Ubiquitin-regulated nuclear-cytoplasmic trafficking of the Nipah virus matrix protein is important for viral budding.
Ubiquitin-regulated nuclear-cytoplasmic trafficking of the Nipah virus matrix protein is important for viral budding.
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DOI：
10.1371/journal.ppat.1001186
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发表时间：
2010-11-11
期刊：
影响因子：
6.7
通讯作者：
Lee B
中科院分区：
文献类型：
--
作者：
Wang YE;Park A;Lake M;Pentecost M;Torres B;Yun TE;Wolf MC;Holbrook MR;Freiberg AN;Lee B
已知副粘病毒在细胞质中复制并从质膜出芽。基质是副粘病毒中介导病毒组装和出芽的主要结构蛋白。奇怪的是，在细胞核中发现了一些副粘病毒的基质蛋白，尽管与这种核定位相关的生物学功能仍然不清楚。我们在这里报告，尼帕病毒基质（NiV-M）蛋白的核细胞质运输和相关的翻译后修饰在基质介导的病毒出芽中发挥着关键作用。尼帕病毒 (NiV) 是一种高致病性新兴副粘病毒，可引起人类致命性脑炎，被列为生物安全 4 级 (BSL4) 病原体。在活 NiV 感染期间，NiV-M 在感染早期首先在细胞核中检测到，然后定位到细胞质和质膜。 NiV-M 中发现的假定二分核定位信号 (NLS) 和富含亮氨酸的核输出信号 (NES) 的突变损害了其核-细胞质运输，并消除了 NiV-M 出芽。 NLS 中高度保守的赖氨酸残基具有双重功能：其正电荷对于介导核输入很重要，而且它也是调节蛋白质核输出的单泛素化的潜在位点。一致地，泛素的过度表达增强了 NiV-M 出芽，而细胞中游离泛素的消耗（通过蛋白酶体抑制剂）导致 NiV-M 的核滞留并阻止病毒出芽。活尼帕病毒出芽对蛋白酶体抑制剂极其敏感：硼替佐米（bortezomib）是 FDA 批准的一种用于治疗多发性骨髓瘤的蛋白酶体抑制剂，可降低病毒滴度，IC50 为 2.7 nM，比人类可达到的血浆峰值浓度低 100 倍。这开启了使用“现成”疗法对抗急性 NiV 感染的可能性。尼帕病毒（NiV）是一种致命的新出现的病毒，会引起致命的大脑炎症，并且感染人类的死亡率很高。 NiV 和密切相关的亨德拉病毒 (HeV) 也可以感染猪和马等重要农业牲畜。由于缺乏有效的疫苗和治疗方法，以及它们对农业和公共卫生构成的持续威胁，导致 NiV 和 HeV 被归类为生物安全 4 级 (BSL4) 病原体。已知副粘病毒如 NiV 在细胞质中复制并从质膜出芽。病毒的组装和出芽是由基质结构蛋白介导的。然而，我们出乎意料地发现，NiV 的基质蛋白需要穿过细胞核才能获得定位和从质膜出芽的功能。尽管 NiV-M 具有假定的核输入和输出信号，但我们还发现 NiV-M 中保守的赖氨酸残基的泛素化对于核输出、随后的膜定位和病毒出芽至关重要。蛋白酶体抑制剂可以消耗细胞中的游离泛素，有效降低活 NiV 感染期间的病毒滴度，为急性 NiV 感染的治疗开辟了新的可能性。
Paramyxoviruses are known to replicate in the cytoplasm and bud from the plasma membrane. Matrix is the major structural protein in paramyxoviruses that mediates viral assembly and budding. Curiously, the matrix proteins of a few paramyxoviruses have been found in the nucleus, although the biological function associated with this nuclear localization remains obscure. We report here that the nuclear-cytoplasmic trafficking of the Nipah virus matrix (NiV-M) protein and associated post-translational modification play a critical role in matrix-mediated virus budding. Nipah virus (NiV) is a highly pathogenic emerging paramyxovirus that causes fatal encephalitis in humans, and is classified as a Biosafety Level 4 (BSL4) pathogen. During live NiV infection, NiV-M was first detected in the nucleus at early stages of infection before subsequent localization to the cytoplasm and the plasma membrane. Mutations in the putative bipartite nuclear localization signal (NLS) and the leucine-rich nuclear export signal (NES) found in NiV-M impaired its nuclear-cytoplasmic trafficking and also abolished NiV-M budding. A highly conserved lysine residue in the NLS served dual functions: its positive charge was important for mediating nuclear import, and it was also a potential site for monoubiquitination which regulates nuclear export of the protein. Concordantly, overexpression of ubiquitin enhanced NiV-M budding whereas depletion of free ubiquitin in the cell (via proteasome inhibitors) resulted in nuclear retention of NiV-M and blocked viral budding. Live Nipah virus budding was exquisitely sensitive to proteasome inhibitors: bortezomib, an FDA-approved proteasome inhibitor for treating multiple myeloma, reduced viral titers with an IC50 of 2.7 nM, which is 100-fold less than the peak plasma concentration that can be achieved in humans. This opens up the possibility of using an “off-the-shelf” therapeutic against acute NiV infection. Nipah virus (NiV) is a lethal, newly emerging virus that causes fatal inflammation of the brain and has a high death rate in infected humans. NiV and the closely related Hendra virus (HeV) can also infect agriculturally important livestock such as pigs and horses. The lack of effective vaccines and treatments, and the ongoing threat they pose to both agriculture and public health, have led to the classification of NiV and HeV as Biosafety Level 4 (BSL4) pathogens. Paramyxoviruses such as NiV are known to replicate in the cytoplasm and bud from the plasma membrane. Viral assembly and budding is mediated by the matrix structural protein. However, we found, quite unexpectedly, that the matrix protein of NiV needs to transit through the nucleus before gaining the functional ability to localize and bud from the plasma membrane. Although NiV-M has putative nuclear import and export signals, we also found that ubiquitination of a conserved lysine residue in NiV-M is critical for nuclear export, subsequent membrane localization and viral budding. Proteasome inhibitors, which deplete cellular pools of free ubiquitin, potently reduce viral titers during live NiV infection, opening up new possibilities for therapeutics against acute NiV infection.
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