Plasma membrane calcium-ATPase in cultured human retinal pigment epithelium

The present work demonstrates the presence of plasma membrane (Ca(2+) + Mg2+)-ATPase (PMCA) activity in cultured human retinal pigment epithelium (HRPE). Whole-cell HRPE homogenates exhibited Ca(2+)-stimulated ATPase activity that was inhibited with high affinity (IC50 = 60 nM) by eosin, a potent inhibitor of the erythrocyte PMCA. This activity was not inhibited by thapsigargin, a selective inhibitor of the sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase (SERCA). A PMCA-specific monoclonal antibody exhibited staining in attached HRPE monolayers. By Western blot analysis of SDS-PAGE separations of whole cell lysates of HRPE, this antibody identified a single band at approximately 145 kD. Finally, a Ca(2+)-dependent, La(3+)-augmented phosphoprotein, which comigrated with anti-PMCA immunoreactivity, was also detected. Taken together, these results show that cultured HRPE express PMCA activity.