An unusual internal ribosome entry site in the herpes simplex virus thymidine kinase gene

Anthony Griffiths, Donald M. Coen

Research output: Contribution to journalArticlepeer-review

32 Scopus citations

Abstract

We have investigated a herpes simplex virus mutant that expresses low levels of thymidine kinase (TK), a phenotype associated with drug resistance and pathogenicity, despite a single-base deletion in the gene. Using a dual-reporter system, a 39-nt sequence including the mutation was shown to direct expression of the downstream reporter gene in reticulocyte lysate. Translation of the downstream reporter was not impaired when the mRNA lacked a 5′ cap or had a stable stem loop 5′ of the upstream reporter and was relatively resistant to edeine, an antibiotic that prevents AUG codon recognition by the 40S-elF2-GTP/Met-tRNAi complex. Twelve nucleotides were as active as the original sequence for translation of the downstream reporter. Surprisingly, this sequence lacks an AUG codon. Analysis of point mutations showed that a CUG codon in the sequence was important. However, many single-base changes had only limited effects, and introduction of AUG codons did not increase translation. A mutant virus containing both the single-base deletion and a mutation that reduced downstream translation in vitro had significantly less TK activity than a virus with the single-base deletion alone. Thus, a remarkably short internal ribosome entry site (IRES) that lacks an AUG codon resides in the viral tk gene. The IRES appears to be responsible for TK expression from a drug-resistant mutant that would otherwise express no TK, which may contribute to pathogenicity. Because we found numerous short sequences with IRES activity, there might be many hitherto unrecognized polypeptides expressed at low levels from eukaryotic mRNAs.

Original languageEnglish (US)
Pages (from-to)9667-9672
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume102
Issue number27
DOIs
StatePublished - Jul 5 2005
Externally publishedYes

Keywords

  • Acyclovir
  • Drug resistance
  • Pathogenesis
  • Proteomics
  • Translation

ASJC Scopus subject areas

  • General

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