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Human cytomegalovirus UL28 and UL29 open reading frames encode a spliced mRNA and stimulate accumulation of immediate-early RNAs. J Virol 2009 Oct;83(19):10187-97

Date

07/25/2009

Pubmed ID

19625400

Pubmed Central ID

PMC2748044

DOI

10.1128/JVI.00396-09

Scopus ID

2-s2.0-70349290582 (requires institutional sign-in at Scopus site)   34 Citations

Abstract

We have identified a spliced transcript that contains sequences from the HCMV UL29 and UL28 open reading frames. It contains amino-terminal UL29 sequences followed by UL28 sequences, and it includes a poly(A) signal derived from the 3'-untranslated region following the UL26 open reading frame. UL29/28 RNA is expressed with early kinetics, and a virus containing a FLAG epitope inserted at the amino terminus of UL29 expressed a tagged approximately 79-kDa protein, pUL29/28, that was detected at 6 h postinfection. The virus also expressed a less-abundant tagged 41-kDa protein, which corresponds in size to a protein that could be produced by translation of an unspliced UL29/28 transcript. Consistent with this prediction, both unspliced and spliced UL29/28 transcript was present in RNA isolated from polysomes. FLAG-tagged protein from the UL29/28 locus accumulated within nuclear viral replication centers during the early phase of infection. Late after infection it was present in the cytoplasm as well, and the protein was present and resistant to proteinase treatment in partially purified preparations of viral particles. Disruption of the UL29/28 locus by mutation resulted in a 10-fold decrease in the levels of DNA replication along with a similar reduction in virus yield. Quantitative reverse transcription-PCR analysis revealed an approximately 2-fold decrease in immediate-early gene expression at 4 to 10 h postinfection compared to the wild-type virus, and transient expression of pUL29/28 activated the major immediate-early promoter. Our results argue that the UL29/28 locus contributes to activation of immediate-early gene expression.

Author List

Mitchell DP, Savaryn JP, Moorman NJ, Shenk T, Terhune SS

Author

Scott Terhune PhD Professor in the Microbiology and Immunology department at Medical College of Wisconsin




MESH terms used to index this publication - Major topics in bold

Amino Acid Sequence
Cell Nucleus
Cytomegalovirus
Fibroblasts
Genes, Immediate-Early
Humans
Models, Biological
Molecular Sequence Data
Open Reading Frames
Plasmids
Promoter Regions, Genetic
RNA Splicing
RNA, Viral
Reverse Transcriptase Polymerase Chain Reaction
Viral Proteins
Virus Replication