<br>核输出选择性抑制剂 (SINE) 化合物改变新世界甲病毒衣壳定位并减少哺乳动物细胞中的病毒复制。,PLOS Neglected Tropical Diseases

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核输出选择性抑制剂 (SINE) 化合物改变新世界甲病毒衣壳定位并减少哺乳动物细胞中的病毒复制。
PLOS Neglected Tropical Diseases ( IF 3.4 ) Pub Date : 2016-11-30 , DOI: 10.1371/journal.pntd.0005122
Lindsay Lundberg ₁ , Chelsea Pinkham ₁ , Cynthia de la Fuente ₁ , Ashwini Brahms ₁ , Nazly Shafagati ₁ , Kylie M Wagstaff ₂ , David A Jans ₂ , Sharon Tamir ₃ , Kylene Kehn-Hall ₁

Affiliation

新世界甲病毒、委内瑞拉马脑炎病毒 (VEEV) 的衣壳结构蛋白与输入 α/β1 和 CRM1 的宿主核转运蛋白相互作用。新型选择性核输出抑制剂 (SINE) 化合物 KPT-185、KPT-335 (verdinexor) 和 KPT-350 以与典型抑制剂 Leptomycin B 类似的方式靶向宿主的主要核输出蛋白 CRM1。 Leptomycin B 的局限性在于它与 CRM1 不可逆结合； SINE 化合物可以缓解这种情况，因为它们是缓慢可逆的。免疫荧光共聚焦显微镜表明，与无活性的化合物 KPT-301 相比，用这些化合物化学抑制 CRM1 增强了衣壳向细胞核的定位。细胞外与细胞内病毒RNA的差异，以及无细胞上清液中衣壳的减少，表明抑制剂影响了病毒组装，从而导致病毒滴度降低。使用荧光素酶标记的病毒并通过噬菌斑测定证实了病毒复制的减少。 SINE 化合物对 VEEV TC83_Cm 没有影响，VEEV TC83_Cm 编码无法进入细胞核的突变形式的衣壳。在 KPT-185 存在的情况下连续传代 VEEV 会导致衣壳的核定位和核输出信号发生突变。最后，SINE 复合治疗还降低了相关的东西方马脑炎病毒的病毒滴度，这表明 CRM1 与新世界甲病毒属的衣壳蛋白保持着共同的相互作用。

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Selective Inhibitor of Nuclear Export (SINE) Compounds Alter New World Alphavirus Capsid Localization and Reduce Viral Replication in Mammalian Cells.

The capsid structural protein of the New World alphavirus, Venezuelan equine encephalitis virus (VEEV), interacts with the host nuclear transport proteins importin α/β1 and CRM1. Novel selective inhibitor of nuclear export (SINE) compounds, KPT-185, KPT-335 (verdinexor), and KPT-350, target the host's primary nuclear export protein, CRM1, in a manner similar to the archetypical inhibitor Leptomycin B. One major limitation of Leptomycin B is its irreversible binding to CRM1; which SINE compounds alleviate because they are slowly reversible. Chemically inhibiting CRM1 with these compounds enhanced capsid localization to the nucleus compared to the inactive compound KPT-301, as indicated by immunofluorescent confocal microscopy. Differences in extracellular versus intracellular viral RNA, as well as decreased capsid in cell free supernatants, indicated the inhibitors affected viral assembly, which led to a decrease in viral titers. The decrease in viral replication was confirmed using a luciferase-tagged virus and through plaque assays. SINE compounds had no effect on VEEV TC83_Cm, which encodes a mutated form of capsid that is unable to enter the nucleus. Serially passaging VEEV in the presence of KPT-185 resulted in mutations within the nuclear localization and nuclear export signals of capsid. Finally, SINE compound treatment also reduced the viral titers of the related eastern and western equine encephalitis viruses, suggesting that CRM1 maintains a common interaction with capsid proteins across the New World alphavirus genus.

更新日期：2019-11-01

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