| Project/Area Number |
16370093
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Cell biology
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| Research Institution | Kyoto University (2005)
Biomolecular Engineering Research Institute (BERI) (2004) |
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Principal Investigator |
OKAZAKI Kenji Kyoto University, Graduate School of Science, Research Scientist, 大学院・理学研究科, 研究員(科学研究) (50211115)
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| Co-Investigator(Kenkyū-buntansha) |
AOTA Shinichi Osaka University, Graduate School of Frontier Biosciences, Research Scholar, 生命機能研究科, 特任研究員 (50192456)
坂本 るり子 技術研究組合生物分子工学研究所, 分子機能研究部, 研究員
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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 |
¥15,200,000 (Direct Cost: ¥15,200,000)
Fiscal Year 2005: ¥7,300,000 (Direct Cost: ¥7,300,000)
Fiscal Year 2004: ¥7,900,000 (Direct Cost: ¥7,900,000)
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| Keywords | Cell differentiation / Pax6 / Regulation of activity / Paired-domain / LE9 / Phosphorylation of transcription factors / 蛋白質リン酸化 / 細胞生物学 / 遺伝子発現 / プラコード / 転写因子 / LE9 |
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| Research Abstract |
By using LE9 enhancer element, a target DNA fragment directly bound by Pax6 protein, that is present in the head surface ectoderm-specific enhancer region of the pax6 gene and is responsive to the autoreguration of the pax6 gene expression, we studied regulatory mechanisms of Pax6 protein through its phosphorylation. Our result showed that the activity of Pax6 toward LE9 is highly elevated by ERK_<1/2> or p38MAPK. We identified four phosphorylation sites in the carboxy terminal domain, among which the three sites were able to be directly phosphorylated by p38MAPK in vitro. Furthermore, a Ser-to-Ala mutation of one of these residue resulted in a suppression of the transcriptional response to the activation of the kinase pathways. Because the phosphorylation of Pax6 had little influence to the stability of Pax6 protein, the phosphorylated sites seems to operate by interacting with other factors to contribute to the transcriptional activation of LE9. Analyses of the phosphorylation pattern of in vivo Pax6 became possible by preparation of monoclonal antibodies specific for an amino acid sequence containing the phosphorylated residue. Whereas a growth signal-induced increase of the phosphorylation of nuclear Pax6 supported the significance of ERK pathway, its sustained phosphorylation suggested an involvement of another kinase. Above results demonstrated that the function of Pax6 is regulated by the phosphorylation of its C-terminal domain and that this control is dependent on the MAP kinase pathways. It Will be an important issue to clarify the mechanisms for phosphorylation signaling in the differentiation process of eyes, in which LE9 enhancer is considered to really function.
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