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1993 Fiscal Year Final Research Report Summary

Studies on the molecular mechanism for calcium signaling

Research Project

Project/Area Number 05044159
Research Category

Grant-in-Aid for international Scientific Research

Allocation TypeSingle-year Grants
SectionJoint Research
Research InstitutionKyoto University

Principal Investigator

IMOTO Keiji  Kyoto University, Faculty of Medicine, assitant professor, 医学部, 助教授 (00176512)

Co-Investigator(Kenkyū-buntansha) NAKAI Junichi  Kyoto University Faculty of Medicine, 獣医学部・生理学教室, 研究員 (80237198)
STUHMER Walter  Max-Planck-Institute fur experimentalle Medizine, プランク実験医学研究所・分子神経情報伝達部門, 部長
BEAM Kurt G.  Colorado State University, 獣医学部・生理学教室, 教授
MORI Yasuo  Kyoto University Faculty of Medicine, 医学部, 助手 (80212265)
FUJITA Yoshihiko  Kyoto University Faculty of Medicine, 医学部, 助手 (80192730)
Project Period (FY) 1993
KeywordsCalcium channels / cDNA cloning / cDNA expression / Site-specific mutagenesis / Activation kinetics / Small subunits
Research Abstract

Previous studies demonstrated that the difference in speed of activation between the cardiac and skeletal muscle calcium channels can be attributed to the difference in amino acid sequence of the first repeat of the four repeated units of homology. To identify the region in repeat I responsible for the different activation kinetics, we have made series of 'repeat I chimera' DNA constructs systematically, and expressed them in cultured skeletal muscle cells derived from mice with muscular dysgenesis. Analyses of speed of activation have revealed that the third putative transmembrane region (S3) and the linker region between S3 and S4 is critical to determine the speed of activation. Together with the notion that S4 functions as a voltage sensor, we speculate that a change in membrane potential triggers movement of S4, which is transmitted by affecting S3 and its adjacent portion, finally leading to channel openings.
Voltage-gated calcium channels contain smaller subunits. When the main s … More ubunit (alpha1 subunit) is coexpressed with smaller subunits, functional activity is markedly increased. To understand the molecular mechanism of small subunit function, we have expressed cardiac calcium channel alpha1 subunit alone or together with small subunits. From the results of Northern blots, protein gel electrophoresis, dihydropyridine binding assay, and electrophysiological measurements, we concluded that the small subunits do not affect metabolism of the main subunit, and that they are necessary for proper functional conformation.
From cDNA libraries of rabbit brain, we have cloned cDNA encoding a new calcium channel (named BIII). By sequencing the cDNA, we have elucidated the primary structure of BIII calcium channel. This calcium channel showed homology to other voltage-dependent calcium channel. Especially, BIII showed higher similarity to BI and BII brain calcium channel. These three types of calcium channel form a subgroup distinct from dihydropyridine-sensitive L-type calcium channel. When BIII channel is expressed by injecting DNA into cultured skeletal muscle cells derived from mice with muscular dysgenesis, it showed omega-conotoxin sensitive calcium channel activity. Its kinetics and voltage-dependence were similar to those reported for N-type calcium channels. Since the N-type calcium channel is important for neurotransmitter release, cloning of an N-type calcium channel BIII will contribute to future studies of brain function. Less

  • Research Products

    (8 results)

All Other

All Publications (8 results)

  • [Publications] Fujita,Y.et al.: "Primary structure and functional expression of the ω-conotoxin sensitive N-type calcium channel from rabbit brain" Neuron. 10. 585-598 (1993)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nishimura,S.et al.: "Requirement of the calcium channel β subunit for functional conformation" FEBS Lett.324. 283-286 (1993)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Imoto,K.: "Ion channels:molecular basis of ion selectivity(MiniReview)" FEBS Lett.325. 100-103 (1993)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nakai,J.et al.: "Critical roles of the S3 segment and S3-S4 linker of repeat I in activation of L-type calcium channels" Proc.Natl.Acad.Sci.USA. 91. 1014-1018 (1994)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Fujita,Y. et al.: "Primary structure and functional expression of the omega-conotoxin sensitive N-type calcium channel from rabbit brain." Neuron. 10. 585-598 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nishimura,S. et al.: "Requirement of the calcium channel beta subunit for functional conformation" FEBS Lett.324. 283-286 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Imoto,K.: "Ion channels : molecular basis of ion selectivity (Mini Review)" FEBS Lett.325. 100-103 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nakai,J. et al.: "Critical roles of the S3 segment and S3-S4 linker of repeat l in activation of L-type calcium channels." Proc. Natl. Acad. Sci. USA. 91. 1014-1018 (1994)

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

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Published: 1995-02-07  

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