Berven L A, Crouch M F, Katsis F, Kemp B E, Harland L M, Barritt G J
Department of Medical Biochemistry, School of Medicine, Flinders University, Adelaide, South Australia.
J Biol Chem. 1995 Oct 27;270(43):25893-7. doi: 10.1074/jbc.270.43.25893.
The role of a trimeric GTP-binding protein (G-protein) in the mechanism of vasopressin-dependent Ca2+ inflow in hepatocytes was investigated using both antibodies against the carboxyl termini of trimeric G-protein alpha subunits, and carboxyl-terminal alpha-subunit synthetic peptides. An anti-Gi1-2 alpha antibody and a Gi2 alpha peptide (Gi2 alpha) Ile345-Phe355), but not a Gi3 alpha peptide (Gi3 alpha Ile344-Phe354), inhibited vasopressin- and thapsigargin-stimulated Ca2+ inflow, had no effect on vasopressin-stimulated release of Ca2+ from intracellular stores, and caused partial inhibition of thapsigargin-stimulated release of Ca2+. An anti-Gq alpha antibody also inhibited vasopressin-stimulated Ca2+ inflow and partially inhibited vasopressin-induced release of Ca2+ from intracellular stores. Immunofluorescence measurements showed that Gi2 alpha is distributed throughout much of the interior of the hepatocyte as well as at the periphery of the cell. By contrast, Gq/11 alpha was found principally at the cell periphery. It is concluded that the trimeric G-protein, Gi2, is required for store-activated Ca2+ inflow in hepatocytes and acts between the release of Ca2+ from the endoplasmic reticulum (presumably adjacent to the plasma membrane) and the receptor-activated Ca2+ channel protein(s) in the plasma membrane.
利用针对三聚体G蛋白α亚基羧基末端的抗体以及羧基末端α亚基合成肽，研究了三聚体GTP结合蛋白（G蛋白）在肝细胞中血管加压素依赖性Ca2+内流机制中的作用。抗Gi1-2α抗体和Gi2α肽（Gi2α Ile345-Phe355），而非Gi3α肽（Gi3α Ile344-Phe354），抑制了血管加压素和毒胡萝卜素刺激的Ca2+内流，对血管加压素刺激的细胞内钙库Ca2+释放没有影响，并部分抑制了毒胡萝卜素刺激的Ca2+释放。抗Gqα抗体也抑制了血管加压素刺激的Ca2+内流，并部分抑制了血管加压素诱导的细胞内钙库Ca2+释放。免疫荧光测量显示，Gi2α分布于肝细胞内部的大部分区域以及细胞周边。相比之下，Gq/11α主要位于细胞周边。得出的结论是，三聚体G蛋白Gi2是肝细胞中储存激活的Ca2+内流所必需的，并且在Ca2+从内质网（可能与质膜相邻）释放与质膜中受体激活的Ca2+通道蛋白之间起作用。
