1997 Fiscal Year Final Research Report Summary
Analysis of the mechanisms of neuronal cell-type diversification by the transcription factor UNC-86
| Project/Area Number |
08680847
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Neurochemistry/Neuropharmacology
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| Research Institution | Tokyo Women's Medical College |
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Principal Investigator |
MITANI Shohei Tokyo Women's Medical College, Department of Medicine, Associate Professor, 医学部, 助教授 (90192757)
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| Project Period (FY) |
1996 – 1997
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| Keywords | nematode / neurodevelopment / transcription factors / epitope-tagging |
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| Research Abstract |
We developed a novel method (epitope-tagging of target genes of transcription factors) to isolate in vivo binding sites of transcription factors in the nematode C.elegans. Generally, this kind of experiments are very difficult so far. We cloned the whole DNA fragment for unc-86 gene. We inserted GFP (green fluorescent protein) cDNA in frame to the coding region of the transcription factor, and used the DNA construct to rescue unc-86 mutant phenotypes. We isolated nuclei from such a strain, cross-linked the nuclear protein to DNA.Then, the nuclei were fragmented to nucleosomes. The UNC-86/GFP-target DNA sequence complexes were immuno-purified with an anti-GFP monoclonal antibody. The DNA fragments isolated in this way were cloned and sequenced. So far, 29 unique sequences corresponding to certain genome 1oci on the C.elegans chromosomes were identified using the database by C.elegans genome sequence consortium. Twenty-four of which appeared regulatory sequences of certain genes. Four of 24 corresponding to known genes, three of which are unc-86 gene itself and one known as a gene which works downstream of unc-86 in a genetic pathway for HSN neuron differentiation. Other genes are categorized into two. First are genes sharing somehow homologies to known genes which can work in nervous systems. Others didn't share any homology to known genes and are to be analyzed in the future on the possible functions in the nervous systems.
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