Mikoshiba K, Furuichi T, Miyawaki A, Yoshikawa S, Nakagawa T, Yamada N, Hamanaka Y, Fujino I, Michikawa T, Ryo Y
Department of Molecular Neurobiology, University of Tokyo, Japan.
Philos Trans R Soc Lond B Biol Sci. 1993 Jun 29;340(1293):345-9. doi: 10.1098/rstb.1993.0077.
Inositol 1,4,5-trisphosphate (InsP3) is a second messenger that releases Ca2+ from the intracellular stores. The InsP3 receptor (InsP3-R) was purified and its cDNA was cloned. We have found that InsP3-R is identical to the P400 protein identified as a protein enriched in the cerebellar Purkinje cells. We generated an L fibroblast cell transfectant that produced cDNA derived InsP3-R. The expressed protein displays high affinity and specificity for InsP3. InsP3 induces Ca2+ release from the membrane vesicles of the transfected cells. Incorporation of purified InsP3-R into a lipid bilayer showed InsP3 induced Ca2+ release. These result suggest that InsP3-R is a Ca2+ release channel. Immunogold method using monoclonal antibodies against the receptor showed that it is highly condensed on the smooth surfaced endoplasmic reticulum (ER) and slightly on the outer nuclear membrane and rough ER. Cross linking experiments show that the InsP3-R forms a homotetramer. The approximately 650 N-terminal amino acids are highly conserved between mouse and Drosophila melanogaster, and this region has the critical sequences for InsP3 binding. We found novel subtypes of the InsP3-R resulting from RNA-splicing that are expressed in a tissue-specific and developmentally specific manner and also resulting from different genes. It is believed that there are two Ca2+ release mechanisms, InsP3-induced Ca2+ release (IICR) and Ca(2+)-induced Ca2+ release (CICR). Eggs are good materials to analyse the machanism of Ca2+ signalling: fertilized hamster eggs exhibit repetitive Ca2+ transients as well as the Ca2+ wave.(ABSTRACT TRUNCATED AT 250 WORDS)
肌醇 1,4,5 -三磷酸（InsP3）是一种从细胞内储存中释放 Ca2+ 的第二信使。InsP3 受体（InsP3 - R）已被纯化，其 cDNA 也已被克隆。我们发现 InsP3 - R 与一种被鉴定为在小脑浦肯野细胞中富集的 P400 蛋白相同。我们构建了一个产生源自 cDNA 的 InsP3 - R 的 L 成纤维细胞转染体。所表达的蛋白对 InsP3 显示出高亲和力和特异性。InsP3 诱导转染细胞的膜囊泡释放 Ca2+。将纯化的 InsP3 - R 整合到脂质双分子层中显示 InsP3 诱导 Ca2+ 释放。这些结果表明 InsP3 - R 是一个 Ca2+ 释放通道。使用针对该受体的单克隆抗体的免疫金法表明它高度聚集在光滑表面的内质网（ER）上，在外核膜和粗面 ER 上则较少。交联实验表明 InsP3 - R 形成同四聚体。小鼠和黑腹果蝇之间大约 650 个 N 端氨基酸高度保守，并且该区域具有 InsP3 结合的关键序列。我们发现了由 RNA 剪接产生的 InsP3 - R 的新型亚型，它们以组织特异性和发育特异性的方式表达，并且也由不同基因产生。据信存在两种 Ca2+ 释放机制，即 InsP3 诱导的 Ca2+ 释放（IICR）和 Ca(2+) 诱导的 Ca2+ 释放（CICR）。卵子是分析 Ca2+ 信号传导机制的良好材料：受精的仓鼠卵表现出重复的 Ca2+ 瞬变以及 Ca2+ 波。（摘要截短于 250 字）
