| 研究生: |
鄒毅寰 Tsou, Yi-Huan |
|---|---|
| 論文名稱: |
探討核仁蛋白 MSP58 調控核醣體 DNA 基因轉錄之角色 To study the role of nucleolar protein MSP58 in ribosomal DNA (rDNA) transcription |
| 指導教授: |
林鼎晏
Lin, Ding-Yen |
| 學位類別: |
碩士 Master |
| 系所名稱: |
醫學院 - 藥理學研究所 Department of Pharmacology |
| 論文出版年: | 2012 |
| 畢業學年度: | 100 |
| 語文別: | 中文 |
| 論文頁數: | 74 |
| 中文關鍵詞: | MSP58 、核仁 、核醣體 DNA 基因轉錄作用 |
| 外文關鍵詞: | MSP58, nucleoli, ribosomal DNA gene transcription |
| 相關次數: | 點閱:104 下載:2 |
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MSP58/MCRS1 過去報導與許多蛋白質有交互作用關係,如核仁蛋白 p120、Mi-2β及轉錄因子 Daxx、STRA13、Nrf1、SWI/SNF 染色質重組複合體次單元 BRG1 或是 RNA 結合蛋白 FMR等,這些發現說明了 MSP58 在細胞核以及核仁內為重要的轉錄調節者。在本研究我們探討 MSP58 調控核醣體 DNA (rDNA) 基因轉錄所扮演的角色,利用本實驗室設計與製作的 MSP58 多株抗體進行免疫螢光染色,我們發現 MSP58 呈現微小球顆粒狀分布在核仁內,細胞質內的分佈較少,並且 MSP58 會與核仁轉錄因子 UBF 結合且共位。透過報告基因分析法,我們發現細胞過度表現 MSP58 會抑制 rDNA 啟動子的轉錄活性,然而利用 siRNA 抑制內生性 MSP58 的表現則會提升 rDNA 轉錄活性。此外,以酵母菌雙雜交技術我們發現一些 MSP58 的新穎結合蛋白。由以上發現指出 MSP58 蛋白質複合體於細胞內參與核醣體 DNA 基因的轉錄調控。
MSP58/MCRS1 has been reported to interact with several proteins, such as nucleolar protein p120, Mi-2β; transcription factors Daxx, STRA13, Nrf1, SWI/SNF chromatin remodeling subunit BRG1, and also RNA-binding protein FMR. These findings imply that MSP58 proteins are crucial for transcriptional regulation in nuclei and nucleoli. In this study, we examined the role of MSP58 in the transcriptional modulation of the ribosomal genes. Initially, we generated and characterized the polyclonal antibodies against MSP58. The main localization of MSP58 was consistent in nuclei and microspherule speckles of nucleoli with faint cytoplasmic staining. Interestingly, MSP58 was associated and co-localized with endogenous nucleolar transcription factor UBF in the nucleoli. Analysis by luciferase reporter assay revealed that ectopic expression of MSP58 suppressed transcriptional activity of the rDNA promoter, whereas knocking-down MSP58 expression by siRNA treatment up-regulated ribosomal RNA transcription. In addition, we identified several nucleolar proteins as novel MSP58-interacting proteins by yeast two-hybrid screening. These results indicated that MSP58 complexes are involved in the regulation of ribosomal gene transcription in vivo.
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