Characterization of Ribosomal Frameshifting in Theiler's Murine Encephalomyelitis Virus
| Autor: | Leanne K, Finch, Roger, Ling, Sawsan, Napthine, Allan, Olspert, Thomas, Michiels, Cécile, Lardinois, Susanne, Bell, Gary, Loughran, Ian, Brierley, Andew E, Firth |
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| Přispěvatelé: | Napthine, Sawsan [0000-0001-7404-8494], Brierley, Ian [0000-0003-3965-4370], Firth, Andrew [0000-0002-7986-9520], Apollo - University of Cambridge Repository |
| Rok vydání: | 2015 |
| Předmět: |
viruses
Immunology Immunoblotting Viral Plaque Assay Biology Microbiology Mass Spectrometry Cell Line 03 medical and health sciences Mice Theilovirus Virology Rosaniline Dyes Animals Spotlight Luciferases Biological sciences 030304 developmental biology Genetics Recombination Genetic 0303 health sciences Translational frameshift 030306 microbiology Frameshifting Ribosomal Murine encephalomyelitis virus Genome Replication and Regulation of Viral Gene Expression Research council Mutagenesis Insect Science |
| Zdroj: | Journal of Virology |
| ISSN: | 1098-5514 0022-538X |
| DOI: | 10.1128/jvi.01043-15 |
| Popis: | Theiler's murine encephalomyelitis virus (TMEV) is a member of the genus Cardiovirus in the Picornaviridae , a family of positive-sense single-stranded RNA viruses. Previously, we demonstrated that in the related cardiovirus, Encephalomyocarditis virus , a programmed −1 ribosomal frameshift (−1 PRF) occurs at a conserved G_GUU_UUU sequence within the 2B-encoding region of the polyprotein open reading frame (ORF). Here we show that −1 PRF occurs at a similar site during translation of the TMEV genome. In addition, we demonstrate that a predicted 3′ RNA stem-loop structure at a noncanonical spacing downstream of the shift site is required for efficient frameshifting in TMEV and that frameshifting also requires virus infection. Mutating the G_GUU_UUU shift site to inhibit frameshifting results in an attenuated virus with reduced growth kinetics and a small-plaque phenotype. Frameshifting in the virus context was found to be extremely efficient at 74 to 82%, which, to our knowledge, is the highest frameshifting efficiency recorded to date for any virus. We propose that highly efficient −1 PRF in TMEV provides a mechanism to escape the confines of equimolar expression normally inherent in the single-polyprotein expression strategy of picornaviruses. IMPORTANCE Many viruses utilize programmed −1 ribosomal frameshifting (−1 PRF) to produce different protein products at a defined ratio, or to translate overlapping ORFs to increase coding capacity. With few exceptions, −1 PRF occurs on specific “slippery” heptanucleotide sequences and is stimulated by RNA structure beginning 5 to 9 nucleotides (nt) downstream of the slippery site. Here we describe an unusual case of −1 PRF in Theiler's murine encephalomyelitis virus (TMEV) that is extraordinarily efficient (74 to 82% of ribosomes shift into the alternative reading frame) and, in stark contrast to other examples of −1 PRF, is dependent upon a stem-loop structure beginning 14 nt downstream of the slippery site. Furthermore, in TMEV-based reporter constructs in transfected cells, efficient frameshifting is critically dependent upon virus infection. We suggest that TMEV evolved frameshifting as a novel mechanism for removing ribosomes from the message (a “ribosome sink”) to downregulate synthesis of the 3′-encoded replication proteins. |
| Databáze: | OpenAIRE |
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