Identification of functionally important domains of human cytomegalovirus gO that act after trimer binding to receptors.
Identification of functionally important domains of human cytomegalovirus gO that act after trimer binding to receptors.
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鉴定人类巨细胞病毒的功能重要结构域在三聚体与受体结合后起作用。
DOI：
10.1371/journal.ppat.1010452
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发表时间：
2022-04
期刊：
影响因子：
6.7
通讯作者：
中科院分区：
文献类型：
--
作者：
人巨细胞病毒（HCMV）的进入涉及三聚体（gH/gL/gO）与成纤维细胞中的PDGFRα相互作用。进入上皮细胞和内皮细胞需要结合未知受体的三聚体和结合neuropilin-2的五聚体（gH/gL/UL128-131）。为了确定三聚体中功能重要的结构域，我们筛选了一个重叠的20-mer氧化棕黄烯肽文库，并鉴定了两组可以阻止病毒进入的肽：19/20 （a.a.235 - 267）和32/33 （a.a.404 - 436）。含有野生型gO的可溶性三聚体阻断HCMV的进入，而含有19/20或32/33序列突变的可溶性三聚体不阻断HCMV的进入。有趣的是，突变三聚体保留了与包括PDGFRα在内的细胞受体结合的能力。肽19/20和32/33序列分别形成从氧化石墨烯表面延伸的叶状结构和相邻的凹状结构。这两组序列均未与PDGFRα接触。相反，我们的数据支持一个模型，其中19/20和32/33三聚体序列在受体结合的下游起作用，例如将HCMV运输到核内体或与gB结合以进入融合。我们还在人血清中筛选了与抗体结合的肽，发现肽20和肽26与抗体结合，这些肽在兔免疫后产生中和抗体（nab），并能从人血清中提取nab。肽20和26序列是第一个三聚体鉴定的NAb表位。这些研究描述了氧化石墨烯上的两个重要表面：i)肽19/20和32/33，它们明显作用于受体结合的下游；ii)肽26，与PDGFRα相互作用。这两个表面都是nab的靶标。人类巨细胞病毒（HCMV）感染了世界上80%的人口，在移植患者中引起严重的发病率和死亡率，并可传播给发育中的胎儿，导致严重的神经缺陷。目前用于治疗HCMV的抗病毒药物不是很有效，因为病毒可能产生耐药性，而且目前还没有获得许可的HCMV疫苗。最近，人们对HCMV包膜糖蛋白作为疫苗成分参与进入的研究产生了浓厚的兴趣。一种糖蛋白复合物，gH/gL/gO三聚体特别有趣，因为它是所有细胞类型的细胞外病毒感染所必需的。在这里，我们确定了三聚体中的结构域，这些结构域在受体结合的进入下游具有重要功能，也是自然诱导的中和抗体识别的表位。这些结果将对推进开发新型HCMV疗法的努力产生影响。
Human cytomegalovirus (HCMV) entry involves trimer (gH/gL/gO) that interacts with PDGFRα in fibroblasts. Entry into epithelial and endothelial cells requires trimer, which binds unidentified receptors, and pentamer (gH/gL/UL128-131), which binds neuropilin-2. To identify functionally important domains in trimer, we screened an overlapping 20-mer gO peptide library and identified two sets of peptides: 19/20 (a.a. 235–267) and 32/33 (a.a. 404–436) that could block virus entry. Soluble trimer containing wild type gO blocked HCMV entry, whereas soluble trimers with the 19/20 or 32/33 sequences mutated did not block entry. Interestingly, the mutant trimers retained the capacity to bind to cellular receptors including PDGFRα. Peptide 19/20 and 32/33 sequences formed a lobe extending from the surface of gO and an adjacent concave structure, respectively. Neither of these sets of sequences contacted PDGFRα. Instead, our data support a model in which the 19/20 and 32/33 trimer sequences function downstream of receptor binding, e.g. trafficking of HCMV into endosomes or binding to gB for entry fusion. We also screened for peptides that bound antibodies (Abs) in human sera, observing that peptides 20 and 26 bound Abs. These peptides engendered neutralizing Abs (NAbs) after immunization of rabbits and could pull out NAbs from human sera. Peptides 20 and 26 sequences represent the first NAb epitopes identified in trimer. These studies describe two important surfaces on gO defined by: i) peptides 19/20 and 32/33, which apparently act downstream of receptor binding and ii) peptide 26 that interacts with PDGFRα. Both these surfaces are targets of NAbs. Human cytomegalovirus (HCMV) infects 80% of the world population, causing severe morbidity and mortality in transplant patients and can be transmitted to the developing fetus leading to severe neurological defects. The current anti-viral agents used to treat HCMV are not very effective as viruses can develop resistance and there is no licensed HCMV vaccine available. Recently, there has been intense interest in the HCMV envelope glycoproteins involved in entry as a component of vaccines. One glycoprotein complex, the gH/gL/gO trimer is especially intriguing as it is required for infection of extracellular virus in all cell types. Here, we identify domains in the trimer that have an essential function in entry downstream of receptor binding and are also epitopes recognized by naturally induced neutralizing antibodies. These results will have implications for advancing the efforts to develop novel HCMV therapeutics.
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影响因子：
5.4
作者：
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通讯作者：
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Perez L
影响因子：
3.8
作者：
Dolan, A;Cunningham, C;Davison, AJ
通讯作者：
Davison, AJ