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1990 Fiscal Year Final Research Report Summary

Insulin Resistance Due to Unprocessed Insulin Proreceptor

Research Project

Project/Area Number 63480269
Research Category

Grant-in-Aid for General Scientific Research (B)

Allocation TypeSingle-year Grants
Research Field 内分泌・代謝学
Research InstitutionShiga University of Medical Science

Principal Investigator

KOBAYASHI Masashi  Shiga University of Medical Science, Assistant Professor., 医学部, 講師 (80115758)

Co-Investigator(Kenkyū-buntansha) TERAOKA Hiroshi  Shionogi Research Institute Senior Researcher., 主任研究員
KOBAYASHI Masashi  Shiga University of Medical Science, Assistant Professor. (80115758)
Project Period (FY) 1988 – 1990
KeywordsInsulin receptor disease / PCR / point mutation / Insulin receptor / Insulin resistance
Research Abstract

We reported the siblings with severe insulin resistance because of unprocessed insulin proreceptors. To determine the structural abnormalities, we analyzed DNA sequence of the cleavage site by direct sequence method using PCR technique. The structural change of the cleavage site from Arg-Lys-Arg-Arg to Arg-Lys-Arg-Ser due to G-T point mutation appeared to be the cause for failure to process the proreceptors. To study whether the mutation of insulin proreceptors at the cleavage site was responsible for unprocessed insulin proreceptors and to elucidate the structural and binding characteristics of the proreceptors, we performed transfection of cDNA with the mutation in COS 7 cells and examined the expressed insulin receptors. At 72 hours after transfection, insulin binding increased to the maximum in both cells transfected with normal cDNA or mutated cDNA, and they were 40 times and 14 times of control, non-transfected cells, respectively. Affinity cross-linking and surface labeling showed 135 kDa normal alpha subunit in cells transfected with normal cDNA and 210 kDa proreceptor in mutant cells. The proreceptors were cleaved by trypsin to alpha and beta subunit at the concentration of 0.025%. Auto-phosphorylation study showed decreased ^<32>P incorporation to proreceptors of cells transfected with mutated cDNA at both basal and insulin stimulated states without change in the sensitivity to insulin compared with cells transfected with normal cDNA. Competitive binding study with insulin, proinsulin and miniproinsulin showed that the proreceptors had lower relative affinity to proinsulin, but this characteristics disappeared by trypsin treatment. These results suggest that the mutation was the cause for unprocessed insulin proreceptors in the patients with insulin resistance, and that the expressed proreceptors of the COS 7 cells transfected with the mutated cDNA had similar binding specificity and sensitivity to trypsin.

  • Research Products

    (14 results)

All Other

All Publications (14 results)

  • [Publications] M.Kobayashi,M.Sugibayashi,T.Sasaoka,K.Egawa Y,Sigeta,M.Tamaki,E.Nakamura,H.Teraoka.: "Transfection of cDNA with GーT point mutation at the cleavage site of insulin receptors to COC 7 cells." Biochem Biophys Res Commun. 167. 1073-78 (1990)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] H.Maegawa,M.Kobayashi K.Egawa,D.A.McClain,J.M.Olefsky,Y.Shigeta: "The ligand characteristics of a kinaseーdefective A/K]0]8 human insulin receptor expressed in Rat 1 fibrolasts." Biochim Biophys Acta. 1053. 185-88 (1990)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M.Sugibayashi,Y.Shigeta,H.Teraoka^★,M.Kobayashi: "Characterization of unprocessed insulin proreceptors in cos 7 cells transfected with cDNA with Arg735ーSer735point mutation at the cleavage site." Metabolism. (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] H.Maegawa,Y.Shigeta K.Egawa,M.Kobayashi.: "Impaired Autophosphorylation of Insurin Receptors from Abdominal Skeletal Muscles in Nonーobese Subjects with NIDDM." Diabetes. (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] 杉林 正章・小林 正・繁田 幸男: "遺伝子塩基配列の解析およびそのトランスフェクションによる発現と機能特性" 糖尿病. 34. 113-124 (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M.Kobayashi,T.Sasaoka,Y.Takata,O.Ishibashi,M.Sugibayashi,A.Hisatomi.: "Unprocessed insulin proreceptors due to point mutation at the cleavage site." Diabetes Res Clin Prac. 7. 35-9 (1989)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M.Kobayashi,M.Iwanishi,K.Egawa,H.Maegawa,Y.Shigeta.: "The proceeding of diabetes and.obesity." A new oral antidiabetic agent(CSー045)increases kinase activity of insulin receptors from skeletal muscles of zucker fatty rats., (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M.kobayashi,T.Sasaoka,M.Sugibayashi,Y.Shigeta.: "Recent advance in insulin action and its disorders." Insulin resistance by unprocessed insulir proreceptors due to point mutation at the cleavage site., (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kobayashi M, et al.: "Transfection of cDNA with G-T point mutation at the cleavage site of insulin receptors to COS 7 cells." Biochem Biophys Res Commun. 167 ;. 1073-78 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Maegawa H, Kobayashi M, et al.: "The ligand characteristics of a kinase-defective A/K 1018 human insulin receptor expressed in Rat 1 fibroblasts." Biochim Biophys Acta. 1053. 185-88, (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Sugibayashi M, Shigeta Y, Teraoka H, Kobayashi M.: "Characterization of un-processed insulin proreceptors in COS 7 cells transfected with cDNA with Arg^<735>-Ser^<735> point mutation at the cleavage site." Metabolism.

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Maegawa H, Shigeta Y, Egawa K, Kobayashi M.: "Impaired autophosphorylation of insulin receptors from abdominal skeletal muscles in non-obese subjects with NIDDM." Diabetes.

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kobayashi M, Sasaoka, T, Takata Y, et al.: "Unprocessed insulin proreceptors due to point mutation at the cleavage site." Diabetes Res Clin Prac. 7 :. 35-9, (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kobayashi M et al.: "Insulin resistance by unprocessed insulin pro receptors due to point mutation at the cleavage site." Recent advances in insulin action and its disorders. (Ed. Kobayashi Met al) Elasevier Science publishers.(1991)

    • Description
      「研究成果報告書概要(欧文)」より

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Published: 1993-08-12  

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