Differential regulation of Ca2+ release-activated Ca2+ influx by heterotrimeric G proteins.
The least understood aspect of the agonist-induced Ca2+ signal is the activation and regulation of the Ca2+ release-activated Ca2+ influx (CRAC) across the plasma membrane. To explore the possible role of heterotrimeric G proteins in the various regulatory mechanisms of CRAC, continuous renal epithelial cell lines stably expressing alpha 13 and the constitutively active alpha qQ209L were isolated and used to measure CRAC activity by the Mn2+ quench technique. Release of intracellular Ca2+ by agonist stimulation or thapsigargin was required for activation of CRAC in all cells. Although the size of the internal stores was similar in all cells, CRAC was 2-3-fold higher in alpha 13- and alpha qQ209L-expressing cells. However, the channel was differentially regulated in the two cell types. Incubation at low [Ca2+]i, inhibition of the NOS pathway, or inhibition of tyrosine kinase inhibited CRAC activity in alpha 13 but not alpha qQ209L cells. Treatment with okadaic acid prevented inhibition of the channel by low [Ca2+]i and the protein kinase inhibitors in alpha 13 cells. These results suggest that expression of alpha qQ209L dominantly activates CRAC by stabilizing a phosphorylated state, whereas expression of alpha 13 makes CRAC activation completely dependent on phosphorylation by several kinases. G proteins may also modulate CRAC activity independently of the phosphorylation/dephosphorylation state of the pathway to increase maximal CRAC activity. Furthermore, our results suggest a general mechanism for regulation of CRAC that depends on coupling of receptors to specific G proteins.
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002461:Cell Line, Transformed; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000502:physiology*; D019833:Genistein; D007529:Isoflavones / Q000494:pharmacology; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000378:metabolism; D051379:Mice; D009569:Nitric Oxide / Q000502:physiology; D010766:Phosphorylation
D007529:Isoflavones; D009569:Nitric Oxide; D019833:Genistein; D019204:GTP-Binding Proteins; D002118:Calcium