Canine Distemper Virus Fusion Activation: Critical Role of Residue E123 of CD150/SLAM
Canine Distemper Virus Fusion Activation: Critical Role of Residue E123 of CD150/SLAM
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DOI：
10.1128/jvi.02405-15
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发表时间：
2016-02-01
影响因子：
5.4
通讯作者：
Plattet, Philippe
中科院分区：
文献类型：
--
作者：
Khosravi, Mojtaba;Bringolf, Fanny;Plattet, Philippe
麻疹病毒（MeV）和犬瘟热病毒（CDV）具有四聚体附着蛋白（H）和三聚体融合蛋白，其与SLAM或nectin 4受体合作以触发膜融合以进入细胞。虽然MeV H-SLAM共晶结构揭示了结合界面，但也确定了两种不同的低聚H组装体。在一种构象中，两个SLAM单元夹在两个离散的H头结构域之间，从而突出两个结合界面（“前”和“后”）。在这里，我们研究了激活CDV膜融合机制的两个接口的功能相关性。虽然丙氨酸扫描诱变在SLAM的前H-结合位点中鉴定了五个关键的调节残基，但保守的谷氨酸残基（位置123处的E，被A [E123 A]取代）的取代导致对融合促进的最显著影响。有趣的是，虽然使用可溶性构建体测定H与受体的相互作用揭示了所鉴定的SLAM突变体的结合减少，但当用两种分子的全长对应物研究物理相互作用时，没有记录到任何影响。相反，尽管SLAM的后H结合位点内的三个策略性选择的残基的诱变基本上不影响融合触发，然而，突变体减弱了用膜锚定蛋白构建体记录的H-SLAM相互作用。总的来说，我们的研究结果支持的模式之间的连接蛋白和V结构域的SLAM是常见的所有麻疹病毒，并建议的SLAM残基E123，位于前面的H-结合位点，在触发融合机制的主要作用。然而，我们的数据还支持这一假设，即可能需要两种糖蛋白的其他微结构域（包括后H-结合位点）来实现完全有效的H-SLAM相互作用。
Measles virus (MeV) and canine distemper virus (CDV) possess tetrameric attachment proteins (H) and trimeric fusion proteins, which cooperate with either SLAM or nectin 4 receptors to trigger membrane fusion for cell entry. While the MeV H-SLAM co-crystal structure revealed the binding interface, two distinct oligomeric H assemblies were also determined. In one of the conformations, two SLAM units were sandwiched between two discrete H head domains, thus spotlighting two binding interfaces ("front" and "back"). Here, we investigated the functional relevance of both interfaces in activating the CDV membrane fusion machinery. While alanine-scanning mutagenesis identified five critical regulatory residues in the front H-binding site of SLAM, the replacement of a conserved glutamate residue (E at position 123, replaced with A [E123A]) led to the most pronounced impact on fusion promotion. Intriguingly, while determination of the interaction of H with the receptor using soluble constructs revealed reduced binding for the identified SLAM mutants, no effect was recorded when physical interaction was investigated with the full-length counterparts of both molecules. Conversely, although mutagenesis of three strategically selected residues within the back H-binding site of SLAM did not substantially affect fusion triggering, nevertheless, the mutants weakened the H-SLAM interaction recorded with the membrane-anchored protein constructs. Collectively, our findings support a mode of binding between the attachment protein and the V domain of SLAM that is common to all morbilliviruses and suggest a major role of the SLAM residue E123, located at the front H-binding site, in triggering the fusion machinery. However, our data additionally support the hypothesis that other microdomain(s) of both glycoproteins (including the back H-binding site) might be required to achieve fully productive H-SLAM interactions.