EB病毒EBNA2蛋白質對細胞週期進行及PML核體結構之影響

Abstract

Epstein-Bar virus (EBV)是一種廣泛分布於全世界的人類皰疹病毒(herpes virus)，主要感染人類B淋巴球與上皮細胞，目前被認為與多種人類惡性腫瘤有關，像是巴氏淋巴瘤(Berkitt’s lymphoma)及鼻咽癌(nasopharyngeal carcinoma)。Epstein-Barr virus nuclear antigen 2 (EBNA2)是細胞感染EBV後最早表現的病毒轉錄活化子，已知在EBV引發的B淋巴球轉型過程中是個不可或缺的因子，而且在基因轉殖鼠中表現能誘發腫瘤形成。雖然已有研究顯示EBNA2的轉錄活化功能與其引起細胞轉形癌化有關，但對EBNA2是如何參與EBV引發腫瘤形成目前仍未完全清楚。有研究指出，EBV感染B淋巴球後會啟動細胞內的DNA損傷反應(DNA damage response; DDR)導致大部分受感染的B淋巴球進入生長停滯狀態。在上皮細胞，我們實驗室已先觀察到EBNA2會造成DNA雙股斷裂的累積、p53的活化並抑制細胞生長。在本研究中，我們顯示EBNA2在上皮細胞及B細胞中能夠活化DDR並可能藉此活化p53使上皮細胞細胞週期停滯在G1 phase。我們也進一步發現EBNA2引發的G1 phase停滯需要PML。重要的是，由於我們觀察到EBNA2表現會導致PML蛋白質表現量上升並有利PML進行polysumoylation，透過三維影像重建我們確認EBNA2在人類上皮細胞中會影響PML-NB結構。由於目前PML-NBs被廣泛認為在腫瘤抑制及抗病毒反應上都是重要的樞紐，我們相信這些研究結果將有助於了解EBNA2在EBV引發腫瘤形成及EBV的生命週期中所扮演的角色。

Epstein-Barr virus (EBV), an ubiquitous human herpes virus which preferentially infects human B lymphocytes and epithelial cells, is being causally implicated in numerous human malignancies, notably Burkett’s lymphoma and nasopharyngeal carcinoma. Among the first viral proteins expressed after EBV infection, Epstein-Barr virus nuclear antigen 2 (EBNA2), a viral transactivator, is known for its indispensability in initiating in vitro B cell transformation and for inducing tumor in transgenic mice. Despite accumulating evidences linking transactivation activity to EBNA2 tumorigenecity, elusiveness still remain in how the viral protein contributes to EBV’s oncogenic nature. A recent study, which pointed out a growth-arresting DNA damage response (DDR) was provoked after EBV infection of primary human B cells, provided a clue. In our previous studies, upon EBNA2 expression in human epithelial cells accumulation of DNA double-strand breaks could be observed and a p53-mediated growth retardation also became apparent. In the present study, we showed that EBNA2 expression alone in human epithelial and B cell lines could lead to DDR activation and result in a G1 growth arrest, which was possibly mediated by p53 activation. Moreover, we demonstrated that PML was required for this EBNA2-induced G1 arrest. And most importantly, we found that EBNA2 expression led to upregulation of PML protein levels and increased PML polysumoylation. Accordingly, by conducting 3D reconstuction, we confirmed that EBNA2 expression could alter PML-NB structure in human epithelial cell. Since PML-NBs are now widely appreciated as crucial nodes in both tumor suppression and anti-viral response, we believe that our findings may shed light on the role of EBNA2 in EBV-mediated tumorigenesis and EBV life cycle.