Porcine MKRN1 Modulates the Replication and Pathogenesis of Porcine Circovirus Type 2 by Inducing Capsid Protein Ubiquitination and Degradation
Porcine MKRN1 Modulates the Replication and Pathogenesis of Porcine Circovirus Type 2 by Inducing Capsid Protein Ubiquitination and Degradation
复制标题
猪 MKRN1 通过诱导衣壳蛋白泛素化和降解来调节猪圆环病毒 2 型的复制和发病机制
DOI：
10.1128/jvi.00100-18
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发表时间：
2018-06-01
影响因子：
5.4
通讯作者：
Huang, Yong
中科院分区：
文献类型：
--
作者：
Wang, Tongtong;Du, Qian;Huang, Yong
猪圆环病毒2型（Porcine circovirus type 2，PCV 2）衣壳蛋白（Capsidprotein，Cap）是一种独特的结构蛋白，在病毒的复制和致病过程中起着关键作用。在此，我们的特点是一个假定的猪Makorin环指蛋白1（pMKRN 1）的变体，一个N-末端截短的变体假定的全尺寸猪MKRN 1具有独特的表达模式，从猪mkrn 1基因，并与PCV 2帽相互作用。结构域作图分析表明，这种相互作用需要pMKRN 1的C末端和Cap的片段（氨基酸108至198）。PCV 2瞬时上调PK-15细胞中的pMKRN 1，但持续病毒感染下调PCV 2感染仔猪的主要病理组织中的pMKRN 1。过表达pMKRN 1可通过泛素化和Cap降解显著抑制PCV 2子代的产生，而敲除pMKRN 1可阻断Cap降解并促进子代病毒的复制。pMKRN 1特异性靶向PCV 2 Cap赖氨酸残基164、179和191，以诱导多聚泛素化和随后的降解。Cap蛋白中三个赖氨酸残基的突变或pMKRN 1的RING指结构域内残基243处组氨酸的突变废除了pMKRN 1的E3连接酶活性，使得细胞不能诱导Cap泛素化和降解。与这一发现相一致，Cap泛素化缺陷型PCV 2株与野生型PCV 2相比，显示出增强的病毒复制，并在肺和淋巴结组织中产生严重的组织学病变。综上所述，本研究结果表明，PCV 2下调pMKRN 1变异体以避免pMKRN 1介导的Cap泛素化和降解，从而促进病毒在其靶组织中的复制和致病。重要性猪圆环病毒2型是猪最易感的病原体，给全球养猪业造成巨大的经济损失，但宿主细胞是否已经发展出一些策略来阻止病毒复制仍不清楚。在此，我们发现猪MKRN 1（pMKRN 1）在PCV 2感染的早期阶段上调，并介导Cap蛋白的多聚泛素化和降解以阻断PCV 2复制，而持续的PCV 2感染下调pMKRN 1水平以避免降解，促进病毒在其靶组织中的复制和致病。这些数据为pMKRN 1 E3连接酶在PCV 2感染期间的抗病毒作用的分子机制提供了新的见解，也为PCV 2爆发提出了潜在的新控制措施。
Porcine circovirus type 2 (PCV2) capsid protein (Cap) is a unique structure protein that plays pivotal roles in the process of viral replication and pathogenesis. Herein, we characterized a putative porcine Makorin RING finger protein 1 (pMKRN1) variant, an N-terminal-truncated variant of putative full-size porcine MKRN1 which has a unique expression pattern resulting from the porcine mkrn1 gene and which interacts with PCV2 Cap. A domain mapping assay showed that the C terminus of pMKRN1 and fragments (amino acids 108 to 198) of Cap are required for this interaction. PCV2 transiently upregulated pMKRN1 in PK-15 cells, but persistent viral infection downregulated pMKRN1 in major pathological tissues of PCV2-infected piglets. Overexpression of pMKRN1 significantly inhibited the generation of progeny PCV2 via ubiquitination and degradation of Cap, whereas knockout of pMKRN1 blocked Cap degradation and promoted progeny virus replication. pMKRN1 specifically targeted PCV2 Cap lysine residues 164, 179, and 191 to induce polyubiquitination and subsequent degradation. Mutation of either of the three lysine residues in the Cap protein or mutation of the histidine at residue 243 within the RING finger domain of pMKRN1 abrogated the E3 ligase activity of pMKRN1, rendering cells incapable of inducing Cap ubiquitination and degradation. Consistent with this finding, a Cap ubiquitination-deficient PCV2 strain showed enhanced virus replication and produced severe histological lesions in the lung and lymph node tissues compared with wild-type PCV2. Taken together, the results presented here suggest that PCV2 downregulates the pMKRN1 variant to avoid pMKRN1-mediated Cap ubiquitination and degradation, thus promoting viral replication and pathogenesis in its targeted tissues.IMPORTANCE Porcine circovirus type 2 is the pathogen to which pigs are the most susceptible, causing immense economic losses in the global swine industry, but whether host cells have developed some strategies to prevent viral replication is still unclear. Here, we found that porcine MKRN1 (pMKRN1) was upregulated in the early stage of PCV2 infection and mediated the polyubiquitination and degradation of Cap protein to block PCV2 replication, yet persistent PCV2 infection downregulated pMKRN1 levels to avoid degradation, promoting viral replication and pathogenesis in its targeted tissues. These data present new insight into the molecular mechanisms underlying the antiviral effects of pMKRN1 E3 ligase during PCV2 infection and also suggest potential new control measures for PCV2 outbreaks.