Department of Life Sciences, Presidency University, West Bengal, India.
Laboratory of Molecular Biology, School of Biological Sciences, Indian Association for the Cultivation of Science, Jadavpur, Kolkata, India.
PLoS Pathog. 2020 Feb 24;16(2):e1008105. doi: 10.1371/journal.ppat.1008105. eCollection 2020 Feb.
Epstein-Barr virus (EBV) nuclear oncoprotein EBNA3C is essential for B-cell transformation and development of several B-cell lymphomas particularly those are generated in an immuno-compromised background. EBNA3C recruits ubiquitin-proteasome machinery for deregulating multiple cellular oncoproteins and tumor suppressor proteins. Although EBNA3C is found to be ubiquitinated at its N-terminal region and interacts with 20S proteasome, the viral protein is surprisingly stable in growing B-lymphocytes. EBNA3C can also circumvent autophagy-lysosomal mediated protein degradation and subsequent antigen presentation for T-cell recognition. Recently, we have shown that EBNA3C enhances autophagy, which serve as a prerequisite for B-cell survival particularly under growth deprivation conditions. We now demonstrate that proteasomal inhibition by MG132 induces EBNA3C degradation both in EBV transformed B-lymphocytes and ectopic-expression systems. Interestingly, MG132 treatment promotes degradation of two EBNA3 family oncoproteins-EBNA3A and EBNA3C, but not the viral tumor suppressor protein EBNA3B. EBNA3C degradation induced by proteasomal inhibition is partially blocked when autophagy-lysosomal pathway is inhibited. In response to proteasomal inhibition, EBNA3C is predominantly K63-linked polyubiquitinated, colocalized with the autophagy-lysosomal fraction in the cytoplasm and participated within p62-LC3B complex, which facilitates autophagy-mediated degradation. We further show that the degradation signal is present at the first 50 residues of the N-terminal region of EBNA3C. Proteasomal inhibition reduces the colony formation ability of this important viral oncoprotein, induces apoptotic cell death and increases transcriptional activation of both latent and lytic gene expression which further promotes viral reactivation from EBV transformed B-lymphocytes. Altogether, this study offers rationale to use proteasome inhibitors as potential therapeutic strategy against multiple EBV associated B-cell lymphomas, where EBNA3C is expressed.
爱泼斯坦-巴尔病毒 (EBV) 核癌蛋白 EBNA3C 是 B 细胞转化和几种 B 细胞淋巴瘤发展所必需的，特别是在免疫功能低下的背景下产生的那些淋巴瘤。EBNA3C 募集泛素-蛋白酶体机制来调节多种细胞癌蛋白和肿瘤抑制蛋白。尽管已经发现 EBNA3C 在其 N 端区域被泛素化，并与 20S 蛋白酶体相互作用，但在生长的 B 淋巴细胞中，病毒蛋白惊人地稳定。EBNA3C 还可以规避自噬溶酶体介导的蛋白降解和随后的抗原呈递，以进行 T 细胞识别。最近，我们已经证明 EBNA3C 增强了自噬作用，这是 B 细胞在生长剥夺条件下生存的前提。我们现在证明，蛋白酶体抑制剂 MG132 可诱导 EBV 转化的 B 淋巴细胞和异位表达系统中的 EBNA3C 降解。有趣的是，MG132 处理促进了两种 EBNA3 家族癌蛋白-EBNA3A 和 EBNA3C 的降解，但不促进病毒肿瘤抑制蛋白 EBNA3B 的降解。当自噬溶酶体途径被抑制时，蛋白酶体抑制诱导的 EBNA3C 降解部分受阻。在蛋白酶体抑制的反应中，EBNA3C 主要被 K63 连接的多泛素化，与细胞质中的自噬溶酶体部分共定位，并参与 p62-LC3B 复合物中，促进自噬介导的降解。我们进一步表明，降解信号存在于 EBNA3C N 端区域的前 50 个残基中。蛋白酶体抑制降低了这种重要病毒癌蛋白的集落形成能力，诱导细胞凋亡死亡，并增加潜伏和裂解基因表达的转录激活，这进一步促进了 EBV 转化的 B 淋巴细胞中的病毒重新激活。总之，这项研究为使用蛋白酶体抑制剂作为针对多种 EBV 相关 B 细胞淋巴瘤的潜在治疗策略提供了依据，其中表达 EBNA3C。
