and extracellular K+([K+]o) concentrations. From −70 to +60 mV, neither [Na+]pipnor [K+]orequired to attain half-maximal Ip was significantly different between control and PLM myocytes. This phenotype of decreased Vmaxwithout appreciable changes in Kmfor Na+and K+in PLM-overexpressed myocytes was similar to that observed in MI rat myocytes. Inhibition of Ip by PLM overexpression was not due to decreased Na+-K+-ATPase expression because there were no changes in either protein or messenger RNA levels of either α1- or α2-isoforms of Na+-K+-ATPase. In native rat cardiac myocytes, PLM coimmunoprecipitated with α-subunits of Na+-K+-ATPase. Inhibition of Na+-K+-ATPase by PLM overexpression, in addition to previously reported decrease in Na+-K+-ATPase expression, may explain altered Vmaxbut not Kmof Na+-K+-ATPase in postinfarction rat myocytes.