| Project/Area Number |
11680670
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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 |
Molecular biology
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| Research Institution | University of Tsukuba |
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Principal Investigator |
KOBAYASHI Makoto Institute of Basic Medical Sciences, Assistant Professor, 基礎医学系, 講師 (50254941)
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| Co-Investigator(Kenkyū-buntansha) |
YAMAMOTO Masayuki Inst.Basic Med.Sci, Univ.Tsukuba, Professor, 基礎医学系, 教授 (50166823)
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| Project Period (FY) |
1999 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥3,900,000 (Direct Cost: ¥3,900,000)
Fiscal Year 2000: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 1999: ¥2,600,000 (Direct Cost: ¥2,600,000)
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| Keywords | Zebrafish / GATA-1 / Erythropoiesis / Transcriptional Regulation / GFP / Transgenic fish / Positive autoregulation / GTAT-1 / 橋渡し分子 |
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| Research Abstract |
Transcription factor GATA-1 is known to interact with various factors and form different protein complexes which may play distinct roles during erythropoiesis. GATA-1 gene itself is one of the candidates for GATA-1 target genes, while molecular basis of a putative GATA-1 complex regulating GATA-1 expression is not well understood.
In this study, we used the zebrafish system to elucidate regulatory mechanisms of the GATA-1 gene. To this end, we isolated and analyzed zebrafish GATA-1 genomic DNA, resulting in a finding of a novel intron that was unknown in the previous analysis. This intron corresponds to the first intron of other vertebrate GATA-1 genes. GFP reporter analyses revealed that this intron and a distal double GATA motif in the regulatory region are important for the regulation of zebrafish GATA-1 gene expression. To examine whether GATA-1 regulates its own gene expression, we microinjected into embryos a GFP reporter gene linked to the GATA-1 gene regulatory region and the GATA-1 mRNA successively.
Surprisingly, ectopic expression of the reporter gene was induced at the site of GATA-1 overexpression and was dependent on the distal dotuble GATA motif. Functional domain analyses using transgenic fish lines harboring the GATA-1-GFP reporter construct revealed that both the amino- and carboxyl-zinc finger domains of GATA-1, hence the intact GATA-1 function, are required for the ectopic GFP expression. These results provide the first in vivo evidence that GATA-1 gene expression undergoes positive autoregulation.
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