Sousa Ivanildo P, Carvalho Carlos A M, Mendes Ygara S, Weissmuller Gilberto, Oliveira Andréa C, Gomes Andre M O
Instituto de Bioquímica Médica Leopoldo de Meis, Centro de Ciências da Saúde and ‡Instituto de Biofísica Carlos Chagas Filho, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro , Rio de Janeiro, Rio de Janeiro 21941-902, Brazil.
Biochemistry. 2017 Oct 31;56(43):5823-5830. doi: 10.1021/acs.biochem.7b00650. Epub 2017 Oct 18.
Alphaviruses are enveloped arboviruses mainly proposed to infect host cells by receptor-mediated endocytosis followed by fusion between the viral envelope and the endosomal membrane. The fusion reaction is triggered by low pH and requires the presence of both cholesterol and sphingolipids in the target membrane, suggesting the involvement of lipid rafts in the cell entry mechanism. In this study, we show for the first time the interaction of an enveloped virus with membrane microdomains isolated from living cells. Using Mayaro virus (MAYV), a New World alphavirus, we verified that virus fusion to these domains occurred to a significant extent upon acidification, although its kinetics was quite slow when compared to that of fusion with artificial liposomes demonstrated in a previous work. Surprisingly, when virus was previously exposed to acidic pH, a condition previously shown to inhibit alphavirus binding and fusion to target membranes as well as infectivity, and then reneutralized, its ability to fuse with membrane microdomains at low pH was retained. Interestingly, this observation correlated with a partial reversion of low pH-induced conformational changes in viral proteins and retention of virus infectivity upon reneutralization. Our results suggest that MAYV entry into host cells could alternatively involve internalization via lipid rafts and that the conformational changes triggered by low pH in the viral spike proteins during the entry process are partially reversible.
甲病毒是包膜虫媒病毒，主要通过受体介导的内吞作用感染宿主细胞，随后病毒包膜与内体膜融合。融合反应由低pH触发，且需要靶膜中同时存在胆固醇和鞘脂，这表明脂筏参与了细胞进入机制。在本研究中，我们首次展示了包膜病毒与从活细胞中分离出的膜微区的相互作用。使用新世界甲病毒马亚罗病毒（MAYV），我们证实，病毒与这些微区的融合在酸化时会大量发生，尽管与先前工作中展示的与人工脂质体的融合动力学相比，其动力学相当缓慢。令人惊讶的是，当病毒先前暴露于酸性pH（先前已证明该条件会抑制甲病毒与靶膜的结合、融合以及感染性）然后再中和时，其在低pH下与膜微区融合的能力得以保留。有趣的是，这一观察结果与病毒蛋白中低pH诱导的构象变化的部分逆转以及再中和后病毒感染性的保留相关。我们的结果表明，MAYV进入宿主细胞可能还涉及通过脂筏内化，并且在进入过程中低pH在病毒刺突蛋白中触发的构象变化是部分可逆的。
