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PMID: 8627778 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Human cytomegalovirus uracil DNA glycosylase is required for the normal temporal regulation of both DNA synthesis and viral replication.

Journal of virology ·Vol. 70 ·No. 5 ·1996-05-00 ·Pages 3018-25

Prichard MN, Duke GM, Mocarski ES

Abstract

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.

MeSH Terms
Base Sequence Bromodeoxyuridine/toxicity Cell Division Cells, Cultured Conserved Sequence Cytomegalovirus/enzymology,genetics,physiology DNA Glycosylases DNA Repair DNA Replication/drug effects DNA, Viral/biosynthesis Escherichia coli Fibroblasts Gene Expression Regulation, Viral Genes, Viral Herpesviridae/genetics Humans Kinetics Male N-Glycosyl Hydrolases/genetics,metabolism Open Reading Frames Phenotype Sequence Deletion Skin Time Factors Uracil-DNA Glycosidase Virus Replication/drug effects
Chemicals
DNA, Viral DNA Glycosylases N-Glycosyl Hydrolases Uracil-DNA Glycosidase Bromodeoxyuridine
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Prichard M N
Department of Microbiology and Immunology, Stanford University School of Medicine, California 94305-5402, USA.
Duke G M
Mocarski E S
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Article Info
Journal
Journal of virology
Abbr.
J Virol
ISSN
0022-538X
DOI
10.1128/JVI.70.5.3018-3025.1996
Published
1996-05-00
Pages
3018-25
Language
English
Region
United States
NLM ID
0113724
PMCID
PMC190161
Subset
IM
Grants
NIAID NIH HHS · R01AI20211 · United States
NIAID NIH HHS · AI08771 · United States
NIAID NIH HHS · F32 AI009008-02 · United States
NIAID NIH HHS · AI09009 · United States
NIAID NIH HHS · F32 AI009008 · United States
External Links
PubMed source DOI: 10.1128/JVI.70.5.3018-3025.1996 PMC Full Text View journal details
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