Thapsigargin discriminates strongly between Ca(2+)-ATPase phosphorylated intermediates with different subcellular...

We studied the effects of thapsigargin on the formation of the phosphorylated intermediates (E approximately Ps) of endoplasmic reticulum Ca(2+)-ATPases in microsomes from bovine adrenal medulla. When submicrosomal fractions were separated on a sucrose gradient, two components of 100 kDa Ca(2+)-ATPase E approximately P displaying distinct subcellular distributions were resolved. The first component was defined by Ca(2+)-induced protection against thapsigargin inhibition. The second component did not display such protection, with a 3 orders of magnitude difference in thapsigargin inhibitory potency towards the 2 components. In the absence of Ca2+, both E approximately P components were highly sensitive to thapsigargin inhibition, revealing the presence of high-affinity thapsigargin-binding sites characteristic of SERCA ATPases. These data demonstrate a new level of molecular heterogeneity among Ca(2+)-ATPases of endoplasmic reticulum, and provide the first evidence of differential subcellular localization of individual Ca2+ pump subtypes in cells of neural origin.