Thapsigargin discriminates strongly between Ca(2+)-ATPase phosphorylated intermediates with different subcellular distributions in bovine adrenal chromaffin cells.
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.
Department of Medical Physiology, University of Copenhagen, Denmark.
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Caspersen
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Treiman
D000313:Adrenal Medulla / Q000201:enzymology / Q000648:ultrastructure*; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000032:analysis / Q000037:antagonists & inhibitors* / Q000378:metabolism; D002417:Cattle; D002458:Cell Fractionation; D002838:Chromaffin System / Q000201:enzymology / Q000648:ultrastructure*; D004721:Endoplasmic Reticulum / Q000201:enzymology* / Q000648:ultrastructure; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism; D008274:Magnesium / Q000494:pharmacology; D008861:Microsomes / Q000201:enzymology* / Q000648:ultrastructure; D010446:Peptide Fragments / Q000378:metabolism; D010766:Phosphorylation; D013729:Terpenes / Q000378:metabolism / Q000494:pharmacology*; D019284:Thapsigargin; D014357:Trypsin / Q000378:metabolism
D007527:Isoenzymes; D010446:Peptide Fragments; D013729:Terpenes; D019284:Thapsigargin; D014357:Trypsin; D000252:Calcium-Transporting ATPases; D008274:Magnesium; D002118:Calcium