1998 年度実績報告書

情報伝達機能分子の細胞内動態の画像化とレーザー光分子活性化法による解析技術開発

研究課題

研究課題/領域番号	10558112
研究機関	東京大学
研究代表者	御子柴克彦東京大学, 医科学研究所, 教授 (30051840)
研究分担者	道川貴章東京大学, 医科学研究所, 助手 (90282516)
キーワード	マイクロCALI / 機能分子不活化法 / GFP / 蛍光エネルギー転移 / IP3
研究概要	マイクロCALIでは、1〜2μm位の小さなスポットでの不活性化しかできないため、より高いエネルギーのCALIを組み立てて、細胞の機能分子を不活性化させる技術開発をおこなう。高エネルギー基準にするため、新しいレーザー光源の購入が必須となる。また、光源のスポットの均一性、スポットの形状の調査を必要とする。我々は、何らconfactorを必要とせずに、励起によって蛍光を発する蛋白質、GFP(green-flourescence-protein)に注目している。これをコードする遺伝子は既にとられ、遺伝子工学的手を用いてGFPを他の蛋白質分子に融合して細胞内で発現することが可能になった。 IP3の結合がIP3結合蛋白質に高次構造変化をもたらすことがゲル濾過実験により証明された。蛍光ラベルしたIP3結合蛋白質の構造変化を蛍光エネルギー転移を利用して検出する技術を開発し、更に細胞内導入によって細胞内IP3濃度をリアルタイムでモニターする。最近米国のTsien博士の研究室では、GFPを改変するプロジェクトを進めている。様々な改変GFP蛋白質は、異なる吸収、蛍光スペクトラムを示し、蛍光エネルギー転移を可能にする幾つかの組み合わせが見つかっている。我々は、これら改変GFP遺伝子をTsien博士と共同で、IP3結合蛋白質の両末端に連結させる研究を進行中である。

研究成果

(23件)

すべてその他

すべて文献書誌 (23件)

