Analysis of the function of hPIV2 V protein by using reverse genetics.

利用反向遗传学分析hPIV2 V蛋白的功能。

• 批准号: 15590416
• 负责人: NISHIO Machiko
• 金额: $ 2.3万
• 依托单位: Mie University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15590416/
• 关键词: hPIV2; the V protein; anti-IFN activity; virus growth rate; Cysteine-rich-domain; Trptophan-rich-motif; STAT; recombinant virus; 抗インターフェロン活性; Tet-on system; hPIV4; レコンビナントウイルス; Tryptophan-rich-motif; cysteine-rich-domain; STAT蛋白; hPIV2; the V protein; anti-IFN activity; virus growth rate; Cysteine-rich-domain; Trptophan-rich-motif; STAT; recombinant virus; 抗インターフェロン活性; Tet-on system; hPIV4; レコンビナントウイルス; Tryptophan-rich-motif; cysteine-rich-domain; STAT蛋白

1.The V protein of human parainfluenza virus type 2 (hPIV2) inhibits interferon (IFN) -induced innate antiviral responses through STAT protein degradation. I identified hPIV2 V protein residues essential for STAT2 protein degradation in human cells, that are conserved seven cysteine residues, tryptophan-rich-motif, and aa 207 phenylalanine in the C-terminal V-unique domain, and aa 143 phenylalanine in the P/V common domain.2.To conform the function for virus growth of the V protein, I made the recombinant virus which completely lacks expression of the V protein by inactivating the P gene mRNA-editing signal (rPIV2/P-edit). The growth rate of rPIV2/P-edit is very limited even in the Vero cells that cannot induce IFN. Thus, these results suggest that the V protein is essential not only for STAT protein degradation but also for promoting virus growth. The rhPIV2s which have the mutation in the V-specific domain also grew lower, but rPIV2 which has the mutation in the P/V common domain grew similar to wt.3.I investigated whether the V protein of human parainfluenza virus type 4 (hPIV4) has the function to evade the IFN-induced antiviral responses. The hPIV4 V protein has the conserved seven cysteine residues and tryptophan-rich-motif, and can bind to DDB1 and Cul4A that are important for STAT protein degradation. However, the hPIV4 V protein has no function to inhibit IFN signaling. I show that the only paramyxovirus that can't evade the IFN-induced antiviral responses to data is hPIV4.4.I identified that the V-specific region of hPIV2 binds to the N-terminal 80 amino acids on the NP protein, and is critical for self-association. Furthermore, I have identified a host protein, AIP1/Alix, involved in apoptosis and efficient budding of several enveloped viruses, as an interacting partner of the V and NP proteins. My data also suggest that the transiently binding between V and AIP1 is important for virus growth.

1.人副帕氏菌病毒2型（HPIV2）的V蛋白抑制干扰素（IFN）通过Stat蛋白质降解诱导的先天抗病毒药反应。我确定了人类细胞中STAT2蛋白降解必不可少的HPIV2 V蛋白残基，这些残基是保守的七个半胱氨酸残基，富含色氨酸的含量和AA 207苯丙氨酸的C-末端V-Unique结构域中的AA 207苯丙氨酸，而AA 143在P/V普通域中aa 143苯烷氨基烷。通过灭活P基因mRNA编辑信号（RPIV2/p-edit）来缺乏V蛋白的表达。即使在无法诱导IFN的Vero细胞中，RPIV2/P-EDIT的生长速率也非常有限。因此，这些结果表明，V蛋白不仅对于Stat蛋白质降解，而且对于促进病毒生长至关重要。在V特异性结构域中具有突变的RHPIV2也越来越低，但是在P/V公共结构域中突变的RPIV2与WT.3.I相似。3.i研究了人帕氏菌病毒4（HPIV4）的V蛋白（HPIV4）是否具有逃避IFN IFN诱导的抗病毒反应的功能。 HPIV4 V蛋白具有保守的七个半胱氨酸残基和富含色氨酸的含量，并且可以与DDB1和CUL4A结合，这对于Stat蛋白质降解很重要。但是，HPIV4 V蛋白没有抑制IFN信号传导的功能。我表明，唯一无法逃避IFN诱导的抗病毒反应的帕托马病毒对数据的反应是HPIV4.4.I确定HPIV2的V特异性区域与NP蛋白上的N末端80氨基酸结合，并且对于自我相关至关重要。此外，我已经确定了一种宿主蛋白​​AIP1/ALIX，它参与了几种包膜病毒的凋亡和有效的萌芽，是V和NP蛋白的相互作用伴侣。我的数据还表明，V和AIP1之间的瞬时结合对于病毒生长很重要。