title: Functional co-localization of transfected Ca(2+)-stimulable adenylyl cyclases with capacitative Ca2+ entry sites.
abstract: Three adenylyl cyclases (ACI, ACIII, and ACVIII) have been described, which are putatively Ca(2+)-stimulable, based on in vitro assays. However, it is not clear that these enzymes can be regulated by physiological rises in [Ca2+]i when expressed in intact cells. Furthermore, it is not known whether transfected adenylyl cyclases might display the strict requirement for capacitative Ca2+ entry that is shown by the Ca(2+)-inhibitable ACVI, which is indigenous to C6-2B glioma cells (Chiono, M., Mahey, R., Tate, G., and Cooper, D. M. F. (1995) J. Biol. Chem. 270, 1149-1155). In the present study, ACI, ACIII, and ACVIII were heterologously expressed in HEK 293 cells, and conditions were devised that distinguished capacitative Ca2+ entry from both internal release and nonspecific elevation in [Ca2+]i around the plasma membrane. Remarkably, not only were ACI and ACVIII largely insensitive to Ca2+ release from stores, but they were robustly stimulated only by capacitative Ca2+ entry and not al all by a substantial increase in [Ca2+]i at the plasma membrane elicited by ionophore. (ACIII, reflecting its feeble in vitro sensitivity to Ca2+, was unaffected by any [Ca2+]i rise.) These results suggest a quite unsuspected, essential association of Ca(2+)-sensitive adenylyl cyclases with capacitative Ca2+ entry sites, even when expressed heterologously.
authors.affiliation: Department of Pharmacology and Neuroscience Program, University of Colorado Health Sciences Center, Denver 80262, USA.
authors.forename: K A
authors.initials: KA
authors.lastname: Fagan
authors.forename: R
authors.initials: R
authors.lastname: Mahey
authors.forename: D M
authors.initials: DM
authors.lastname: Cooper
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D004789:Enzyme Activation; D006801:Humans; D017136:Ion Transport; D007527:Isoenzymes / Q000378:metabolism*; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D007527:Isoenzymes; D000262:Adenylyl Cyclases; D002118:Calcium