DNAJB6a調控EB病毒溶裂期中引子酶BSLF1核定位與病毒衣殼組裝之機制探討

Abstract

EB病毒 (Epstein-Barr virus) 在感染細胞，進入溶裂期時，有多種缺乏典型核定位訊號 (nuclear localization signal) 的病毒蛋白仍必須進入細胞核才能執行功能。本研究發現宿主的HSP70共伴侶蛋白DNAJB6a是其中EB病毒引子酶BSLF1關鍵的入核調控因子。我們證實DNAJB6a可直接與BSLF1結合，並將其運送至細胞核內；相對地，其細胞質型的同種異構物DNAJB6b則無法執行此功能。BSLF1依賴DNAJB6a入核的過程，不僅依賴DNAJB6a本身的核定位能力，也需要其招募HSP70 (特別是HSPA1A) 的功能。當細胞中DNAJB6a缺失時，即使BSLF1與解旋酶-引子酶複合體成員共表現時具有部分入核的能力，BSLF1的入核能力仍受阻，導致病毒DNA複製減少以及病毒粒子釋放顯著下降。除了在病毒早期複製階段的角色外，DNAJB6a在EBV後期的組裝過程也發揮關鍵的功能。我們的結果顯示，攜帶NLS的病毒次要衣殼蛋白BORF1與DNAJB6a會一同被招募至PML核小體。這些結構被認為是EB病毒衣殼組裝的場所。DNAJB6a與BORF1形成複合體，並可能藉由HSPA1A穩定BORF1蛋白；反之，當細胞缺乏DNAJB6a時，BORF1會出現錯誤定位並快速降解，可能干擾病毒衣殼的正常形成。而DNAJB6a的過度表現則會提升BORF1表現量並促進較大的BORF1核內點狀結構生成，此作用亦依賴其與HSP70結合的能力。除此之外，我們進一步提出BSLF1藉DNAJB6a的幫助進入細胞核後EB病毒的蛋白激酶BGLF4可能藉由競爭BSLF1與DNAJB6a的結合幫助BSLF1從細胞核膜附近重新分布至病毒複製區域。綜合而言，本研究揭示DNAJB6a在EB病毒溶裂期中具備雙重功能：協助病毒複製酶進入細胞核，並協調病毒衣殼在細胞核內的組裝。我們的結果凸顯 EBV 如何挾持宿主伴侶蛋白，以精準調控其生命週期中的關鍵步驟。

During Epstein-Barr virus (EBV) lytic reactivation, several viral proteins lacking classical nuclear localization signals (NLS) must still translocate into the nucleus to perform their functions. In this study, we identify the host HSP70 co-chaperone DNAJB6a as a key mediator of nuclear import for the viral primase BSLF1. We demonstrate that DNAJB6a directly binds BSLF1 and facilitates its nuclear entry, whereas the cytosolic isoform DNAJB6b fails to do so. This nuclear import process requires both the nuclear localization capacity of DNAJB6a and its ability to recruit HSP70, particularly HSPA1A. Notably, depletion of DNAJB6a impairs BSLF1 nuclear accumulation even when its helicase-primase complex partners are co-expressed, leading to reduced viral DNA replication and markedly diminished virion release. Beyond its role in early replication, DNAJB6a also contributes to the late phase of viral assembly. We show that BORF1, a minor capsid protein bearing an NLS, recruits DNAJB6a to promyelocytic leukemia (PML) nuclear bodies, which serve as assembly hubs for EBV capsids. DNAJB6a forms a complex with BORF1 and may stabilize the BORF1 protein through HSPA1A. In contrast, DNAJB6a depletion results in BORF1 mislocalization and rapid degradation, likely disrupting capsid formation. Overexpression of DNAJB6a enhances BORF1 protein levels and promotes the formation of larger nuclear puncta, an effect dependent on its HSP70-binding activity. Furthermore, we propose that once BSLF1 enters the nucleus via DNAJB6a, the viral kinase BGLF4 may facilitate its redistribution from the nuclear rim to replication compartments by displacing DNAJB6a. Together, our findings reveal a dual role for DNAJB6a in EBV lytic replication: directing nuclear import of a key viral replication enzyme and coordinating capsid assembly within the nucleus. This study highlights how EBV hijacks host chaperone machinery to precisely orchestrate critical stages of its life cycle.