In vitro reconstitution of herpes simplex virus 1 fusion identifies low pH as a fusion co-trigger.
In vitro reconstitution of herpes simplex virus 1 fusion identifies low pH as a fusion co-trigger.
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单纯疱疹病毒 1 (HSV-1) 介导的膜融合是一个复杂的多蛋白过程，由受体触发，可发生在细胞表面和内体中。为了解决这种复杂性，我们在体外用合成脂质囊泡重建了 HSV-1 融合体。使用这种简化的可控系统，我们发现 HSV-1 融合不仅需要同源宿主受体，而且需要低 pH 值。在目标膜一侧，有效的融合需要胆固醇、内体膜中发现的带负电的脂质以及脂质有序和无序的最佳平衡。在病毒颗粒方面，四种 HSV-1 进入糖蛋白（gB、gD、gH 和 gL）足以进行融合。我们认为低 pH 值是 HSV-1 融合的生物学相关共同触发因素。融合对低 pH 值和内体脂质的依赖性可以解释为什么 HSV-1 通过内吞作用进入大多数细胞类型。我们假设在中性 pH 条件下，其他但未定义的细胞因素可能充当融合共同触发因素。这里建立的体外融合系统可用于系统地研究HSV-1介导的膜融合。 HSV-1 会导致终身无法治愈的感染和疾病，包括皮肤粘膜病变和致命性脑炎。病毒和宿主膜的融合是HSV-1感染靶细胞的关键步骤，需要病毒和宿主双方的多种因素。由于这种复杂性，许多基本问题仍未得到解答，例如 HSV-1 介导的膜融合所必需和充分的病毒和宿主因子的身份以及融合触发因素的性质。在这里，我们开发了一种简化的体外融合测定来检查融合要求，并确定低 pH 值作为病毒介导的体外融合的共同触发因素。我们假设低 pH 在细胞进入以及潜在的发病机制中具有关键作用。
Membrane fusion mediated by herpes simplex virus 1 (HSV-1) is a complex, multi-protein process that is receptor triggered and can occur both at the cell surface and in endosomes. To deconvolute this complexity, we reconstituted HSV-1 fusion with synthetic lipid vesicles in vitro. Using this simplified, controllable system, we discovered that HSV-1 fusion required not only a cognate host receptor but also low pH. On the target membrane side, efficient fusion required cholesterol, negatively charged lipids found in the endosomal membranes, and an optimal balance of lipid order and disorder. On the virion side, the four HSV-1 entry glycoproteins—gB, gD, gH, and gL—were sufficient for fusion. We propose that low pH is a biologically relevant co-trigger for HSV-1 fusion. The dependence of fusion on low pH and endosomal lipids could explain why HSV-1 enters most cell types by endocytosis. We hypothesize that under neutral pH conditions, other, yet undefined, cellular factors may serve as fusion co-triggers. The in vitro fusion system established here can be employed to systematically investigate HSV-1-mediated membrane fusion. HSV-1 causes lifelong, incurable infections and diseases ranging from mucocutaneous lesions to fatal encephalitis. Fusion of viral and host membranes is a critical step in HSV-1 infection of target cells that requires multiple factors on both the viral and host sides. Due to this complexity, many fundamental questions remain unanswered, such as the identity of the viral and host factors that are necessary and sufficient for HSV-1-mediated membrane fusion and the nature of the fusion trigger. Here, we developed a simplified in vitro fusion assay to examine the fusion requirements and identified low pH as a co-trigger for virus-mediated fusion in vitro. We hypothesize that low pH has a critical role in cell entry and, potentially, pathogenesis.
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