[文献書誌] Aruga,J.et.al.: "Mouse Zic1 is involved in cerebeller development." J.Neurosci.18. 284-293 (1998)
[文献書誌] Monkawa,T.et.al.: "Localization of inositol 1,4,5-trisphophate receptors in the rat kidney." Kidney Int.53. 296-301 (1998)
[文献書誌] Zhao,H.et.al.: "Functional expression of a mammalian odorant receptor." Science. 279. 237-242 (1998)
[文献書誌] Sato,Y.et.al.: "Adenophostin,a potent agonist of the inositol 1,4,5-trisphoshate receptor,is useful for fertilization of mouse oocytes injected with round spermatids leading to normal offspring." Biology of reproduction. 58. 867-873 (1998)
[文献書誌] Miyamura,T.et.al.: "Metabolic labeling of a subset of glia cells by UDP-galactose: implication for astrocyte lineage diversity." J.Neurosci.Res.52. 173-183 (1998)
[文献書誌] Yamamoto-Hino,M.: "Apical vesicles bearing inositol 1,4,5-trisphosphate receptors in the Ca2+ initiation site of ductal epithelium of submandibular gland." J.Cell.Biol.141. 135-142 (1998)
[文献書誌] Ibata,K.et.al.: "Inositol 1,3,4,5-Tetrakisphosphate binding activities of neural and non-neural synaptotagmins." J.Biol.Chem.273. 12267-12273 (1998)
[文献書誌] Inoue,T.et.al.: "Type 1 inositol 1,4,5-trisphosphate recetor is required for induction of long-term depression in cerebellar purkinje neurons." J.Neurosci.18. 5366-5373 (1998)
[文献書誌] Mathias,R.S.et.al.: "IP3 receptor blockade fails to prevent intracellular Ca2+ release by ET-1 and a-thrombin." Am.J.Physiol.274. C1456-C1465 (1998)
[文献書誌] Hirota,J.et.al.: "T-cell receptor signalling in the inositol 1,4,5-trisphosphate receptor(IP3R) type-1-deficient mice : is IP3R type 1 essential for T-cell-receptor signalling?" Biochem.J.333. 615-619 (1998)
[文献書誌] Fukuzono,S.et.al.: "Overproduction and immuno-affinity purification of myelin proteolipidprotein (PLP),aninositol hexakisphosphate binding protein,in a baculovirus expression system." Biochem.Biophys.Res.Commun.249. 66-72 (1998)
[文献書誌] Muto,A.et.al.: "Activation of inositol 1,4,5-trisphosphate recetors induces transient changes in cell shape of fertilized Xenopus eggs." Cell Motility and the Cytoskeleton. 39. 201-208 (1998)
[文献書誌] Nakata,K.et.al.: "Xenopus Zic family and its role in neural and neural crest development." Mechanisms of Development. 75. 43-51 (1998)
[文献書誌] Nakayama,T.et.al.: "Hemizygous deletion of the HPC-1/syntaxin 1A gene(STX1A)in patient with Williams syndrome." Cytogenet Cell Genet.82. 49-51 (1998)
[文献書誌] Yoshida,M.et.al.: "Role of two series of Ca2+ oscillations in activation of ascidian eggs." Developmenatal Biology. 203. 122-133 (1998)
[文献書誌] Futatsugi,A.et.al.: "Muscle-specific mRNA isoform encodes a protein composed mainly of the N-terminal 175 residues of type 2 lns(1,4,5)P3 receptor." Biochem.J.334. 559-563 (1998)
[文献書誌] Hata,R.et.al.: "Brainstem auditory evoked potentials during brainstem ischemia and reperfusion in gerbils." Neuroscience. 83. 201-213 (1998)
[文献書誌] Takei,K.et.al.: "Regulation of nerve growth mediated by inositol 1,4,5-trisphosphate receptor in growth cones." Science. 282. 1705-1708 (1998)
[文献書誌] Tamura,H.et.al.: "Molecular cloning and expression of cDNA encoding an olfactory-specific mouse phenol sulphotransferase." Biochem.J.331. 953-958 (1998)
[文献書誌] Yoshikawa,F.et.al.: "Cooperative formation of the ligand-binding site of the inositol 1,4,5-trisphosphate receptor by two separable domains." J.Biol.Chem.274. 328-334 (1999)
[文献書誌] Yoshikawa,F.et.al.: "Trypsinized cerebellar inositol 1,4,5-trisphosphate receptor." J.Biol.Chem.274. 316-327 (1999)
[文献書誌] Kabayama,H.et.al.: "Functional involvement of synaptotagmin I/II C2A domain in neurite outgrowth of chick dorsal root ganglion neuron." Neuroscience. 88. 999-1003 (1999)
[文献書誌] Zhu,C.et.al.: "Inosiotl 1,4,5-trisphosphate receptor down-regulation is activated directly by inositol 1,4,5-trisphoshate binding." J.Biol.Chem.274. 3476-3484 (1999)

1998 年度 実績報告書

情報伝達機能分子の細胞内動態の画像化とレーザー光分子活性化法による解析技術開発

研究代表者

御子柴 克彦 東京大学, 医科学研究所, 教授 (30051840)

研究成果

[文献書誌] Aruga,J.et.al.: "Mouse Zic1 is involved in cerebeller development." J.Neurosci.18. 284-293 (1998)

[文献書誌] Monkawa,T.et.al.: "Localization of inositol 1,4,5-trisphophate receptors in the rat kidney." Kidney Int.53. 296-301 (1998)

[文献書誌] Zhao,H.et.al.: "Functional expression of a mammalian odorant receptor." Science. 279. 237-242 (1998)

[文献書誌] Sato,Y.et.al.: "Adenophostin,a potent agonist of the inositol 1,4,5-trisphoshate receptor,is useful for fertilization of mouse oocytes injected with round spermatids leading to normal offspring." Biology of reproduction. 58. 867-873 (1998)

[文献書誌] Miyamura,T.et.al.: "Metabolic labeling of a subset of glia cells by UDP-galactose: implication for astrocyte lineage diversity." J.Neurosci.Res.52. 173-183 (1998)

