EB 病毒的 BFRF3 在 VCA 入核機制中所扮演的角色

Abstract

Epstein-BarrVirus(EBV)，屬於皰疹病毒 γ亞科，已被發現和許多腫瘤性癌症有關。 EBV 平時處於潛伏期，當受到誘導或環境刺激時，會進入溶裂期並形成病毒顆粒。 在溶裂期的過程中，轉譯出的外鞘蛋白質會進入細胞核進行外鞘組裝。但在六種外 鞘蛋白質中，卻只有 BORF1、BVRF2 和 BdRF1 具有入核序列，並可以藉此序列 進入細胞核，其餘外鞘蛋白質的入核機制則尚未完全明瞭。在簡單皰疹病毒中，架 構蛋白質具有協助主外鞘蛋白質入核的能力 ; 然而在 EBV 中，免疫螢光染色的結 果發現只有在 BFRF3 共同存在的情形下，VCA 才能藉由 BdRF1 進入細胞核。因 此，本研究欲探究 BFRF3 在 VCA 入核機制中所扮演的角色。首先證實了 VCA 會 與 BFRF3 結合，接著蔗糖濃度梯度分析的結果顯示 BFRF3 會解開 VCA 的多聚體， 並以氨基酸剔除的實驗證實 BFRF3 氨基酸序列 1-65 的區段負責解開 VCA 的聚合 體。最後，本研究也發現 VCA、BdRF1 和 BFRF3 入核後會聚集於核內的 PML- NBs。綜合以上結果，本研究對於 EBV 的外鞘蛋白質如何入核進行外鞘的組裝提 出更深入的機制探討。

Epstein-Barr Virus (EBV) belongs to the gammaherpesvirus family, and is known to be associated with several neoplastic diseases. EBV usually remains latent after infection. Upon stimulated by environmental factors, EBV would enter into the lytic cycle and then produce infective viral particles. During the lytic cycle, capsid proteins need to be translocated into the nucleus for capsid assembly. Among them, only BORF1, BVRF2, and BdRF1 contain the nuclear localization signal (NLS), while the translocation pathways of other capsid proteins are unclear. In herpes simplex virus 1 (HSV-1), scaffold protein is responsible for the translocation of major capsid protein. However, the immunofluorescence results indicated that translocation of VCA is not dependent on BdRF1, unless BFRF3 is also present. Therefore, this study aims to clarify the involvement of BFRF3 in the translocation of VCA. This study found that BFRF3 binds to VCA. Sucrose gradient analysis subsequently revealed that BFRF3 is required for the reduction of VCA oligomerization. Deletion analysis indicates that amino acid residues 1-65 of BFRF3 is required for VCA de-oligomerization. Finally, this study confirms that VCA, BFRF3, and BdRF1 aggregate at promyelocytic leukemia nuclear bodies (PML- NBs). The study provides insight into the mechanism how capsid proteins were translocated into the nucleus for capsid assembly.