Grant-in-Aid for international Scientific Research
|Allocation Type||Single-year Grants|
|Research Institution||Chiba University School of Medicine|
SEINO Susumu Chiba University, School of Medicine, Professor, 医学部, 教授 (80236067)
THORENS Bemard Institute of Pharmacology and Toxicology, University of Lausanne, Assistant Prof, 薬理学研究所, 助教授
STOFFEL Markus Laboratory of Metabolic Diseases, The Rockefeller University, Assistant Professo, 代謝疾患研究部門, 助教授
BELL Graeme I. Howard Hughes Medical Institute, .Investigator, Departments of Biochemistry and, ハワードヒューズ研究所, 教授
GONOI Tohru Chiba University, Research Center for Pathogenic Fungi, and Microbial Toxicoses,, 助手 (30134365)
INAGAKI Nobuya Akita University School of Medicine, Professor, 医学部, 教授 (30241954)
KANATSUKA Azima Chiba University School of Medicine, Lecturer (40134366)
THORENS Bern ローザンヌ大学, 薬理学研究所, 助教授
GRAEME I.Bel シカゴ大学, ハワードヒューズ研究所, 教授
|Project Period (FY)
1996 – 1997
Completed(Fiscal Year 1997)
|Budget Amount *help
¥11,800,000 (Direct Cost : ¥11,800,000)
Fiscal Year 1997 : ¥5,900,000 (Direct Cost : ¥5,900,000)
Fiscal Year 1996 : ¥5,900,000 (Direct Cost : ¥5,900,000)
|Keywords||Diabetes / Insulin / ATP-sensitive K+ channels. / Genes / Sulfonylurea receptor / Exocytosis / Familial hypoglycemia / ATP感受性K^+-チャネル / イオンチャネル / カルシウム / カリウム / スルホニール尿素剤 / 膵β細胞|
Understanding of the mechanisms of insulin secretion from pancreatic beta-cells should provide a clue to understand the pathogenesis of diabetes mellitus and hypoglycemia.
We have focused the genes encoding proteins expressed in pancreatic beta-cells that are involved in stimulus-secretion coupling in glucose-induced insulin secretion. We have identified two novel genes, Kir6.2 and Noc2, which play important roles in insulin secretion. We have studied the stuctures and functions of these molecules.
The results are summarized as follows.
1.ATP-sensitive K^+ channels. We have cloned a novel member of the inward rectifier K^+ channel family, Kir6.2. Kir6.2 is a protein of 390 amino acids having two transmembrane segments. By coexpression studies of Kir6.2 and a receptor for sulfonylureas (SUR1), widely used in the treatment of non-insulin dependent diabetes mellitus, we have shown that pancreatic beta-cell ATP-sensitive K^+ channel comprises Kir6.2 and SUR1. While SUR1 confers drug and nucle
otide sensitivities, Kir6.2 forms K^+ ion permeable pore domain. An isoform of SUR1, SUR2, was also cloned. We have shown that mutations of SUR1 gene or Kir6.2 gene cause familial hypoglycemia of infancy. We have generated transgenic mice expressing a dominant-negative form of the K_<ATP> channel subunit Kir6.2 [Kir6.2G132S,substitution of glycine (Gly) with serine (Ser) at position 132] in pancreatic beta-cells. Kir6.2G132S transgenic mice develop hypoglycemia with hyperinsulinemia in neonates and hyperglycemia with hypoinsulinemia and decreased beta-cell population in adults. The transgenic mice exhibited a high frequency of apoptotic beta-cells prior to the appearance of hyperglycemia suggesting that the K_<ATP> channel might play a significant role in beta-cell survival in addition to its role in the regulation of insulin secretion.
2. Noc2. We have cloned a cDNA encoding a novel protein of 302 amino acids (designated Noc2, noC2domain)which has 40.7% amino acid identity with and 77.9% similarity to the N-terminal region of rabphilin-3A,a target molecule of Rab3A.However, unlike rabphilin-3A,Noc2 lacks two C2 domains that are thought to interact with Ca^<2+> and phospholipids. Noc2 is expressed predominantly in endocrine tissues including pancreatic beta-cells. Immunoblot analysis of subcellular fractions of the insulin-secreting cell line MIN6 and immunocytochemistry reveal that Noc2 is a 38 kDa protein present in the cytoplasm. Overexpression of Noc2 in PCl2 cells cotransfected with growth hormone (GH) enhances high K^+-induced GH secretion. Screeniy with the yeast two-hybrid system shows that Noc2 interacts with the LIM domain-containing protein zyxin, a component of the cytoskeleton, and this interaction is further confirmed by the coimmunoprecipitation experiment. Accordingly, Noc2 is probably involved in regulated exocytosis in pancreatic beta-cells by interacting with the cytoskeleton. Less