Role of protein phosphorylation in Myulating RNA polymerase II transcription

蛋白质磷酸化在 Myulate RNA 聚合酶 II 转录中的作用

• 批准号: 11680685
• 负责人: HISATAKE Koji
• 金额: $ 1.02万
• 依托单位: Saitama Medical School
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11680685/
• 关键词: General Transcription Factor; Baculo virus; Protein Phosphorylation; Transcription; TFIIH; Capping Reaction; バキュウロウイル; リン酸化; 伸長反応; CTD; バキュロウイルス

1. Role of ERCC3 in transcriptional activation by GAL4-VP16In basal transcription, ERCC3 helicase acts during promoter opening and promoter escape. We tested at which step ERCC3 helicase acts during transcriptional activation by GAL4-VP16. The effect of TFIIH mutants were first tested using premelted templates, and revealed that TFIIH functions during promoter escape in activated transcription. The effect is specific to activation. Using the various mutant templates, we found that activators act during a very early stage of transcription. The effect requires coactivator PC4. RNA polymerase II is unstable during the early phase of transcription and may abort transcription. With the use of ATP analogs, we found that RNA polymerase II may detach from the template when ERCC3 helicase activity is blocked. Taken together, transcriptional activation involves the enhancement of recruitment of transcriptional machinery and promoter escape. Rate constant of promtoer escape is not stimulated in activated transcription.2. Role of TFIIH in transcription-coupled capping reactionsIn collaborated with Dr.Mizumoto, we have developed a system in which transcription and capping reactions can occur together. The system includes recombinant factors except RNA polymerase II.In this system, capping reaction occurs simultaneously with transcription initiation. The molecular analyses of capping and its relation to transcription initiation are in progress.

1. ERCC 3在GAL 4-VP 16转录激活中的作用在基础转录中，ERCC 3解旋酶在启动子打开和启动子逃逸过程中起作用。我们测试了ERCC 3解旋酶在GAL 4-VP 16的转录激活过程中作用的步骤。TFIIH突变体的作用首先使用预熔化的模板进行测试，并揭示TFIIH在激活转录的启动子逃逸期间起作用。这种效应是激活所特有的。使用各种突变体模板，我们发现激活因子在转录的非常早期阶段起作用。该效应需要辅活化剂PC 4。RNA聚合酶II在转录的早期阶段是不稳定的，并且可以中止转录。通过使用ATP类似物，我们发现当ERCC 3解旋酶活性被阻断时，RNA聚合酶II可以从模板上分离。两者合计，转录激活涉及增强转录机器的募集和启动子逃逸。启动子逃逸的速率常数在激活转录中不受刺激. TFIIH在转录偶联加帽反应中的作用在与Mizumoto博士的合作中，我们开发了一个转录和加帽反应可以同时发生的系统。该系统包括除RNA聚合酶II外的重组因子。在该系统中，加帽反应与转录起始同时发生。加帽及其与转录起始的关系的分子分析正在进行中。

项目成果

Ishiguro,A.: "The Rpb6 subunit of fission yeast RNA polymerase II is a contact target of the transcription elongation factor TFIIS"Molecular and Cellular Biology. 20. 1263-70 (2000)

Ishiguro,A.：“裂殖酵母 RNA 聚合酶 II 的 Rpb6 亚基是转录延伸因子 TFIIS 的接触靶标”《分子与细胞生物学》。

Ishiguro,A.: "The Rpb 6 subunit of fission yeast RNA polymerase II is a contact target of the transcription elongation factor TFIIS"Molecular and Cullular Biology. 20. 1263-70 (2000)

Ishiguro,A.：“裂殖酵母 RNA 聚合酶 II 的 Rpb 6 亚基是转录延伸因子 TFIIS 的接触靶标”《分子与细胞生物学》。

Ishiguro, A: "The Rpb 6 subunit of fission yeart RNA polymerase II is a contant targect of the transcription elongation factor TFIIS"Molecular and Cellular Biology. 20. 1263-70 (2000)

Ishiguro, A：“裂变 Yeart RNA 聚合酶 II 的 Rpb 6 亚基是转录延伸因子 TFIIS 的恒定靶标”《分子和细胞生物学》。

Wada,T.: "FACT relieves DSIF/NELF-mediated inhibition of transcriptional elongation and reveals functional differences between P-TEFb and TFIIH."Molecular Cell. 5. 1067-1072 (2000)

Wada,T.：“FACT 缓解 DSIF/NELF 介导的转录延伸抑制，并揭示 P-TEFb 和 TFIIH 之间的功能差异。”《分子细胞》。

Wada, T: "The Fact uliums DSIF/NELF mediated inhibition of transcriptional elongation and Miuals functional differences between p-TEF6 and TFIIH"Molecular Cell. 5. 1067-1072 (2000)

Wada, T：“事实 uliums DSIF/NELF 介导的转录延伸抑制以及 p-TEF6 和 TFIIH 之间的 Miuals 功能差异”分子细胞。