Internal entry of ribosomes is directed by the 5' noncoding region of classical swine fever virus and is dependent on the presence of an RNA pseudoknot upstream of the initiation codon.

Journal of virology ·Vol. 71 ·No. 1 ·1997-01-00 ·Pages 451-7

Rijnbrand R, van der Straaten T, van Rijn PA, Spaan WJ, Bredenbeek PJ

Abstract

Bicistronic RNAs containing the 373-nucleotide-long 5' nontranslated region (NTR) of the classical swine fever virus (CSFV) genome as intercistronic spacer were used to show the presence of an internal ribosome entry site (IRES) in the 5' end of the CSFV genome. By coexpression of the poliovirus 2A protease it was demonstrated that the CSFV 5' NTR-driven translation is independent of the presence of functional eukaryotic initiation factor eIF-4F. Deletion analysis indicated that the 5' border of the IRES is located between nucleotides 28 and 66. The role of a proposed pseudoknot structure at the 3' end of the CSFV 5' NTR in IRES-mediated translation was investigated by site-directed mutagenesis. Mutant RNAs that had lost the ability to base pair in stem II of the pseudoknot were translationally inactive. Translation to wild-type levels could be restored through the introduction of compensatory complementary base changes that repaired base pairing in stem II. In addition, we showed that the AUG codon, which is located 7 nucleotides upstream of the polyprotein initiation site and is conserved in pestiviruses, could not be used to initiate translation. Also, an AUG codon introduced downstream of the polyprotein initiation site was not recognized as an initiation site by ribosomes. These data suggest that after internal entry on the CSFV 5' NTR, ribosomal scanning for the initiation codon is limited to a small region.

MeSH Terms

Animals Classical Swine Fever Virus/genetics Codon, Initiator Cysteine Endopeptidases/metabolism Genes Nucleic Acid Conformation Protein Biosynthesis RNA, Messenger RNA, Viral Regulatory Sequences, Nucleic Acid Ribosomes Structure-Activity Relationship Swine/virology Tumor Cells, Cultured Viral Proteins

Chemicals

Codon, Initiator RNA, Messenger RNA, Viral Viral Proteins Cysteine Endopeptidases picornain 2A, Picornavirus

Authors & Affiliations

5 authors, click to expand affiliations / ORCID

Rijnbrand R

Department of Virology, Leiden University, The Netherlands.

van der Straaten T

van Rijn P A

Spaan W J

Bredenbeek P J

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32 references, click to expand

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Article Info

Journal

Journal of virology

Abbr.

J Virol

ISSN

0022-538X

DOI

10.1128/JVI.71.1.451-457.1997

Published

1997-01-00

Pages

451-7

Language

English

Region

United States

NLM ID

0113724

PMCID

PMC191071

Subset

External Links

PubMed source DOI: 10.1128/JVI.71.1.451-457.1997 PMC Full Text View journal details

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Internal entry of ribosomes is directed by the 5' noncoding region of classical swine fever virus and is dependent on the presence of an RNA pseudoknot upstream of the initiation codon.

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