Silva Laurie A, Khomandiak Solomiia, Ashbrook Alison W, Weller Romy, Heise Mark T, Morrison Thomas E, Dermody Terence S
Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, Tennessee, USA.
J Virol. 2014 Mar;88(5):2385-97. doi: 10.1128/JVI.03116-13. Epub 2013 Dec 26.
Chikungunya virus (CHIKV) is a reemerging arbovirus responsible for outbreaks of infection throughout Asia and Africa, causing an acute illness characterized by fever, rash, and polyarthralgia. Although CHIKV infects a broad range of host cells, little is known about how CHIKV binds and gains access to the target cell interior. In this study, we tested whether glycosaminoglycan (GAG) binding is required for efficient CHIKV replication using CHIKV vaccine strain 181/25 and clinical isolate SL15649. Preincubation of strain 181/25, but not SL15649, with soluble GAGs resulted in dose-dependent inhibition of infection. While parental Chinese hamster ovary (CHO) cells are permissive for both strains, neither strain efficiently bound to or infected mutant CHO cells devoid of GAG expression. Although GAGs appear to be required for efficient binding of both strains, they exhibit differential requirements for GAGs, as SL15649 readily infected cells that express excess chondroitin sulfate but that are devoid of heparan sulfate, whereas 181/25 did not. We generated a panel of 181/25 and SL15649 variants containing reciprocal amino acid substitutions at positions 82 and 318 in the E2 glycoprotein. Reciprocal exchange at residue 82 resulted in a phenotype switch; Gly(82) results in efficient infection of mutant CHO cells but a decrease in heparin binding, whereas Arg(82) results in reduced infectivity of mutant cells and an increase in heparin binding. These results suggest that E2 residue 82 is a primary determinant of GAG utilization, which likely mediates attenuation of vaccine strain 181/25.
Chikungunya virus (CHIKV) infection causes a debilitating rheumatic disease that can persist for months to years, and yet there are no licensed vaccines or antiviral therapies. Like other alphaviruses, CHIKV displays broad tissue tropism, which is thought to be influenced by virus-receptor interactions. In this study, we determined that cell-surface glycosaminoglycans are utilized by both a vaccine strain and a clinical isolate of CHIKV to mediate virus binding. We also identified an amino acid polymorphism in the viral E2 attachment protein that influences utilization of glycosaminoglycans. These data enhance an understanding of the viral and host determinants of CHIKV cell entry, which may foster development of new antivirals that act by blocking this key step in viral infection.
基孔肯雅病毒（CHIKV）是一种再次出现的虫媒病毒，在亚洲和非洲引发感染疫情，导致以发热、皮疹和多关节痛为特征的急性疾病。尽管CHIKV可感染多种宿主细胞，但对于CHIKV如何结合并进入靶细胞内部知之甚少。在本研究中，我们使用CHIKV疫苗株181/25和临床分离株SL15649测试了高效CHIKV复制是否需要糖胺聚糖（GAG）结合。用可溶性GAG对181/25株（而非SL15649株）进行预孵育，导致感染呈剂量依赖性抑制。虽然亲本中国仓鼠卵巢（CHO）细胞对这两种毒株均易感，但两种毒株均不能有效结合或感染缺乏GAG表达的突变CHO细胞。尽管GAG似乎是两种毒株有效结合所必需的，但它们对GAG的需求存在差异，因为SL15649能轻易感染表达过量硫酸软骨素但缺乏硫酸乙酰肝素的细胞，而181/25则不能。我们构建了一组181/25和SL15649变体，这些变体在E2糖蛋白的第82位和318位含有相互的氨基酸替换。在第82位残基处进行相互交换导致了表型转换；甘氨酸（82）导致对突变CHO细胞的有效感染，但肝素结合减少，而精氨酸（82）导致突变细胞的感染性降低和肝素结合增加。这些结果表明，E2残基82是GAG利用的主要决定因素，这可能介导了疫苗株181/25的减毒。
基孔肯雅病毒（CHIKV）感染会导致一种使人衰弱的风湿性疾病，可持续数月至数年，然而目前尚无获批的疫苗或抗病毒疗法。与其他甲病毒一样，CHIKV表现出广泛的组织嗜性，这被认为受病毒-受体相互作用的影响。在本研究中，我们确定CHIKV的疫苗株和临床分离株均利用细胞表面糖胺聚糖来介导病毒结合。我们还在病毒E2附着蛋白中鉴定出一种氨基酸多态性，其影响糖胺聚糖的利用。这些数据增进了对CHIKV细胞进入的病毒和宿主决定因素的理解，这可能有助于开发通过阻断病毒感染这一关键步骤起作用的新型抗病毒药物。
