2 CALCIUM-BINDING SITES MEDIATE THE INTERCONVERSION OF LIVER INOSITOL 1,4,5-TRISPHOSPHATE RECEPTORS BETWEEN 3 CONFORMATIONAL STATES

Cytosolic CA(2+) biphasically regulates Ins(1,4,5)P-3-stimulated Ca2+ mobilization in liver [Marshall and Taylor (1993) J. Biol. Chem. 268, 13214-13220]. We have investigated the mechanisms underlying this biphasic control of Ca2+ mobilization in permeabilized hepatocytes by comparing the effects of Sr2+, Ba2+ and Ca2+ on the liver Ins(1,4,5)P-3 receptor. Both Ca2+ and Sr2+ increased the binding of [H-3]Ins(1,4,5)P-3 to liver membranes by converting receptors from a low-affinity (K-D similar to 35 nM) to a high-affinity (K-D similar to 5 nM) state. Ba2+ (less than or equal to 20 mu M) did not affect [H-3]Ins(1,4,5)P-3 binding. At concentrations similar to those that caused an enhancement of [H-3]Ins(1,4,5)P-3 binding, Sr2+ (EC(50) = 570 nM) and Ca2+ (EC(50) = 200 nM) increased the sensitivity of the intracellular Ca2+ stores to Ins(1,4,5)P-3. Further modest elevations in [Ca2+] (EC(50) = 1.5 mu M) inhibited Ins(1,4,5)P-3-stimulated Ca2+ mobilization, whereas Sr2+ caused inhibition only when its concentration was very substantially increased (EC(50) similar to 900 mu M). Sr2+ is therefore only 3-fold less potent than Ca2+ in causing sensitization of Ins(1,4,5)P-3-stimulated Ca2+ release, but 600-fold less potent in causing inhibition. Ba2+ neither sensitized ([Ba2+] less than or equal to 20 mu M) nor inhibited ([Ba2+] less than or equal to 1 mM) Ins(l,4,5)P-3-stimulated Ca2+ release, and did not inhibit either the sensitization of Ca2+ release evoked by Sr2+ or the inhibition of Ca2+ release evoked by Ca2+. Our results suggest that two distinct Ca2+-binding sites, which differ in their selectivities for bivalent cations. mediate the interconversion of Ins(1,4,5)P-3 receptors between at least three different conformational states. These two Ca2+-binding sites, which may reside either on the Ins(1,4,5)P-3 receptor itself or on distinct regulatory proteins, can be distinguished by their different selectivities for bivalent cations.