Vanadate-Sensitive Microsomal ATPases and Microsomal ⁴⁵Ca²⁺ Uptake in Tracheal Epithelial Cells

Abstract

Cytosolic free Ca²⁺ plays important roles in the regulation of physiological processes in tracheal epithelial cells and is probably regulated by many ion-transporting ATPases in these cells. Therefore, the effect of vanadate was investigated to characterize microsomal ion-transporting ATPases. Dose response experiments showed that vanadate had a biphasic effect on the microsomal ATPase activity: a decrease at the vanadate concentration below 100 μM, and a steep decrease at the concentration above 100 μM. The dose response data were fitted to two sigmoidal functions, corresponding to a low-affinity vanadate-sensitive (LAVS) ATPase and a high-affinity vanadate-sensitive (HAVS) ATPase. In ⁴⁵Ca²⁺ uptake experiments, both LAVS and HAVS ATPases mediated microsomal ⁴⁵Ca²⁺ uptake. The LAYS ATPase was selectively sensitive to thapsigargin in both ATPase activity and ⁴⁵Ca²⁺ uptake, suggesting that it is an ER/SR-type intracellular Ca²⁺-ATPase. Although the HAVS ATPase mediated one-fourth of microsomal ⁴⁵Ca²⁺ uptake, its activity was not sensitive to thapsigargin. These results indicate that the activities of these two vanadate-sensitive ATPases are mediated by different enzymes, since thapsigargin only blocks the activity of LAYS ATPase. In conclusion, there are two types of vanadate-sensitive microsomal ATPases, and these ATPases mediate microsomal ⁴⁵Ca²⁺ uptake in airway epithelial cells.