| Project/Area Number |
09470519
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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 |
Human genetics
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| Research Institution | Tokyo Medical and Dental Univerisry |
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Principal Investigator |
SHIGETAKA Kitajima Medical Research Institute, Tokyo Medical and Dental Univerisry Professor, 難治疾患研究所, 教授 (30186241)
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| Co-Investigator(Kenkyū-buntansha) |
ATORU Oshiro Medical Research Institue, Tokyo Medical and Dental Univerisry Assistant Professor, 難治疾患研究所, 助手 (30160485)
TERUMASA Tuchiya Medical Research Insitute, Tokyo Medical and Dental Univerisry Assistant Professor, 難治疾患研究所, 助手 (20242109)
YUKIO Yasukochi Medical Research Institue, Tokyo Medical and Dental Univerisry Professor, 難治疾患研究所, 教授 (60037398)
TEIJIRO Aso Cancer Research Institue, Investigator, 癌研究会研究所, 研究員 (20291289)
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| Project Period (FY) |
1997 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥13,300,000 (Direct Cost: ¥13,300,000)
Fiscal Year 2000: ¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1999: ¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1998: ¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 1997: ¥5,500,000 (Direct Cost: ¥5,500,000)
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| Keywords | TFIIF / Tripartite structure of RAP74 / HIV-Tat / Transcriptional elongation / Transcriptional recycling / Elongin / Testis-specific A2 / Gene targeting of elongin / RNAポリメラーゼII / 組織特異的因子 / 開始型 / 伸長型 / トリプシン分解 / 組織特異的エロンガン / 転写制御 / p32 / p30 / RNAポリメラーゼIIホロ酵素 |
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| Research Abstract |
TFIIF, which recruits RNA polymerase II (Pol II) into preinitiation complex formed at gene promoter, can also function as an elongation factor for mRNA synthesis by Pol II, Elongin is a general elongation factor which may be involved in oncogenesis in VHL disease. To understand regu1atory mechanism of transcriptional elonagtion, we investigated function of TFIIF and elongin in this study.
1) TFIIF
RAP74 subunit of TFIIF is predicted to have tri-partite structure ; N- and C-terminal domains, and central region. We identified autoantibodies against RAP74 subunit of TFIIF in sera from autoimmune disease patients which selectively recognize the central portion of RAP74. By 2-hybrid screen, p32 was cloned as RAP74-interacting gene. p32 could bind to central region of RAP74 and HIV-Tat in vitro, and was capable for accelerating elongation by Pol II in the presence of HIV-Tat. p32 may be implicated in replicative growth of HIV.We could not clone FCP1, which binds to C-terminal of RAP74 and dephosphorylates CTD of Pol II by 2-hybrid screening. FCP1 was reported by Greenblatt et al. We expressed and purified recombinant TFIIF through co-infection of baculoviruses for RAP30 and 74. Using this, structural analysis of free, initiation, and elongation forms of TFIIF was performed. Tripartite structure of RAP74, its outer localization in TFIIF, and inner location of RAP30 was revealed. We are trying to crystallize TFIIF molecule for structural analysis.
2) Elongin
We cloned a novel form of elongin, A2, from EST on database. A2 is expressed preferentially in the testis while A is ubiquitously expressed. Recombinant A2 is capable for stimulating elongation of Pol II in vitro. A2 may function in expression of testis-specific genes. Furthermore, we identified A3, another novel form of elongin family. We are now characterizing its strucutre and function. Our gene knock-out project is under investigation, We screened ES cells which are heterologous for elongin A gene.
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