| Project/Area Number |
16570185
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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 |
Developmental biology
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| Research Institution | Osaka University (2005)
Biomolecular Engineering Research Institute (BERI) (2004) |
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Principal Investigator |
AOTA Shinichi Osaka University, Graduate School of Frontier Biosciences, researcher, 大学院・生命機能研究科, 特任研究員 (50192456)
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| Co-Investigator(Kenkyū-buntansha) |
OKAZAKI Kenji Kyoto University, Faculty of Science, researcher, 理学部, 研究員 (50211115)
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| Project Period (FY) |
2004 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 2005: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 2004: ¥1,600,000 (Direct Cost: ¥1,600,000)
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| Keywords | regulation of expression / transcription factor / regulatory network / feedback regulation / placode / Pax6 / differentiation |
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| Research Abstract |
To elucidate the genetic network controlling the lens placode development, we first optimized a high-speed enhancer assay utilizing the chicken embryo electroporation technology. We developed a reporter plasmid ptk2GFP, which resulted in the lowest background activity and good responses to various enhancers. By cloning various DNA fragments from the control region of the Pax6 gene, which plays central roles in lens development, we screened Pax6 enhancers active in the head surface ectoderm and the lens placode. In addition to the already-known enhancer Pax6EE, in which we previously identified PAX6 and SOX2 binding sites, we identified a novel enhancer termed Pax6LS in the downstream region of the Pax6 locus. We extensively analyzed Pax6EE and Pax6LS by site-directed mutagenesis and chicken embryo enhancer assays, and found evidences that PAX6 activates Pax6EE in vivo by synergistic interactions with many transcription factors. We also identified important sites for Pax6LS enhancer activity by site-directed mutagenesis. Pax6LS is active in the lens placode and lens vesicle. Since the lens placode is not formed in Sey homozygous mutant mice, of which Pax6 alleles are mutated, Pax6LS enhancer activity is dependent on Pax6. We also found that Pax6LS is downstream of BMP signaling, which has been implicated for roles in lens placode development. These results altogether indicated the presence of multiple feedback loops regulating the Pax6 expression during lens placode development.
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