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Investigating molecular mechanisms of 2A-stimulated ribosomal pausing and frameshifting in Theilovirus.

Published version
Peer-reviewed

Type

Article

Change log

Authors

Napthine, Sawsan 
Kibe, Anuja 
Brown, Katherine 

Abstract

The 2A protein of Theiler's murine encephalomyelitis virus (TMEV) acts as a switch to stimulate programmed -1 ribosomal frameshifting (PRF) during infection. Here, we present the X-ray crystal structure of TMEV 2A and define how it recognises the stimulatory RNA element. We demonstrate a critical role for bases upstream of the originally predicted stem-loop, providing evidence for a pseudoknot-like conformation and suggesting that the recognition of this pseudoknot by beta-shell proteins is a conserved feature in cardioviruses. Through examination of PRF in TMEV-infected cells by ribosome profiling, we identify a series of ribosomal pauses around the site of PRF induced by the 2A-pseudoknot complex. Careful normalisation of ribosomal profiling data with a 2A knockout virus facilitated the identification, through disome analysis, of ribosome stacking at the TMEV frameshifting signal. These experiments provide unparalleled detail of the molecular mechanisms underpinning Theilovirus protein-stimulated frameshifting.

Description

Funder: Helmholtz Association


Funder: Royal Society

Keywords

Frameshifting, Ribosomal, Ribosomes, Theilovirus, Viral Proteins

Journal Title

Nucleic Acids Res

Conference Name

Journal ISSN

0305-1048
1362-4962

Volume Title

49

Publisher

Oxford University Press (OUP)
Sponsorship
Biotechnology and Biological Sciences Research Council (BB/L000334/1)
European Research Council (646891)
Wellcome Trust (202797/Z/16/Z)
Wellcome Trust (106207/Z/14/Z)
Wellcome Trust (203864/Z/16/Z)
Wellcome Trust (098406/Z/12/B)
Biotechnology and Biological Sciences Research Council (BB/J007072/1)
Wellcome Trust (098406/Z/12/Z)
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