Regulatory mechanism of intracellular Ca^<2+> dynamics

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The inositol 1,4,5-trisphosphate receptor (IP_3R) exisits as a tetrameric complex to form a functional inositol 1,4,5-trisphosphate-gated Ca^<2+> channel. Molecular cloning studies have shown that there are at least three types of IP_3R subunits, designated type 1, type 2, and type 3. The levels of expression of IP_3R subunits in various cell lines were investigated by Western blot analysis using type-specific antibodies against 15 C-terminal amino acids of each IP_3R subunits. We found that all the three types of IP_3R subunits were expressed in each cell line examined, but their levels of expression varied. To determine whether IP_3R from heterotetramers, we employed immunoprecipitation experiments using Chinese hamster ovary cells (CHO-K1 cells), in which all three types are abundantly expressed. Each type-specific antibody immunoprecipitated not only the respective cognate type but also the other two types. This result suggests that distinct types of IP_3R subunits assemble to form … More heterotetramers in CHO-K1 cells. We also detected heterotetramers in rat liver, in which IP_3R type 1 and type 2 are expressed abundantly. Previous studies have shown some functional differences among IP_3R types, suggesting the possibility that various compositions of submits show distinct channel properties. The diversity of IP_3R channels may be further increased by the co-assembly of different IP_3R subunits to form homo-or heterotetramers.Kinetics of Ca^<2+> release by adenophostin, a novel gaonist of inositol 1,4,5-trisphosphate (IP_3) receptor, in the purified and reconstituted IP_3 receptor type 1 (IP_3R1) was investigated using the fluorescent Ca^<2+> indicator fluo-3. Submaximal concentrations of adenophostin caused quantal Ca^<2+> release from the purified IP_3R1 as IP_3 did. Adenophostin-induced Ca^<2+> release by the purified IP_3R1 exhibited a high positive cooperativity (nH=3.9(]SY.+-.])0.2, EC_<50>=11 nM), whereas the IP_3-induced Ca^<2+> release exhibited a moderate one (nH=1.8(]SY.+-.])0.1, EC_<50>=1100 nM). Inhibitation of [^3H] IP_3 binding to the purified IP_3R1 by adenophostin exhibited a positive cooperativity (nH=1.9, K_i=10 nM), whereas IP_3 did not (nH=1.1, K_i=41 nM). Less