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Title: | MCAF1參與Epstein-Barr病毒溶裂期之複製 Involvement of MCAF1 in Epstein-Barr Virus Lytic Replication |
Authors: | Yu-Chun Lee 李宇群 |
Advisor: | 張麗冠(Li-Kwan Chang) |
Keyword: | 病毒,溶裂期複製, MCAF1,EBV,lytic replication, |
Publication Year : | 2012 |
Degree: | 碩士 |
Abstract: | Epstein-Barr病毒 (EBV)屬於人類皰疹病毒科的一種,會感染B細胞或是上皮細胞,其生活史包含了潛伏期與溶裂期。當病毒在宿主細胞內受到刺激便會由潛伏期進入溶裂期,並在約24小時後開始大量複製以產生病毒顆粒,在其溶裂期的生活史中,極早期基因的產物Rta及Zta能夠促使早期基因的活化,而這類早期基因主要功能皆與病毒DNA的複製相關,此外,Rta、Zta亦能夠在Zta Response Element (ZRE)與宿主蛋白質MCAF1形成複合體,幫助病毒協同活化早期基因。本研究的目的是想要了解MCAF1,是否會參與EB病毒的溶裂複製。首先以免疫螢光染色的結果發現,當細胞在加藥誘導進入溶裂期後的12小時開始,MCAF1便與Rta或Zta位於細胞核內相同的位置,而這樣的現象直到加藥後的60小時依然可見;除此之外,在加藥後36小時觀察到Rta與MCAF1和病毒早期蛋白質BBLF2/3、BALF2和BBLF4位於細胞核內相同位置。另外,經由共免疫沉澱分析顯示,MCAF1會與病毒早期基因BALF2和BBLF2/3產生交互作用。最後,利用siRNA將細胞內的MCAF1抑制之後發現,雖然EB病毒早期基因EA-D的表現未受影響,但病毒複製的能力仍因MCAF1表現量的降低而有所下降。綜合上述結果,MCAF1除了能夠在病毒溶裂期的極早期幫助Rta、Zta活化早期基因外,亦藉由與早期蛋白質的交互作用參與EB病毒DNA的複製。 Epstein-Barr virus (EBV) is a human gammaherpesvirus which infects B lymphocytes and epthilial cells with two distinct life cycle, latency and lytic cycle. When a host cell is suffered from stresses, EBV will reactivate from latency and amplify its genome via lytic replication rapidly in 24 hrs. During the lytic cycle, EBV espresses two immediate early proteins, Rta and Zta, which play a key role in the regulation of early genes that are associated with viral DNA replication. In addition, Rta and Zta form a complex via MCAF1 on the Zta response element (ZRE) to accelerate activation of early genes synergistically. This study is to demonstrate whether MCAF1 involves in EBV lytic replication. Firstly, immunofluorescence analysis showed that MCAF1 colocalizes with Rta and Zta after lytic induction for 12 hr - 60 hr. Moreover, Rta and MCAF1 also colocalize with BBLF2/3, BALF2 and BBLF4 at 36 hr after lytic induction. Additionally, MCAF1 interactes with BALF2 and BBLF2/3 by coimmunoprecipitation assay. Furthermore, RNAi assay demonstrated that knock down of MCAF1 decreases EBV lytic replication, although the expression of EA-D remains unchanged. Hence, MCAF1 not only participates in the activation of early gene with Rta and Zta, but also involves in EBV lytic replication by interacting with replication proteins. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/64747 |
Fulltext Rights: | 有償授權 |
Appears in Collections: | 生化科技學系 |
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ntu-101-1.pdf Restricted Access | 8.37 MB | Adobe PDF |
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