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研究生:許妤安
研究生(外文):hsü yü an
論文名稱:第一型干擾素生物功能之研究:IFIT-1生理功能及新的訊息傳導路徑之研究
論文名稱(外文):The biological roles of Type I interferon: biological functions of IFIT-1 and a novel signaling pathway
指導教授:萬磊蔡輔仁蔡輔仁引用關係
指導教授(外文):wan leits''ai fu jên
學位類別:碩士
校院名稱:亞洲大學
系所名稱:生物科技學系碩士班
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:2007
畢業學年度:95
語文別:中文
論文頁數:79
中文關鍵詞:生理功能
外文關鍵詞:biological functions
相關次數:
  • 被引用被引用:0
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  • 下載下載:28
  • 收藏至我的研究室書目清單書目收藏:1
干擾素,是一種能夠在多種細胞型態上進行活化調節的細胞激素,通常使用於癌症與感染性疾病的治療。在受到病毒的感染,干擾素刺激或雙股RNA 的誘導,會使得細胞內名為Viral-Stress Inducible Gene(VSIG)這類型的基因表現。其中之一的p56 蛋白便是由Interferon Stimulate Gene 56(ISG56)基因(又稱為interferon-induced protein with tetratricopeptide repeats 1; IFIT-1)轉錄成mRNA 561 基因後再轉譯而成的蛋白質。它在序列結構上具有6 個tetratricopeptide repeats (TPR),通常是透過這些TPR 而與細胞內的其他蛋白質產生交互作用。在先前的研究中發現,p56 能夠透過eIF3 上的次單位p48 與eIF3 結合,抑制了三元複合體(ternary complex)形成,因而抑制下游蛋白質合成。但在我們的研究中,利用噬菌體表現系統發現另一個會與p56 產生交互作用的蛋白:RPL15,並以沈澱試驗的方法證明之。此外我們還經由MTT assay 得知,當細胞在RPL15 被抑制或者p56 表現量增加的情況下,都會造成細胞的生長較為緩慢以及對於干擾素的刺激變得較為敏感。在另一方面,前人的研究指出,細胞受到干擾素α 刺激活化了STAT6 是只有在B 細胞上才有的專一性反應,在我們的實驗中首度發現肝癌細胞能透過第一型的干擾素刺激,在活化STAT2 後而活化STAT6,並形成STAT6:STAT2 異質複合體進入核內,使IL-1Ra 基因表現。
Interferon-(IFN), a cytokine with modulatory activities on many cell types, is useful for treating many types of cancer and infectious diseases. The common set of genes induced by virus infection, IFN, and dsRNA is called viral stress-inducible genes (VSIG). One of VSIG, P56 is the product of mRNA 561 encode by the ISG56 gene (also known as IFIT1). It contains six tetratricopeptide repeats (TPRs) spaced evenly along the entire protein, which are known to mediated protein-protein interactions. Previous studies we known that P56 can bind to the large eIF-3 complex that contains P48 subunit and to inhibit protein synthesis, which blocks the formation of stable eIF3‧ternary complex. With six TPRs, P56 may interact with various proteins and influence some other biological process. In our study, we found the ribosomal protein 15 (RPL15) interacted with p56 by phage display experiments in vitro. The p56-RPL15 interaction was confirmed by pull-down assay in vivo. Furthermore, we found to down-regulation of RPL15 expression or overexpression of p56 was inhibited cell growth and more sensitivity with IFN by MTT assay. Previous study indicated that induction of STAT6 complexes by IFNα appears to be cell type specific. STAT6 in response to IFNα is predominantly detected in B cells. It was first proved in our research that type I IFN can activate STAT6 in hepatoma cells and lead to the formation of stat2 : stat6 complexes. The STAT6 activation is mediated by the associating with activated STAT2 and it will result in the production of IL1-Ra.
目錄-1 ~ 2
中文摘要-3
英文摘要-4
第一章 前言-5 ~ 15
1. 干擾素的簡介
2. 第一型干擾素的臨床應用
3. JAK-STAT signaling pathway
4. 第一型干擾素的訊息傳遞路徑
5. IFIT1 簡介
6. RPL15 簡介
7. 研究主題
第二章 材料與實驗方法-16 ~ 23
1. 噬菌體篩選(phage display)
2. 沈澱試驗(GST pull-down assay)
3.西方墨點法(Western Blot)
4.建構質體DNA
5.細胞培養
6.Transfection
7.MTT assay
8.定量PCR 分析
9.免疫沈澱法
10.基因敲除技術
11.ELISA (Enzyme-link immunosorbent assay)
12.EMSA (electrophoretic mobility shift assay)
第三章 結果-24 ~ 29
第四章 討論-30 ~ 33
參考文獻-34 ~ 48
圖表-49 ~ 76
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