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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 張麗冠 | zh_TW |
| dc.contributor.advisor | Li-Kwan Chang | en |
| dc.contributor.author | 胡家瑜 | zh_TW |
| dc.contributor.author | Jia-Yu Hu | en |
| dc.date.accessioned | 2024-08-15T16:11:22Z | - |
| dc.date.available | 2024-08-16 | - |
| dc.date.copyright | 2024-08-15 | - |
| dc.date.issued | 2024 | - |
| dc.date.submitted | 2024-08-06 | - |
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Regulation of the stability of Epstein-Barr virus BFRF3 by TRIM5α-mediated TRIMosome. 臺灣大學生化科技學系碩士論文 蔡曉涵. (2016). Role of Rta of Epstein-Barr virus in mitochondria. 臺灣大學生化科技學系碩士論文. | - |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/94200 | - |
| dc.description.abstract | EB 病毒 (Epstein-Barr virus, EBV)，感染全球約90%以上的人口，與許多腫瘤及自體免疫疾病相關。Rta 是 EB 病毒在溶裂期的極早期表現的蛋白質，具有轉錄活化的能力，調控許多病毒與宿主的基因表現。先前我們發現反轉錄病毒的限制因子 TRIM5α 會與 Rta 結合並促進其泛素化，因此抑制EB 的病毒顆粒產生。近期研究顯示 TRIM5α 會參與粒線體自噬 (Mitophagy)，以維持粒線體品質。而我們發現 Rta 也位在粒線體，因此本研究將探討 Rta 與TRIM5α 如何調控粒線體自噬。首先以粒線體分離和免疫螢光染色證明 Rta 位於粒線體外膜，並與 TOM20 及 TRIM5α 共定位在粒線體上。Rta 也會讓粒線體中磷酸化的 Drp1 增加，同時 LC3-I 轉換成 LC3-II 的量也會增加。接著以 shRNA 抑制 Drp1 表現後，LC3-II 的量下降，顯示 Rta 透過 Drp1 誘導粒線體自噬。分析 Rta 的粒線體導向序列 (Mitochondrial targeting sequences, MTS) 後，將 Rta的 N 端氨基酸突變，以免疫螢光染色分析，結果顯示Rta(MTS)突變株在粒線體的分布下降且 LC3B 與 TOM20 的共定位也會降低。另外，透過shRNA抑制TRIM5α表現，也觀察到在粒線體的Rta所誘導的LC3-II 的量會減少。最後，若是將HEK293(2089)細胞中Drp1的表現量抑制，EB 病毒的早期蛋白質 EA-D 的表現會減少，表示溶裂期受到抑制。綜合上述結果，本研究證實 Rta 是一個位在粒線體的病毒蛋白質，會經由 Drp1 誘發粒線體自噬，同時發現TRIM5α會參與Rta誘導的粒線體自噬，且EB 病毒會透過調控粒線體動態平衡來維持溶裂期。未來將進一步闡述EB 病毒的Rta和TRIM5α如何調節粒線體動態平衡的機制。 | zh_TW |
| dc.description.abstract | Epstein-Barr virus (EBV) infects over 90% population and is associated with many tumor and autoimmune diseases. Rta encoded by EBV is an immediate-early transcription factor, which activates the transcription of EBV lytic genes as well as cellular genes. Our earlier study found that a retrovirus restriction factor, TRIM5α interacts with Rta and promotes its ubiquitination, thus inhibiting EBV virions production. Recent studies indicated that TRIM5α regulates mitochondria quality control via mitophagy. We also found that Rta is localized at mitochondria. Therefore, this study aims to explore how Rta and TRIM5α regulate mitophagy. This study found by indirect immunofluorescence analysis that Rta colocalizes with TOM20 and TRIM5α at the outer membrane of mitochondria and enriches the levels of phosphorylated Drp1(S616) and accumulates the levels of LC3-II on mitochondria. Knockdown of the expression of Drp1 also decreases the levels of LC3-II. By analyzing the sequence of the possible mitochondrial targeting sequence (MTS) in Rta, we generated an Rta mutant, Rta(MTS), with mutated MTS at the N-terminal amino acid residues. Confocal microscopy analysis revealed that the mutation descreases the colocalization of Rta and TOM20; the colocalization of LC3B and TOM20 in Rta(MTS) is also reduced. Additionally, knockdown TRIM5α also decreases the amounts of LC3-II induced by Rta in mitochondria. Finally, the inhibition of Drp1 expression in HEK293(2089) cells reduces the levels of EA-D, indicating that the lytic phase is suppressed. Taken together, this study demonstrates that Rta is a viral protein localized in mitochondria, which induces mitophagy via Drp1 and thus influences EBV lytic cycle. TRIM5α also participates in Rta-induced mitophagy. The mechanism by which Rta and TRIM5α regulate mitochondrial dynamics will be further elucidated. | en |
