Prichard M N, Duke G M, Mocarski E S
Department of Microbiology and Immunology, Stanford University School of Medicine, California 94305-5402, USA.
J Virol. 1996 May;70(5):3018-25. doi: 10.1128/JVI.70.5.3018-3025.1996.
Human cytomegalovirus (CMV) encodes a gene, UL114, whose product is homologous to the uracil DNA glycosylase and is highly conserved in all herpesviruses. This DNA repair enzyme excises uracil residues in DNA that result from the misincorporation of dUTP or spontaneous deamination of cytosine. We constructed a recombinant virus, RC2620, that contains a large deletion in the UL114 open reading frame and carries a 1.2-kb insert containing the Escherichia coli gpt gene. RC2620 retains the capacity to replicate in primary human fibroblasts and reaches titers that are similar to those produced by the parent virus but exhibits a significantly longer replication cycle. Although the rate of expression of alpha and beta gene products appears to be unaffected by the mutation, DNA synthesis fails to proceed normally. Once initiated, DNA synthesis in mutant virus-infected cells proceeds at the same rate as with wild-type virus, but initiation is delayed by 48 h. The mutant virus also exhibits two predicted phenotypes: (i) hypersensitivity to the nucleoside analog 5-bromodeoxyuridine and (ii) retention of more uracil residues in genomic DNA than the parental virus. Together, these data suggest UL114 is required for the proper excision of uracil residues from viral DNA but in addition plays some role in establishing the correct temporal progression of DNA synthesis and viral replication. Although such involvement has not been previously observed in herpesviruses, a requirement for uracil DNA glycosylase in DNA replication has been observed in poxviruses.
人类巨细胞病毒（CMV）编码一个基因UL114，其产物与尿嘧啶DNA糖基化酶同源，并且在所有疱疹病毒中高度保守。这种DNA修复酶能切除DNA中因dUTP错掺入或胞嘧啶自发脱氨基而产生的尿嘧啶残基。我们构建了一种重组病毒RC2620，它在UL114开放阅读框中存在大片段缺失，并携带一个包含大肠杆菌gpt基因的1.2 kb插入片段。RC2620保留了在原代人成纤维细胞中复制的能力，其滴度与亲本病毒产生的滴度相似，但复制周期明显延长。尽管α和β基因产物的表达速率似乎不受该突变影响，但DNA合成无法正常进行。一旦启动，突变病毒感染细胞中的DNA合成速率与野生型病毒相同，但启动延迟了48小时。突变病毒还表现出两种预测的表型：（i）对核苷类似物5-溴脱氧尿苷高度敏感，（ii）基因组DNA中保留的尿嘧啶残基比亲本病毒更多。这些数据共同表明，UL114是从病毒DNA中正确切除尿嘧啶残基所必需的，而且在确立DNA合成和病毒复制的正确时间进程中也发挥了一定作用。虽然这种参与此前在疱疹病毒中未被观察到，但在痘病毒中已观察到DNA复制需要尿嘧啶DNA糖基化酶。
