The ubiquitous cellular transcriptional factor USF targets the varicella-zoster virus open reading frame 10 promoter and determines virulence in human skin xenografts in SCIDhu mice in vivo
The ubiquitous cellular transcriptional factor USF targets the varicella-zoster virus open reading frame 10 promoter and determines virulence in human skin xenografts in SCIDhu mice in vivo
复制标题
DOI：
10.1128/jvi.02537-06
复制
发表时间：
2007-04-01
影响因子：
5.4
通讯作者：
Arvin, Ann M.
中科院分区：
文献类型：
--
作者：
Che, Xibing;Berarducci, Barbara;Arvin, Ann M.
水痘-带状疱疹病毒（VZV）开放阅读框10（ORF 10）在SCIDhu皮肤异种移植物中是毒力的决定因素，但在体内人T细胞中不是。在这个分析的调节ORF 10转录，我们已经确定了四个ORF 10相关的转录，包括一个主要的1.3 kb的RNA跨越ORF 10只和其他三个通读转录。cDNA末端快速扩增实验表明，ORF 10的1.3kb转录本具有单一的起始和终止位点。在瞬时表达试验中，ORF 10启动子被主要的VZV反式激活因子JE 62强烈刺激。缺失分析显示，相对于ORF 10转录起始位点，完整ORF 10启动子活性的大致边界在-75和-45之间以及+5和-8之间。ORF 10启动子缺失的重组病毒POKA 10-Delta pro阻断了ORF 10以及ORF 9A和ORF 9 mRNA的转录，而通读ORF 9A/9/10和ORF 9/10转录物的表达增加，补偿了单顺反子mRNA的损失。细胞因子USF特异性结合到ORF 10启动子内的共识位点，并且是IE62反式激活所需的，而破坏预测的TATA盒或Oct-1结合元件则没有影响。USF结合位点在重组病毒POKA 10-pro Delta USF中被破坏，并且不产生ORF 10蛋白。两种ORF 10启动子突变体均降低了SCIDhu皮肤异种移植物中的VZV复制。这些观察结果提供了ORF 10蛋白对皮肤中VZV发病机制的贡献的进一步证据，并证明VZV依赖于细胞转录因子USF来支持其在人体皮肤中的体内毒力。
Varicella-zoster virus (VZV) open reading frame 10 (ORF10) is a determinant of virulence in SCIDhu skin xenografts but not in human T cells in vivo. In this analysis of the regulation of ORF10 transcription, we have identified four ORF10-related transcripts, including a major 1.3-kb RNA spanning ORF10 only and three other read-through transcripts. Rapid-amplification-of-cDNA-ends experiments indicated that the 1.3-kb transcript of ORF10 has single initiation and termination sites. In transient expression assays, the ORF10 promoter was strongly stimulated by the major VZV transactivator, JE62. Deletion analyses revealed approximate boundaries for the full ORF10 promoter activity between -75 and -45 and between +5 and -8, relative to the ORF10 transcription start site. The recombinant virus POKA10-Delta pro, with the ORF10 promoter deletion, blocked transcription of ORF10 and also of ORF9A and ORF9 mRNAs, whereas expression of read-through ORF9A/9/10 and ORF9/10 transcripts was increased, compensating for the loss of the monocistronic mRNAs. The cellular factor USF bound specifically to its consensus site within the ORF10 promoter and was required for IE62 transactivation, whereas disrupting the predicted TATA boxes or Oct-1 binding elements had no effect. The USF binding site was disrupted in the recombinant virus, POKA10-pro Delta USF, and no ORF10 protein was produced. Both ORF10 promoter mutants reduced VZV replication in SCIDhu skin xenografts. These observations provided further evidence of the contribution of the ORF10 protein to VZV pathogenesis in skin and demonstrated that VZV depends upon the cellular transcriptional factor USF to support its virulence in human skin in vivo.