Inhibition by calyculin A and okadaic acid of the Ca(2+) release-activated Ca(2+) entry pathway in rat basophilic leukemia cells: evidence for regulation by type 1/2A serine/threonine phosphatase activity.
Using a combination of fluorescence measurements of intracellular Ca(2+) ion concentration ([Ca(2+)](i)) and membrane potential we have investigated the sensitivity to serine/threonine phosphatase inhibition of Ca(2+) entry stimulated by activation of the Ca(2+) release-activated Ca(2+) (CRAC) entry pathway in rat basophilic leukemia cells. In both suspension and adherent cells, addition of the type 1/2A phosphatase inhibitor calyculin A, during activation of CRAC uptake, resulted in a fall in [Ca(2+)](i) to near preactivation levels. Pre-treatment with calyculin A abolished the component of the Ca(2+) rise associated with activation of CRAC uptake and inhibited Mn(2+) entry, consistent with a requirement of phosphatase activity for activation of the pathway. Depletion of intracellular Ca(2+) stores is accompanied by a large depolarisation which is absolutely dependent upon Ca(2+) entry via the CRAC uptake pathway. Application of calyculin A or okadaic acid, a structurally unrelated phosphatase antagonist inhibits this depolarisation. Taken in concert, these data demonstrate a marked sensitivity of the CRAC entry pathway to inhibition by calyculin A and okadaic acid.
Evans
Nicholas E
NE
Department of Physiology, University of Cambridge, Downing Street, Cambridge CB2 3EG, UK.
Forth
Mark K L
MK
Simpson
Anna K
AK
Mason
Michael J
MJ
D000818:Animals; D001896:Boron Compounds / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects*; D002413:Cations, Divalent / Q000378:metabolism; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D017136:Ion Transport / Q000187:drug effects; D015471:Leukemia, Basophilic, Acute; D008345:Manganese / Q000378:metabolism; D008387:Marine Toxins; D008564:Membrane Potentials / Q000187:drug effects; D019319:Okadaic Acid / Q000494:pharmacology*; D010080:Oxazoles / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D054645:Protein Phosphatase 1; D051381:Rats; D019284:Thapsigargin / Q000494:pharmacology
D001896:Boron Compounds; D002413:Cations, Divalent; D004791:Enzyme Inhibitors; D008387:Marine Toxins; D010080:Oxazoles; D019319:Okadaic Acid; D008345:Manganese; D019284:Thapsigargin; C059041:calyculin A; C109986:2-aminoethoxydiphenyl borate; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D002118:Calcium