2001 Fiscal Year Final Research Report Summary
Biochemical analysis of Wnt/Wingless signal transduction pathway with tissue culture system
| Project/Area Number |
12680635
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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 |
Functional biochemistry
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
YANAGAWA Shin-ichi Inst. for Virus Dept. of Research Viral Oncology, Kyoto University, Instructor, ウイルス研究所, 助手 (70183978)
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| Project Period (FY) |
2000 – 2001
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| Keywords | Wnt / Development / Casein kinase I / Signal transduction / β-catenin / Drosophila |
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| Research Abstract |
The Wnt family of secretory protein plays pivotal roles in a number of basic developmental processes. Components of the Wnt/Wingless (Wg) signaling pathway are Frizzled, Dishevelled, Glycogen synthase kinase-38, Axin, APC tumor suppressor, β-catenin, and T-cell transcription factor. These genes conserved in vertebrates and invertebrates. Using cell culture assay for Wnt/Wg that we have established, we are analyzing biochemical interactions among these components of the Wnt/Wg pathway. Casein kinase I (CKI) was recently reported as a positive regulator of Wnt signaling in vertebrates and C. elegans (Peters, J. M. et al : Nature 1999 401 : 345-350) To elucidate the function of Drosophila CKI in the wingless (wg) pathway, we have disrupted its function by double-stranded RNA-mediated interference (RNAi). While previous studies mainly based on CKI overexpression, this is the first convincing loss-of-function approach of CKI. Surprisingly, CKI_α- or CKI_ε-RNAi markedly elevated the Armadillo (Arm) protein levels in Drosophila Schneider S2R+ cells, without affecting its mRNA levels. Pulse-chase analysis showed that CKI-RNAi indeed stabilizes Arm protein. Moreover Drosophila embryos injected with CKI_α-double-stranded RNA showed a naked cuticle phenotype, which is associated with activation of Wg signaling. These results indicate that CKI functions as a negative regulator of Wg/Arm signaling. Overexpression of CKI_α induced hyper-phosphorylation of both Arm and Dishevelled in S2R+ cells and conversely, CKI_α-RNAi reduced the amount of hyper-modified forms. His-tagged Arm was phosphorylated by CKI_α in vitro on a set of serine and threonine residues that are also phosphorylated by Zeste-white 3. Thus, we propose that CKI phosphorylates Arm and stimulates its degradation.
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Research Products
(8 results)
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[Publications] Shimada, Y., Usui, T., Yanagawa, S.-i., Takeichi, M., and Uemura, T.: "Asymmetric colocalization of Flamingo, a seven-pass transmembrane cadherin, and Dishevelled in planar cell polarization"Current Biology. 11. 859-863 (2001)
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[Publications] Chen, W., Hu, L.A., Semenov, M.V., Yanagawa. S.-i., Kikuchi, A., Lefkowitz. R.J., and Miller, W.E.: "β-arrestin1 modulates lymphoid enhancer factor transcriptional activity through interaction with phosphorylated dishevelled proteins"Proc. Natl. Acad. Sci. USA. 98. 14889-14894 (2001)
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「研究成果報告書概要(欧文)」より
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[Publications] Yanagawa, S.-i., Matsuda. Y., Lee, J.-S., Matsubayashi. H., Sese, S., Kadowaki, T., and Ishimoto, A.: "Casein kinase I phosphoryltaes the Armadillo protein and induces its degradation in Drosophila"EMBO. J.. 21 (in press). (2002)
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「研究成果報告書概要(欧文)」より
