Origin of Expression of the Herpes Simplex Virus Type 1 Protein US1.5
Origin of Expression of the Herpes Simplex Virus Type 1 Protein US1.5
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DOI：
10.1128/jvi.00984-09
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发表时间：
2009-09-15
影响因子：
5.4
通讯作者：
Schaffer, Priscilla A.
中科院分区：
文献类型：
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作者：
Bowman, J. Jason;Schaffer, Priscilla A.
ICP22是1型单纯疱疹病毒（HSV-1）的一种即时早期蛋白，是病毒在非允许细胞类型中的复制和一类晚期病毒蛋白（包括糖蛋白c）的表达所必需的。由于全长蛋白与框架内c端特异性蛋白U(S)1.5的共表达，对ICP22功能机制的理解变得更加复杂。在本报告中，我们证实U(S)1.5蛋白是一个真正的翻译产物，因为它在三种实验室菌株和两种低传代临床HSV-1分离株的感染中被检测到。为了阐明ICP22和U(S)1.5蛋白的表达模式，我们在瞬时表达实验中检测了它们在质粒中的合成。由于之前的研究已经确定了两个不同的U(S)1.5翻译起始位点，我们试图通过研究一系列缺失、无义和蛋氨酸替换对U(S)1.5表达的影响来确定哪一个是正确的。首先，由ICP22开放阅读框（ORF）编码的氨基酸90 ~ 420在十二烷基硫酸钠-聚丙烯酰胺凝胶中以U(S)1.5的迁移率迁移。其次，在M90下游引入一个停止密码子，可以抑制ICP22和U(S)1.5的表达。最后，M90突变为丙氨酸（M90A），使全长ICP22得以表达，同时显著降低U(S)1.5的表达。在感染M90A突变病毒的细胞中，U(S)1.5蛋白表达水平降低，但ICP22蛋白表达水平不降低。因此，我们得出结论，IC22和U(S)1.5的表达可以相互独立地发生，U(S)1.5的翻译始于ICP22 ORF的M90。
ICP22, an immediate-early protein of herpes simplex virus type 1 (HSV-1), is required for viral replication in nonpermissive cell types and for expression of a class of late viral proteins which includes glycoprotein C. An understanding of the mechanism of ICP22 function has been complicated by the coexpression of the full-length protein with an in-frame, C-terminus-specific protein, U(S)1.5. In this report, we confirm that the U(S)1.5 protein is a bona fide translation product since it is detected during infections with three laboratory strains and two low-passage clinical isolates of HSV-1. To clarify the expression patterns of the ICP22 and U(S)1.5 proteins, we examined their synthesis from plasmids in transient expression assays. Because previous studies had identified two different U(S)1.5 translational start sites, we attempted to determine which is correct by studying the effects of a series of deletion, nonsense, and methionine substitutions on U(S)1.5 expression. First, amino acids 90 to 420 encoded by the ICP22 open reading frame (ORF) migrated at the mobility of U(S)1.5 in sodium dodecyl sulfate-polyacrylamide gels. Second, introduction of a stop codon downstream of M90 ablated expression of both ICP22 and U(S)1.5. Finally, mutation of M90 to alanine (M90A) allowed expression of full-length ICP22 while dramatically reducing expression of U(S)1.5. Levels of U(S)1.5 but not ICP22 protein expression were also reduced in cells infected with an M90A mutant virus. Thus, we conclude that expression of IC22 and that of U(S)1.5 can occur independently of each other and that U(S)1.5 translation initiates at M90 of the ICP22 ORF.