| Project/Area Number |
07044236
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| Research Category |
Grant-in-Aid for international Scientific Research
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| Allocation Type | Single-year Grants |
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| Section | Joint Research |
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| Research Field |
Neurochemistry/Neuropharmacology
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| Research Institution | Niigata University |
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Principal Investigator |
KUWANO Ryozo Niigata University Research Laboratory for Molecular genetics Associate Professor, 遺伝子実験施設, 助教授 (20111734)
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| Co-Investigator(Kenkyū-buntansha) |
HERRUP Karl Case Western Reseve University Professor, Professor
SAUER Brian National Institute of Health Expert, Expert
NISHIYAMA Akiko Cleaveland clinic Foundation Research Associate, Research A
HANAOKA Kazunori Kitasato University Faculty of Science Professor, 理学部, 教授 (40189577)
KUMANISHI Toshiro Niigata University Brain Research Professor, 脳研究所, 教授 (40018601)
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| Project Period (FY) |
1995 – 1997
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| Project Status |
Completed (Fiscal Year 1997)
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| Budget Amount *help |
¥12,600,000 (Direct Cost: ¥12,600,000)
Fiscal Year 1997: ¥3,800,000 (Direct Cost: ¥3,800,000)
Fiscal Year 1996: ¥4,400,000 (Direct Cost: ¥4,400,000)
Fiscal Year 1995: ¥4,400,000 (Direct Cost: ¥4,400,000)
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| Keywords | astrocytes / S100 protein / neuronal function / diphteria toxin / Cre-lox / transgenic mice / gene expression / immunohistochemisty / GFAP |
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| Research Abstract |
We focused on the role of astrocytes in maintaining of neuronal functions and attempted to introduce the strategy of cell ablation using a adiphtheia toxin A fragment (DT-A) gene in transgenic mice.
1) Prior to create transgenic mice astrocyte specific gene expression was examined in primary culture cells prepared from mouse embryos. In the course of this project we purified several proteins involved in synapse formation from never growth cone-enriched fraction, determined the amino acid sequence, prepared specific antibodies against these peptides and analyzed subcellular localization in the cultured cells.
2) Based on spatial-temporal specific gene expression using as appropriate promoter such as S100 or GFAP promoter in astrocyte, the DT-A gene is activated in the restricted cells in the central nervous system. In addition we used cre-lox system for control of the target gene expression. Cre-recombinase mediated site specific recombination occurs on the restricted DNA sequence composed of 34 bp (loxP). DT-A activity was suppressed by insertion of transcription termination signal down stream of murine molony leukemia virus promoter. The DT-A activated by co-transfection with cre-recombinase resulted in cell death.
3) Gene trapping provided novel promoters expressed in the central nervous system. We have found the trapped gene related to mental retardation and characterize an affect of the gene expression on maintaining neuronal functions.
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