Expression and functional characterization of isoforms 4 of the plasma membrane calcium pump.

PMCA isoforms 4CII (generated by splicing at the C-terminus) and 4BICI (a pump version lacking the 10th transmembrane domain) were expressed in Sf9 cells using the baculovirus system. The purified PMCA4CII had a 20-fold lower affinity for calmodulin than the PMCA4CI, the PMCA4 isoform of the erythrocytes' membranes, but had a higher activity in the absence of calmodulin. The amount of phosphoenzyme intermediate formed by PMCA4CII in the presence of Ca2+ alone was almost 3 times higher than in PMCA4CI and was increased by La3+ less than in the PMCA4CI. The isoform lacking the 10th transmembrane domain (PMCA4BICI) had no Ca2+-dependent ATPase activity, but was still able to form the phosphoenzyme intermediate starting from phosphate. When expressed in COS cells, this isoform was retained in the endoplasmic reticulum; changes in membrane architecture apparently occurred during its expression; the C-terminal portion of the isoform was located in the cytosol, indicating that the deletion of the 10th transmembrane domain resulted in the loss of at least another transmembrane domain.