Host Proteolytic Activity Is Necessary for Infectious Bursal Disease Virus Capsid Protein Assembly
Host Proteolytic Activity Is Necessary for Infectious Bursal Disease Virus Capsid Protein Assembly
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DOI：
10.1074/jbc.m112.356113
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发表时间：
2012-07-13
影响因子：
4.8
通讯作者：
Rodriguez, Jose F.
中科院分区：
文献类型：
--
作者：
Irigoyen, Nerea;Caston, Jose R.;Rodriguez, Jose F.
在许多病毒中，前体颗粒或原衣壳被组装并经历大量的化学和物理修饰以产生感染性衣壳。衣壳组装和成熟是病毒和宿主因子参与的精细调节过程。我们表明，前体的VP 2衣壳蛋白（pVP 2）的传染性法氏囊病病毒（IBDV），一种双链RNA病毒，在C-末端结构域（CTD）由宿主蛋白酶，嘌呤霉素敏感的氨肽酶（PurSA）处理。pVP 2 CTD（71个残基）在决定构建T = 13复合衣壳的VP 2（441个残基）的各种构象中具有重要作用。pVP 2 CTD活性由VP 4病毒蛋白酶和VP 2本身对不同靶点的共翻译和翻译后蛋白水解修饰控制，以产生成熟的VP 2 -441种类。嘌呤霉素敏感性氨肽酶负责切割Arg-452-Arg-453键以产生中间体pVP 2 -452多肽的肽酶活性。pVP 2 R453 A取代消除PurSA活性。我们使用基于杆状病毒的系统在昆虫细胞中表达IBDV多蛋白，并发现与IBDV病毒体相似的病毒样颗粒的形成效率低下，这与这些细胞中缺乏嘌呤霉素敏感的氨肽酶有关。然而，通过共表达鸡PurSA或pVP 2 -452蛋白，有效地挽救了病毒样颗粒组装。在允许IBDV复制的细胞系中沉默或药理学抑制嘌呤霉素敏感性氨肽酶活性导致感染性IBDV颗粒的组装和/或成熟的主要阻断，因为病毒产量显著降低。因此PurSA活性对于IBDV复制是必需的。
In many viruses, a precursor particle, or procapsid, is assembled and undergoes massive chemical and physical modification to produce the infectious capsid. Capsid assembly and maturation are finely tuned processes in which viral and host factors participate. We show that the precursor of the VP2 capsid protein (pVP2) of the infectious bursal disease virus (IBDV), a double-stranded RNA virus, is processed at the C-terminal domain (CTD) by a host protease, the puromycin-sensitive aminopeptidase (PurSA). The pVP2 CTD (71 residues) has an important role in determining the various conformations of VP2 (441 residues) that build the T = 13 complex capsid. pVP2 CTD activity is controlled by co- and posttranslational proteolytic modifications of different targets by the VP4 viral protease and by VP2 itself to yield the mature VP2-441 species. Puromycin-sensitive aminopeptidase is responsible for the peptidase activity that cleaves the Arg-452-Arg-453 bond to generate the intermediate pVP2-452 polypeptide. A pVP2 R453A substitution abrogates PurSA activity. We used a baculovirus-based system to express the IBDV polyprotein in insect cells and found inefficient formation of virus-like particles similar to IBDV virions, which correlates with the absence of puromycin-sensitive aminopeptidase in these cells. Virus-like particle assembly was nonetheless rescued efficiently by coexpression of chicken PurSA or pVP2-452 protein. Silencing or pharmacological inhibition of puromycin-sensitive aminopeptidase activity in cell lines permissive for IBDV replication caused a major blockade in assembly and/or maturation of infectious IBDV particles, as virus yields were reduced markedly. PurSA activity is thus essential for IBDV replication.