Please use this identifier to cite or link to this item:
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/90625
Title: | EB病毒之BGLF1與出核相關蛋白質的結合 Interaction of Epstein-Barr virus BGLF1 with proteins involved in nuclear egress |
Authors: | 張舒茵 Shu-Yin Chang |
Advisor: | 張麗冠 Li-Kwan Chang |
Keyword: | EB病毒,BGLF1,出核(nuclear egress),BFLF2,BFRF1,殼體組裝(capsid assembly), Epstein-Barr virus,BGLF1,nuclear egress,BFLF2,BFRF1,capsid assembly, |
Publication Year : | 2023 |
Degree: | 碩士 |
Abstract: | Epstein-Barr virus為第四型人類皰疹病毒,同時也是第一個被發現與人類癌症相關的病毒,並且全球九成的人口被 EB 病毒感染。EB 病毒的生活史可以分成潛伏期 (latency) 以及溶裂期 (lytic cycle) ,在潛伏期只會有少量的病毒基因表現,以維持病毒基因體保留在宿主細胞中,而在經過適當的刺激後,會使 EB 病毒進入溶裂期,產生成熟的病毒顆粒從宿主細胞釋放。目前已知病毒顆粒的 Capsid-associated tegument complex (CATC) 對於病毒殼體的穩定性及病毒顆粒出核 (nuclear egress) 具有重要功能。而 EB 病毒的 CATC 是由 BGLF1、BPLF1 及 BVRF1 所組成,本研究室先前的研究發現 BGLF1 除了能與殼體蛋白結合之外,並且能與在出核過程中扮演重要角色的 BFLF2 結合,因此推測 BGLF1 可能會參與出核的過程,不過詳細的機制尚不清楚。為了進一步釐清 BGLF1 在出核過程中跟相關蛋白質之間的結合,本研究透過 GST pull-down assay 證明 BGLF1 與 BFLF2 和 BFRF1 之間是直接結合,並且透過免疫螢光染色發現在 nuclear egress complex (NEC) 存在的情況下,會共定位在核膜。接著為了確認在出核過程中,procapsid 是否必須藉由 BGLF1 與 NEC 之間的結合,才能將其順利移動到核膜,同樣藉由 GST pull-down assay 釐清殼體蛋白與組成 NEC 的蛋白質之間的結合情形,結果發現組成病毒殼體的蛋白質包含 VCA、BFRF3、BDLF1 及 BORF1,在體外條件下皆能直接與 BFLF2 和 BFRF1 結合。另外本研究透過免疫螢光染色觀察帶有標籤的 BGLF1、BFLF2、BFRF3 以及 Rta 在 HeLa 細胞的分佈,確認在細胞內 BGLF1 與 BFLF2、BFRF3 會在細胞質共定位,而與 Rta 在細胞核以及細胞質共定位。希望透過本研究能幫助釐清 BGLF1 與其他病毒蛋白的結合關係,進一步推測 BGLF1 在 EB 病毒出核時所扮演的角色。 Epstein-Barr virus (EBV) is a type 4 human herpesvirus and the first virus discovered to be associated with human cancers. It infects approximately 90% of the global population. The life cycle of EBV can be divided into latency and lytic phase. During latency, only few viral genes are expressed to maintain viral genome replication. Upon appropriate stimulation, EBV enters the lytic phase, leading to the production and release of mature viral particles from host cells. Capsid-associated tegument complex (CATC) have been found to play important roles in stabilizing the viral capsid and facilitating nuclear egress of viral particles. The CATC proteins of EBV include BGLF1, BPLF1, and BVRF1. Previous study in our laboratory has shown that BGLF1 not only interacts with capsid proteins but also associates with BFLF2, which plays a crucial role in the nuclear egress. This suggests that BGLF1 is involved in the nuclear egress process, although the detailed mechanism remains unclear. To further elucidate the interactions between BGLF1 and relevant proteins during nuclear egress, this study implemented GST pull-down assays to demonstrate direct interactions between BGLF1 and BFLF2 as well as BFRF1. Immunofluorescence analysis revealed that BGLF1 colocalizes with nuclear egress complex (NEC) at the nuclear membrane. Then, in order to confirm whether BGLF1 is indispensable for recruitment of procapsid to the nuclear membrane during nuclear egress, GST pull-down assays indicated the interactions between capsid proteins and NEC components. The results showed that viral capsid proteins including VCA, BFRF3, BDLF1, and BORF1 directly interact with BFLF2 and BFRF1 under in vitro conditions. Additionally, this study examined the distribution of tagged BGLF1, BFLF2, BFRF3, and Rta in HeLa cells through immunofluorescence analysis and found that BGLF1 colocalized with BFLF2 as well as BFRF3 in the cytoplasm, while colocalized with Rta in both the nucleus and cytoplasm. Collective of this study clarifies the interactions between BGLF1 and other viral proteins, further elucidating the role of BGLF1 in EBV nuclear egress. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/90625 |
DOI: | 10.6342/NTU202302766 |
Fulltext Rights: | 同意授權(限校園內公開) |
Appears in Collections: | 生命科學系 |
Files in This Item:
File | Size | Format | |
---|---|---|---|
ntu-111-2.pdf Restricted Access | 3.46 MB | Adobe PDF | View/Open |
Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.