Molecular Determinants of Proteolytic Disassembly of the Reovirus Outer Capsid
Molecular Determinants of Proteolytic Disassembly of the Reovirus Outer Capsid
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DOI：
10.1074/jbc.m111.334854
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发表时间：
2012-03-09
影响因子：
4.8
通讯作者：
Dermody, Terence S.
中科院分区：
文献类型：
--
作者：
Doyle, Joshua D.;Danthi, Pranav;Dermody, Terence S.
在附着和内化之后，哺乳动物呼肠孤病毒经历细胞内蛋白水解分解，然后渗透到细胞质中。呼肠孤病毒解体的起始事件是组织蛋白酶介导的病毒外衣壳蛋白sigma 3的蛋白水解降解。菌株3型Dearing (T3D) sigma 3的氨基酸354 （Y354H）上的单个酪氨酸-组氨酸突变增强了呼肠孤病毒的分解，并赋予其对蛋白酶抑制剂（如E64）的抗性。菌株3 Abney （T3A）与T3D的sigma 3氨基酸序列在Y354H等8个位置存在差异。然而，T3A表现出与T3D相当的拆卸动力学和蛋白酶敏感性。我们假设一个或多个额外的sigma 3多态性抑制Y354H表型并恢复T3D拆卸特性。为了验证这一假设，我们设计了一组具有T3A sigma 3多态性的呼肠孤病毒变体，将其单独引入T3D-sigma 3Y354H。我们评估了这些病毒对E64的抗性和体外组织蛋白酶L的敏感性，发现在198位含有甘氨酸到谷氨酸取代的病毒（G198E）表现出与T3A和T3D相似的分解动力学和E64敏感性。此外，含有233位和347位（S233L和I347T）变化的病毒在198位发生了新的补补性突变，这加强了残基198是sigma 3蛋白水解易感性的关键决定因素的结论。在热处理后，sigma 3中携带Y354H的变异比T3A或T3D更快地丧失感染性，G198E消除了这一效应。这些结果确定了控制sigma 3切割和衣壳稳定性的残基调控网络，从而为非包膜病毒分解的调控提供了见解。
Following attachment and internalization, mammalian reoviruses undergo intracellular proteolytic disassembly followed by viral penetration into the cytoplasm. The initiating event in reovirus disassembly is the cathepsin-mediated proteolytic degradation of viral outer capsid protein sigma 3. A single tyrosine-to-histidine mutation at amino acid 354 (Y354H) of strain type 3 Dearing (T3D) sigma 3 enhances reovirus disassembly and confers resistance to protease inhibitors such as E64. The sigma 3 amino acid sequence of strain type 3 Abney (T3A) differs from that of T3D at eight positions including Y354H. However, T3A displays disassembly kinetics and protease sensitivity comparable with T3D. We hypothesize that one or more additional sigma 3 polymorphisms suppress the Y354H phenotype and restore T3D disassembly characteristics. To test this hypothesis, we engineered a panel of reovirus variants with T3A sigma 3 polymorphisms introduced individually into T3D-sigma 3Y354H. We evaluated E64 resistance and in vitro cathepsin L susceptibility of these viruses and found that one containing a glycine-to-glutamate substitution at position 198 (G198E) displayed disassembly kinetics and E64 sensitivity similar to those properties of T3A and T3D. Additionally, viruses containing changes at positions 233 and 347 (S233L and I347T) developed de novo compensatory mutations at position 198, strengthening the conclusion that residue 198 is a key determinant of sigma 3 proteolytic susceptibility. Variants with Y354H in sigma 3 lost infectivity more rapidly than T3A or T3D following heat treatment, an effect abrogated by G198E. These results identify a regulatory network of residues that control sigma 3 cleavage and capsid stability, thus providing insight into the regulation of nonenveloped virus disassembly.