Distribution of two distinct Ca2+ -ATPase-like proteins and their relationships to the agonist-sensitive calcium store in adrenal chromaff in cells

MANY cellular functions are regulated by activation of cell-surface receptors that mobilize calcium from internal stores sensitive to inositol 1,4,5-trisphosphate (Ins(l,4,5)P 3 ) (ref. 1). The nature of these internal calcium stores and their localization in cells is not clear and has been a subject of debate. It was originally suggested 2–5 that the Ins(l,4,5)P 3 -sensitive store is the endoplasmic reticulum, but a new organelle, the calciosome, identified by its possession of the calcium-binding protein, calsequestrin 6 , and a Ca 2+ -ATPase-like protein 7 of relative molecular mass 100,000 (100K), has been described as a potential Ins(l,4,5)P 3 -sensitive calcium store. Direct evidence on whether the calciosome is the Ins(l,4,5)P 3 -sensitive store is lacking. Using monoclonal anti-bodies raised against the Ca 2+ -ATPase of skeletal muscle sarcoplasmic reticulum 8 , we show that bovine adrenal chromaffin cells contain two Ca 2+ -ATPase-like proteins with distinct subcellular distributions. A 100K Ca 2+ -ATPase-like protein is diffusely distributed, whereas a 140K Ca 2+ -ATPase-like protein is restricted to a region in close proximity to the nucleus. In addition, Ins(l,4,5)P 3 -generating agonists result in a highly localized rise in cytosolic calcium concentration ([Ca 2+ ] i ) initiated in a region close to the nucleus, whereas caffeine results in a rise in [Ca 2+ ] i throughout the cytoplasm. Our results indicate that chromaffin cells possess two calcium stores with distinct Ca 2+ -ATPases and that the organelle with the 100K Ca 2+ -ATPase is not the Ins(l,4,5)P 3 -sensitive store.