An exploration of conditions proposed to trigger the Ebola virus glycoprotein for fusion
An exploration of conditions proposed to trigger the Ebola virus glycoprotein for fusion
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DOI：
10.1371/journal.pone.0219312
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发表时间：
2019-07-05
期刊：
影响因子：
3.7
通讯作者：
White, Judith M.
中科院分区：
文献类型：
--
作者：
Feneant, Lucie;Szymanska-de Wijs, Katarzyna M.;White, Judith M.
埃博拉病毒继续造成可怕的疾病并灌输恐惧。2013-2016年在西非爆发的疫情造成了难以估量的发病率和死亡率（超过1.1万人死亡），第二大疫情正在刚果民主共和国发生。埃博拉病毒感染的第一阶段是进入，最终将病毒基因组送入细胞质以开始复制。在包膜病毒中，埃博拉病毒使用复杂的进入途径：它们与细胞表面的附着因子结合，被巨噬细胞吞噬，并通过内体系统运输。在此过程中，糖蛋白（GP）的受体结合亚基被组织蛋白酶从类似于130 kDa减少到类似于19 kDa。这一事件允许裂解的GP（GP(cl)）与其内体受体Niemann-Pick C1 （NPC1）结合。然后病毒与一个晚期内体膜融合，但这是如何发生的仍然是一个有争议的话题。早期但长期的观察是，携带GP（cl）的颗粒的进入被升高内体pH或抑制半胱氨酸蛋白酶的药物所抑制，这表明需要一个额外的因素。然而，一些人得出结论，NPC1足以触发GP（cl）的融合活性。在这里，我们使用敏感细胞-细胞和假病毒-细胞融合试验重新检查了这个问题。在任何pH下，我们都没有观察到GP（cl）介导的与NPC1或其GP（cl）结合域的融合，而在低pH下，我们一直观察到与淋巴细胞性脉络膜脑膜炎病毒的GP的融合。添加建议的融合增强因子-阳离子（Ca++和K+）、一种还原剂、阴离子脂质双（单酰基甘油）磷酸，以及组织蛋白酶B和l的混合物，都没有诱导到可检测的融合。我们的发现与先前的建议一致，即需要一个额外的因子来触发GP（cl）在与NPC1结合后的完全融合活性。我们讨论了研究的注意事项以及可能缺少的因素。
Ebolaviruses continue to inflict horrific disease and instill fear. The 2013-2016 outbreak in Western Africa caused unfathomable morbidity and mortality (over 11,000 deaths), and the second largest outbreak is on-going in the Democratic Republic of the Congo. The first stage of an Ebolavirus infection is entry, culminating in delivery of the viral genome into the cytoplasm to initiate replication. Among enveloped viruses, Ebolaviruses use a complex entry pathway: they bind to attachment factors on cell surfaces, are engulfed by macropinocytosis, and traffic through the endosomal system. En route, the receptor binding subunit of the glycoprotein (GP) is reduced from similar to 130 to similar to 19 kDa by cathepsins. This event allows cleaved GP (GP(cl)) to bind to Niemann-Pick C1 (NPC1), its endosomal receptor. The virus then fuses with a late endosomal membrane, but how this occurs remains a subject of debate. An early, but standing, observation is that entry of particles bearing GP(cl) is inhibited by agents that raise endosomal pH or inhibit cysteine proteases, suggesting the need for an additional factor(s). Yet, some have concluded that NPC1 is sufficient to trigger the fusion activity of GP(cl). Here, we re-examined this question using sensitive cell-cell and pseudovirus-cell fusion assays. We did not observe detectable GP(cl)-mediated fusion with NPC1 or its GP(cl) binding domain at any pH tested, while robust fusion was consistently observed with GP from lymphocytic choriomeningitis virus at low pH. Addition of proposed fusion-enhancing factors-cations (Ca++ and K+), a reducing agent, the anionic lipid Bis(Monoacylglycero)Phosphate, and a mixture of cathepsins B and L-did not induce detectable fusion. Our findings are in line with the earlier proposal that an additional factor is required to trigger the full fusion activity of GP(cl) after binding to NPC1. We discuss caveats to our study and what the missing factor(s) might be.