Analysis of the function of paramyxovaus V protein

副粘液V蛋白的功能分析

基本信息

批准号：
18590448
负责人：
NISHIO Machiko
金额：
$ 2.55万
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：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18590448/
关键词：
hPIV2 V protein virus growth recombinant virus L protein virus RNA replication Yeast two-hybrid shRNA 抗インターフェロン活性

项目摘要

1. I identified a host protein, AIPI/Alix, involved in apoptosis and efficient budding of several enveloped viruses as an interacting partner of hPIV2 NP and V proteins. To investigate possible involvement of AIPI in hPIV2 virus growth, I established cell lines in which were depleted the intracellular AIP 1 by using short hairpin RNA induced by adding Dox. The wt hPIV2 growth in the established cells was inhibited by 10- to 100-fold, but the growth rates of rPIV2s that have mutant V proteins of inability to bind with AIP I were similar. These results have shown that the association between AIPI/Alix and V proteins is important for virus growth.2. I investigated the role of the V protein in hPIV2 replication, using a mini-genome system free of vaccinia virus. I found that the C-terminal of the V protein was essential for the inhibition and for interaction with the L protein, but not for interaction with the NP protein. These data suggest that the inhibitory effect of the hPIV2 V protein is the result of L protein binding, and not that of NP.3. I found that a homogeneous set of NP, P and L proteins of PIV2 or PIV5 was able to drive reporter gene expression from the hPIV2 mini-genome system, but all heterogeneous combinations were inactive.4. By using a yeast two-hybrid-based screening assay with hPIV2 V as bait and a target cDNA library, I got some positive clones. I would like to investigate the interaction between these clones and V.

1。我确定了一种宿主蛋白AIPI/ALIX，参与了几种包围病毒的凋亡和有效的萌芽，是HPIV2 NP和V蛋白的相互作用伴侣。为了研究AIPI参与HPIV2病毒生长的可能参与，我通过使用通过添加DOX诱导的短发夹RNA来耗尽细胞内AIP 1的细胞系。已建立细胞中的WT HPIV2生长受到10至100倍的抑制，但是RPIV2的生长速率具有突变的V蛋白，其与AIP I结合的突变v蛋白是相似的。这些结果表明，AIPI/ALIX和V蛋白之间的关联对于病毒生长很重要2。我使用没有离甲酸病毒的微型基因组系统研究了V蛋白在HPIV2复制中的作用。我发现，V蛋白的C末端对于抑制和与L蛋白的相互作用至关重要，但对于与NP蛋白的相互作用至关重要。这些数据表明，HPIV2 V蛋白的抑制作用是L蛋白结合的结果，而不是NP.3的结果。我发现PIV2或PIV5的NP，P和L蛋白均一组能够从HPIV2迷你基因组系统中驱动报告基因的表达，但所有异质组合都是不活跃的。4。通过使用HPIV2 V作为诱饵和目标cDNA库的酵母基两杂交筛选测定，我得到了一些阳性克隆。我想研究这些克隆与V之间的相互作用。