Cell Attachment Domains of the Porcine Epidemic Diarrhea Virus Spike Protein Are Key Targets of Neutralizing Antibodies
Cell Attachment Domains of the Porcine Epidemic Diarrhea Virus Spike Protein Are Key Targets of Neutralizing Antibodies
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DOI：
10.1128/jvi.00273-17
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发表时间：
2017-06-01
影响因子：
5.4
通讯作者：
Bosch, Berend-Jan
中科院分区：
文献类型：
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作者：
Li, Chunhua;Li, Wentao;Bosch, Berend-Jan
猪流行性腹泻病毒（Porcine epidemic diarrhea virus，PEDV）是一种引起猪肠道疾病的病毒，在世界范围内给养猪业造成了巨大的经济损失。目前针对这种新出现的冠状病毒的疫苗接种方法仅部分有效，尽管自然感染可保护猪免受再感染，并为哺乳仔猪提供催乳免疫。负责受体结合和细胞进入的病毒刺突（S）糖蛋白是中和抗体的主要靶标。然而，抗体表位的知识，其性质和位置的刺突结构，以及抗体干扰感染的机制是稀缺的。在这里，我们描述了10个中和和非中和小鼠单克隆抗体的产生和表征提出对S蛋白的S1受体结合亚基。通过表达不同的S1蛋白片段，鉴定了分布在S1亚基的5个结构域上的6个抗体表位类别。抗体的交叉反应性和交叉中和的表征揭示了PEDV毒株之间的抗原差异。有效中和抗体的表位分离成两个表位类别，并映射在N-末端唾液酸结合结构域和更C-末端受体结合结构域中。抗体中和逃逸突变体显示出单个氨基酸取代，削弱了抗体结合和中和作用，并定义了表位的位置。我们的观察图片的PEDV S1亚基的抗体表位景观，并揭示其细胞附着域的中和antibody.Importance的关键目标猪流行性腹泻病毒（PEDV），一种新兴的猪冠状病毒，导致经济上重要的肠道疾病的猪。目前缺乏用于疾病控制的有效PEDV疫苗。病毒体表面的刺突（S）糖蛋白是病毒进入细胞的关键因素，因此也是中和抗体的主要靶标。为了了解PEDV刺突蛋白的抗原性，我们开发了针对刺突蛋白的S1受体结合区的单克隆抗体，并表征了它们的表位、中和活性和对多种PEDV毒株的交叉反应性。抗体的表位分离成分散在多结构域S1结构上的六个表位类别。单克隆抗体揭示了PEDV毒株之间B细胞表位的抗原变异性。中和抗体的表位映射到S1中的两个不同的结构域，其分别参与与碳水化合物和蛋白质细胞表面分子的结合，表明PEDV刺突蛋白上的这些细胞附着位点在引发保护性体液免疫应答中的重要性。
Porcine epidemic diarrhea virus (PEDV) causes enteric disease in pigs, resulting in significant economic losses to the swine industry worldwide. Current vaccination approaches against this emerging coronavirus are only partially effective, though natural infection protects pigs against reinfection and provides lactogenic immunity to suckling piglets. The viral spike (S) glycoprotein, responsible for receptor binding and cell entry, is the major target for neutralizing antibodies. However, knowledge of antibody epitopes, their nature and location in the spike structure, and the mechanisms by which the antibodies interfere with infection is scarce. Here we describe the generation and characterization of 10 neutralizing and nonneutralizing mouse monoclonal antibodies raised against the S1 receptor binding subunit of the S protein. By expression of different S1 protein fragments, six antibody epitope classes distributed over the five structural domains of the S1 subunit were identified. Characterization of antibodies for cross-reactivity and cross-neutralization revealed antigenic differences among PEDV strains. The epitopes of potent neutralizing antibodies segregated into two epitope classes and mapped within the N-terminal sialic acid binding domain and in the more C-terminal receptor binding domain. Antibody neutralization escape mutants displayed single amino acid substitutions that impaired antibody binding and neutralization and defined the locations of the epitopes. Our observations picture the antibody epitope landscape of the PEDV S1 subunit and reveal that its cell attachment domains are key targets of neutralizing antibodies.IMPORTANCE Porcine epidemic diarrhea virus (PEDV), an emerging porcine coronavirus, causes an economically important enteric disease in pigs. Effective PEDV vaccines for disease control are currently lacking. The spike (S) glycoprotein on the virion surface is the key player in virus cell entry and, therefore, the main target of neutralizing antibodies. To understand the antigenic landscape of the PEDV spike protein, we developed monoclonal antibodies against the spike protein's S1 receptor binding region and characterized their epitopes, neutralizing activity, and cross-reactivity toward multiple PEDV strains. Epitopes of antibodies segregated into six epitope classes dispersed over the multidomain S1 structure. Monoclonal antibodies revealed antigenic variability in B-cell epitopes between PEDV strains. The epitopes of neutralizing antibodies mapped to two distinct domains in S1 that are involved in binding to carbohydrate and proteinaceous cell surface molecules, respectively, indicating the importance of these cell attachment sites on the PEDV spike protein in eliciting a protective humoral immune response.