2001 Fiscal Year Final Research Report Summary
STUDY FOR MOLECULAR MECHANISM OF MUSCLE-SPECIFIC ALTERNATIVE RNA SPLICING.
| Project/Area Number |
11670132
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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 |
General medical chemistry
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| Research Institution | JICHI MEDICAL SCHOOL |
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Principal Investigator |
ENDO Hitoshi JlCHI MEDICAL SCHOOL SCHOOL OF MEDICINE ASSISTANT PROFESSOR, 医学部, 講師 (50221817)
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| Project Period (FY) |
1999 – 2001
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| Keywords | MYOGENESIS / ATP SYNTHASE / RNA SPLICING / ALTERNATIVE SPLICING / MITOCHONDRIA / SPLICING SILENCER / SPLICING FACTOR / SPLICING REGULATION |
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| Research Abstract |
Mitochondrial ATP synthase provides most ATP in eukaryotic cells. The muscle-specific isoform of ATP synthase γ-subunit (F_1γ) is generated by tissue-specific alternative splicing. In this study, we investigated on the molecular mechanism of muscle-specific alternative splicing in F_1γ pre-mRNA. First, we developed the reversible induction system for muscle-specific alternative splicing using human fibrosarcoma (HT1080) cells, and also constructed an in vitro splicing system reflecting muscle-specific alternative splicing in human F_1γ minigene, by addition with nuclear extracts from muscle-specific splicing-induced HT1080 cells. From mutation analysis of F_1γ minigene using this in vitro system, we identified an Exonic Splicing Enhancer (ESE) and a Muscle-specific Exonic Splicing silencer (MS-ESS) on an alternatively spliced exon. The ESE is required for constitutive exon selection, and the MS-ESS specifically acts for exon exclusion under the muscle-specific splicing condition. From UV cross-linking and North-Western blot analyses using these cis-regulatory elements, a 48 kDa protein and a 42 kDa protein were detected as RNA-binding proteins, which bind to ESE and ME-ESS, respectively. Finally, we also described the difference of muscle-specific splicing reguratory mechanism between muotubes and mature muscle fibers using transgenic mice bearing F_1γ minigene. These splicing reguratory factors must be identified to understand the molecular mechanism of muscle-specific alternative splicing.
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[Publications] Otsuki T., Furukawa Y., Ikeda K., Endo H., Yamashita T., Shinohara A., Iwamatsu A., Ozawa K., Liu J. M.: "Fanconi anemia protein, FANCA, associates with BRG1, a component of the human SWI/SNF complex"Hum. Mol. Genet.. 10. 2651-60 (2001)
Description
「研究成果報告書概要(欧文)」より
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