Differences among type I, II, and III inositol-1,4,5-trisphosphate receptors in ligand-binding affinity influence the sensitivity of calcium stores to inositol-1,4,5-trisphosphate.

Type I, II, and III inositol-1,4,5-trisphosphate (InsP3) receptors are expressed selectively in different cell lines and tissues. We examined whether type I, II, and III InsP3 receptors differ in ligand-binding affinity and whether such differences influence the sensitivity of Ca2+ stores to InsP3. Initially, SH-SY5Y human neuroblastoma cells, AR4-2J rat pancreatoma cells, and RINm5F rat insulinoma cells were studied because these cells express predominantly (>85%) type I, II, and III receptors, respectively. Immunopurification of receptors from these cell lines and measurement of InsP3 binding revealed that the rank order of affinity for InsP3 was type I > type II > type III (binding sites were half-maximally saturated at 1.5, 2.5, and 22.4 nM InsP3, respectively). Examination of Ca2+ store mobilization in permeabilized cells showed that InsP3 was equipotent in SH-SY5Y and AR4-2J cells but was approximately 5-fold less potent in RINm5F cells. In contrast, Ca2+ uptake and InsP3-independent Ca2+ release were very similar in the three cell types. The binding affinity of InsP3 in permeabilized SH-SY5Y, AR4-2J, and RINm5F cells correlated well with its potency as a Ca2+-mobilizing agent and with binding affinity to immunopurified type I, II, and III receptors. Thus, InsP3 receptor binding affinity seems to influence the potency of InsP3 as a Ca2+-mobilizing agent. Finally, immunopurification of type I, II, and III receptors from rat tissues revealed that the affinity differences seen in receptors purified from cultured cells are paralleled in vivo. In combination, the data from cell lines and rat tissues reveal that type I, II, and III receptors bind InsP3 with Kd values of approximately 1, approximately 2, and approximately 40 nM, respectively, and that the selective expression of a particular receptor type will influence the sensitivity of cellular Ca2+ stores to InsP3.