| dc.description.provenance | Submitted by admin ntu (admin@lib.ntu.edu.tw) on 2024-08-15T16:11:22Z No. of bitstreams: 0 | en |
| dc.description.provenance | Made available in DSpace on 2024-08-15T16:11:22Z (GMT). No. of bitstreams: 0 | en |
| dc.description.tableofcontents | 口試委員審定書 i
致謝 ii 中文摘要 iii ABSTRACT iv 目次 v 表次 viii 圖次 ix 第一章 前言 1 1.1 EB 病毒 (Epstein-Barr virus, EBV) 1 1.1.1 EB 病毒的簡介 1 1.1.2 EB 病毒的感染和生活史 2 1.1.3 EB 病毒的極早期基因 4 1.2 Rta 蛋白質 4 1.2.1 Rta 的簡介 4 1.2.2 Rta 對病毒與宿主基因的調控 5 1.2.3 Rta 的轉譯後修飾 6 1.3 粒線體 6 1.3.1 粒線體動態平衡 (Mitochondrial dynamics) 6 1.3.2 粒線體自噬 (Mitophagy) 8 1.3.3 粒線體導向序列 (Mitochondrial targeting sequence, MTS) 8 1.3.4 病毒與粒線體動態平衡的調控 9 1.4 TRIM5α (Tripartite motif-containing protein 5 alpha) 10 1.4.1 TRIM5α 的簡介 10 1.4.2 TRIM5α 的抗病毒作用 11 1.4.3 TRIM5α 與粒線體的品質調節 12 1.5 研究動機 12 第二章 材料與方法 13 2.1 細胞株與細胞培養 13 2.2 菌株與細菌培養 13 2.3 質體 DNA 的萃取 13 2.4 質體與質體的建構 14 2.5 轉型作用 (Transformation) 14 2.6 轉染 (Transfection) 14 2.7 基因靜默分析 (shRNA knockdown) 15 2.8 粒線體分離 (Mitochondrial isolation) 15 2.9 西方墨點法 (Western blot, immunoblotting) 15 2.10 免疫螢光染色 (Immunofluorescence analysis) 16 第三章 結果 17 3.1 Rta 與 TRIM5α 位於粒線體的外膜上 17 3.2 Rta 增加 Drp1 的磷酸化並使其定位至粒線體 18 3.3 Rta 透過 Drp1 誘導粒線體自噬 18 3.4 Rta 的粒線體導向序列 (MTS) 分析以及 Rta(MTS) 突變株設計 19 3.5 Rta(MTS) 突變株在粒線體的定位減少 19 3.6 Rta(MTS) 突變株誘導粒線體自噬的能力降低 20 3.7 TRIM5α 會影響 Rta 誘導的粒線體自噬 20 3.8 Drp1 對 EB 病毒溶裂期的影響 21 第四章 討論 22 4.1 Rta 與 TRIM5α 位於粒線體的外膜上 22 4.2 Rta 會促進粒線體分裂 22 4.3 Rta 透過 Drp1 誘導粒線體自噬 22 4.4 Rta 的 MTS 與 Rta(MTS) 突變株的定位 23 4.5 MTS 的突變影響 Rta 誘導粒線體自噬的能力 23 4.6 TRIM5α 參與 Rta 誘導的粒線體自噬 24 4.7 粒線體自噬對 EB 病毒溶裂期的影響 24 參考文獻 26 第五章 圖表 43 附錄 68 | - |
| dc.language.iso | zh_TW | - |
| dc.subject | EB 病毒 | zh_TW |
| dc.subject | 粒線體自噬 | zh_TW |
| dc.subject | TRIM5α | zh_TW |
| dc.subject | Rta | zh_TW |
| dc.subject | 粒線體 | zh_TW |
| dc.subject | Rta | en |
| dc.subject | Epstein-Barr virus | en |
| dc.subject | Mitophagy | en |
| dc.subject | Mitochondria | en |
| dc.subject | TRIM5α | en |
| dc.title | EB病毒Rta蛋白質對粒線體自噬的影響 | zh_TW |
| dc.title | Role of Epstein-Barr virus Rta on mitophagy | en |
| dc.type | Thesis | - |
| dc.date.schoolyear | 112-2 | - |
| dc.description.degree | 碩士 | - |
| dc.contributor.oralexamcommittee | 劉世東;陳美如;邱亞芳;李家瑋 | zh_TW |
| dc.contributor.oralexamcommittee | Shih-Tung Liu;Mei-Ru Chen;Ya-Fang Chiu;Chia-Wei Lee | en |
| dc.subject.keyword | EB 病毒,Rta,TRIM5α,粒線體,粒線體自噬, | zh_TW |
| dc.subject.keyword | Epstein-Barr virus,Rta,TRIM5α,Mitochondria,Mitophagy, | en |
| dc.relation.page | 71 | - |
| dc.identifier.doi | 10.6342/NTU202403678 | - |
| dc.rights.note | 同意授權(限校園內公開) | - |
| dc.date.accepted | 2024-08-09 | - |
| dc.contributor.author-college | 生命科學院 | - |
| dc.contributor.author-dept | 生化科技學系 | - |
| dc.date.embargo-lift | 2029-08-06 | - |
| 顯示於系所單位： | 生化科技學系 | |
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