| Project/Area Number |
05680679
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Neurochemistry/Neuropharmacology
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| Research Institution | Tokyo Metropolitan University |
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Principal Investigator |
ISOBE Toshiaki Tokyo Metropolitan Univ, Associate professor, 理学部, 助教授 (70106607)
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| Co-Investigator(Kenkyū-buntansha) |
HANAOKA Kazunori Kitazato Univ.Professor, 理学部, 教授 (40189577)
ISOBE Toshiaki Tokyo Metropolitan Univ, Associate professor (70106607)
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| Project Period (FY) |
1993 – 1994
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| Project Status |
Completed (Fiscal Year 1994)
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| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1994: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1993: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Keywords | Neural circuit / Cerebellar morphogenesis / Cell cycle / Signal transduction / Cerebellar protein / molecular cloning / Nuclear factor / Primary structure / cerebellum / neural circuit / development / gene regulation / brain protein / amino acid sequence / molecular cloning / histochemistry / cell-cycle motif / leucine-rich repeat / histockemistry |
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| Research Abstract |
Morphogenesis of the rat cerebellar cortex, including differentiation of neurons and synaptogenesis, occurs at an early postnatal stage, mainly during the first two weeks after birth. To elucidate the molecular mechanism of this dynamic cellular process, we have developed a data base of rat cerebellar proteins detectable by two-dimensional electrophoresis. The data base contains two-dimensional patterns and derived information pertaining to polypeptide constituents of the developing rat cerebellum. Using this data base, we have compared the protein constituents of a series of immature and mature cerebella. A subset of polypeptides were found to be uniquely expressed in the immature cerebellum. A number of polypeptides of this class were digested by proteinases in situ or after extraction from the gel, and the generated fragments were micro-sequenced by conventional Edman degradation or mass spectrometry combined with liquid chromatography. The sequences were then compared with those in
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protein/gene sequence data bases to identify the isolated proteins. The identified proteins include phosphorylated and non-phosphorylated forms of stathmin whichi has been studied extensively in relation to the differentiation of neurons, and fatty acid-binding protein that belongs to a family of retinoic-acid binding proteins and participates in the neuronal differentiation in chick retina. Two proteins analyzed were unique in that no significant homologies were found to any other sequences in the data bases. The amino acid sequence of these proteins termed V-1 and LANP respectively, were determined by direct protein analysis and cDNA cloning. The sequence showed that the V-1 is a typical cytosolic protein of 117 amino acids and has two and a half of 33 amino acids repeats called "cell-cycle motif". LAMP contains 247 amino acids consisting of teh N-teminally located leucine-rich repeats followed by an acidic amino acids tail with nuclear transfer signal. Both the cell-cycle motif and leucine-rich repeats mediate protein-protein interation through binding to specific target molecules, and are found in numbers of proteins that function in cell fate determination in wide organisms including yeast and Drosophila. In situ histochemical analysis postnatal rat cerebella showed the transient expression of V-1 in granular layr and of LANP in Purkinje cells, suggsting the potential roles of V-1 and LANP in the signal transduction pathway that directs the cerebellar morphogenesis. Less
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