[文献書誌] Yamamoto-Hino,M.: "Apical vesicles bearing inositol 1,4,5-trisphosphate receptors in the Ca2+ initiation site of ductal epithelium of submandibular gland." J.Cell.Biol.141. 135-142 (1998)

[文献書誌] Ibata,K.et.al.: "Inositol 1,3,4,5-Tetrakisphosphate binding activities of neural and non-neural synaptotagmins." J.Biol.Chem.273. 12267-12273 (1998)

[文献書誌] Inoue,T.et.al.: "Type 1 inositol 1,4,5-trisphosphate recetor is required for induction of long-term depression in cerebellar purkinje neurons." J.Neurosci.18. 5366-5373 (1998)

[文献書誌] Mathias,R.S.et.al.: "IP3 receptor blockade fails to prevent intracellular Ca2+ release by ET-1 and a-thrombin." Am.J.Physiol.274. C1456-C1465 (1998)

[文献書誌] Hirota,J.et.al.: "T-cell receptor signalling in the inositol 1,4,5-trisphosphate receptor(IP3R) type-1-deficient mice : is IP3R type 1 essential for T-cell-receptor signalling?" Biochem.J.333. 615-619 (1998)

[文献書誌] Fukuzono,S.et.al.: "Overproduction and immuno-affinity purification of myelin proteolipidprotein (PLP),aninositol hexakisphosphate binding protein,in a baculovirus expression system." Biochem.Biophys.Res.Commun.249. 66-72 (1998)

[文献書誌] Muto,A.et.al.: "Activation of inositol 1,4,5-trisphosphate recetors induces transient changes in cell shape of fertilized Xenopus eggs." Cell Motility and the Cytoskeleton. 39. 201-208 (1998)

[文献書誌] Nakata,K.et.al.: "Xenopus Zic family and its role in neural and neural crest development." Mechanisms of Development. 75. 43-51 (1998)

[文献書誌] Nakayama,T.et.al.: "Hemizygous deletion of the HPC-1/syntaxin 1A gene(STX1A)in patient with Williams syndrome." Cytogenet Cell Genet.82. 49-51 (1998)

[文献書誌] Yoshida,M.et.al.: "Role of two series of Ca2+ oscillations in activation of ascidian eggs." Developmenatal Biology. 203. 122-133 (1998)

[文献書誌] Futatsugi,A.et.al.: "Muscle-specific mRNA isoform encodes a protein composed mainly of the N-terminal 175 residues of type 2 lns(1,4,5)P3 receptor." Biochem.J.334. 559-563 (1998)

[文献書誌] Hata,R.et.al.: "Brainstem auditory evoked potentials during brainstem ischemia and reperfusion in gerbils." Neuroscience. 83. 201-213 (1998)

[文献書誌] Takei,K.et.al.: "Regulation of nerve growth mediated by inositol 1,4,5-trisphosphate receptor in growth cones." Science. 282. 1705-1708 (1998)

[文献書誌] Tamura,H.et.al.: "Molecular cloning and expression of cDNA encoding an olfactory-specific mouse phenol sulphotransferase." Biochem.J.331. 953-958 (1998)

[文献書誌] Yoshikawa,F.et.al.: "Cooperative formation of the ligand-binding site of the inositol 1,4,5-trisphosphate receptor by two separable domains." J.Biol.Chem.274. 328-334 (1999)

[文献書誌] Yoshikawa,F.et.al.: "Trypsinized cerebellar inositol 1,4,5-trisphosphate receptor." J.Biol.Chem.274. 316-327 (1999)

[文献書誌] Kabayama,H.et.al.: "Functional involvement of synaptotagmin I/II C2A domain in neurite outgrowth of chick dorsal root ganglion neuron." Neuroscience. 88. 999-1003 (1999)

[文献書誌] Zhu,C.et.al.: "Inosiotl 1,4,5-trisphosphate receptor down-regulation is activated directly by inositol 1,4,5-trisphoshate binding." J.Biol.Chem.274. 3476-3484 (1999)

1998 年度実績報告書

御子柴克彦東京大学, 医科学研究所, 教授 (30051840)