Bimolecular Complementation Defines Functional Regions of Herpes Simplex Virus gB That Are Involved with gH/gL as a Necessary Step Leading to Cell Fusion
Bimolecular Complementation Defines Functional Regions of Herpes Simplex Virus gB That Are Involved with gH/gL as a Necessary Step Leading to Cell Fusion
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DOI：
10.1128/jvi.02687-09
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发表时间：
2010-04-01
影响因子：
5.4
通讯作者：
Eisenberg, Roselyn J.
中科院分区：
文献类型：
--
作者：
Atanasiu, Doina;Whitbeck, J. Charles;Eisenberg, Roselyn J.
单纯疱疹病毒（HSV）进入细胞需要四种膜糖蛋白：gD是受体结合蛋白，gB和gH/gL构成核心融合机制。 gD 及其受体的晶体结构为理解最初的触发步骤提供了基础，但核心融合蛋白如何发挥作用仍然未知。 gB晶体结构表明它是III类融合蛋白，但与其他类成员不同，gB本身不会引起融合。双分子互补 (BiMC) 研究表明，gD 受体结合会触发 gB 和 gH/gL 之间的相互作用，同时触发融合。没有答案的是 BiMC 是否会导致聚变，还是聚变的副产品。我们使用 gB 单克隆抗体 (MAb) 来阻断这些事件的不同方面。针对 gB 的非病毒中和性 MAb 未能阻断 BiMC 或融合。相比之下，中和病毒的 gB 单克隆抗体则阻断融合。这些 MAb 映射到 gB 的三个功能区 (FR)。 FR1 的单克隆抗体含有融合环，而 FR2 则阻断 BiMC 和融合。相比之下，针对 FR3（参与受体结合的区域）的单克隆抗体会阻断融合，但不会阻断 BiMC。因此，FR3 MAb 将 BiMC 相互作用与融合分开，表明 BiMC 发生在融合之前。当取代野生型 (wt) gB 时，融合环突变体阻断融合和 BiMC，表明环插入先于 BiMC。因此，我们假设每个 gB FR 都参与导致融合的路径的不同方面。在 gD 触发后，gB 融合环被插入目标脂质膜中。然后 gB 与 gH/gL 相互作用，这种相互作用最终导致融合。
Herpes simplex virus (HSV) entry into cells requires four membrane glycoproteins: gD is the receptor binding protein, and gB and gH/gL constitute the core fusion machinery. Crystal structures of gD and its receptors have provided a basis for understanding the initial triggering steps, but how the core fusion proteins function remains unknown. The gB crystal structure shows that it is a class III fusion protein, yet unlike other class members, gB itself does not cause fusion. Bimolecular complementation (BiMC) studies have shown that gD-receptor binding triggers an interaction between gB and gH/gL and concurrently triggers fusion. Left unanswered was whether BiMC led to fusion or was a by-product of it. We used gB monoclonal antibodies (MAbs) to block different aspects of these events. Non-virus-neutralizing MAbs to gB failed to block BiMC or fusion. In contrast, gB MAbs that neutralize virus blocked fusion. These MAbs map to three functional regions (FR) of gB. MAbs to FR1, which contains the fusion loops, and FR2 blocked both BiMC and fusion. In contrast, MAbs to FR3, a region involved in receptor binding, blocked fusion but not BiMC. Thus, FR3 MAbs separate the BiMC interaction from fusion, suggesting that BiMC occurs prior to fusion. When substituted for wild-type (wt) gB, fusion loop mutants blocked fusion and BiMC, suggesting that loop insertion precedes BiMC. Thus, we postulate that each of the gB FRs are involved in different aspects of the path leading to fusion. Upon triggering by gD, gB fusion loops are inserted into target lipid membranes. gB then interacts with gH/gL, and this interaction is eventually followed by fusion.