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標題: | 自噬作用對TRIM5α調控EB病毒外鞘蛋白質BORF1的影響 Role of autophagy in the regulation of BORF1 of Epstein-Barr virus by TRIM5α |
作者: | Tse-Yao Chen 陳則堯 |
指導教授: | 張麗冠 |
關鍵字: | Epstein-Barr Virus (EB 病毒),外鞘蛋白質,BORF1,TRIM5α,泛素化修飾,p62,自噬作用, Epstein-Barr Virus (EBV),capsid proteins,BORF1,TRIM5α,ubiquitination,p62,autophagy, |
出版年 : | 2016 |
學位: | 碩士 |
摘要: | EB病毒 (Epstein-Barr Virus),屬於皰疹病毒科γ亞科,其生活史包括潛伏期 (latency) 與溶裂期 (lytic cycle)。病毒在進入溶裂期後會大量表現病毒基因,複製遺傳物質並組裝成為病毒顆粒,此過程中外鞘殼體 (capsid) 的組裝是非常重要的步驟。BORF1是EB病毒的次要外鞘蛋白質 (minor capsid protein),能與BDLF1形成三聚體 (triplex) 並引導其餘的外鞘蛋白質進入細胞核內進行組裝。Tripartite-motif 5 alpha (TRIM5α) 是反轉錄病毒的限制因子 (restriction factor),本實驗室先前研究發現TRIM5α能與BORF1直接結合,並利用其E3連接酶 (E3 ligase) 活性催化BORF1的泛素化修飾使其穩定性下降,進而抑制病毒溶裂期的發展。本研究首先證明TRIM5α B30.2功能區上的surface patch區域對其與EB病毒外鞘蛋白質BORF1的結合相當重要。接著觀察BORF1泛素化修飾的情形,顯示lysine 114為其上重要的泛素化修飾位置,若突變會提昇BORF1蛋白質的穩定性。TRIM5α亦能催化BORF1的K63鍵結長鏈泛素化修飾,顯示其降解可能與選擇性自噬作用有關。本研究進一步發現選擇性自噬作用的受體p62/sequestosome 1 (SQSTM1) 能透過辨認BORF1上的泛素化修飾而與其結合。此外,若抑制溶酶體 (lysosome) 活性則會使受泛素化修飾的BORF1累積,並提昇與p62的結合量。最後,若以siRNA抑制細胞中p62的含量,則BORF1的表現量會上升。綜合以上結果,本研究發現TRIM5α能與p62共同作用,透過選擇性自噬作用降解BORF1。 Epstein-Barr virus (EBV) is a human herpesvirus with two distinct life cycles, latency and lytic cycle. During the lytic cycle, it encodes proteins that are involved in viral lytic DNA replication, capsid assembly and maturation. BORF1, a minor capsid protein of EBV, forms a triplex with BDLF1 and recruits other capsids into the nucleus for capsid assembly. Tripartite-motif 5 alpha (TRIM5α) is a restriction factor with the activity of ubiquitin E3 ligase. Previous study indicated that TRIM5α binds to BORF1 and enhances its ubiquitination, thus inhibiting lytic progression. This study demonstrated the surface patch region on B30.2 in TRIM5α is responsible for the interaction with BORF1. Moreover, lysine 114 of BORF1 is the major ubiquitination site. Mutating Lys-114 stabilizes BORF1. TRIM5α also promotes K63-linked poly ubiquitination of BORF1, which implies the possibility of involvement of autophagy in this process. This study reveals that the cargo receptor of selective autophagy, p62/sequestosome 1 (SQSTM1), recognizes the K63-linked poly ubiquitin chain on BORF1 and target it for autophagic degradation. Inhibition of lysosome activity causes the accumulation of ubiquitinated BORF1, and the interaction between p62 and BORF1 is evident. Finally, knockdown of p62 enhances the expression of BORF1. Taken together, this study demonstrates that TRIM5α functions concordantly with p62 and destabilizes BORF1 via autophagic degradation. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/78170 |
DOI: | 10.6342/NTU201601599 |
全文授權: | 有償授權 |
顯示於系所單位: | 生化科技學系 |
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