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Structural and functional analysis of the roles of the HCV 5′ NCR miR122-dependent long-range association and SLVI in genome translation and replication

The hepatitis C virus RNA genome possesses a variety of conserved structural elements, in both coding and non-coding regions, that are important for viral replication. These elements are known or predicted to modulate key life cycle events, such as translation and genome replication, some involving...

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Autores principales: Bentley, Kirsten, Cook, Jonathan P., Tuplin, Andrew K., Evans, David J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: PeerJ Inc. 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6225842/
https://www.ncbi.nlm.nih.gov/pubmed/30416884
http://dx.doi.org/10.7717/peerj.5870
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author Bentley, Kirsten
Cook, Jonathan P.
Tuplin, Andrew K.
Evans, David J.
author_facet Bentley, Kirsten
Cook, Jonathan P.
Tuplin, Andrew K.
Evans, David J.
author_sort Bentley, Kirsten
collection PubMed
description The hepatitis C virus RNA genome possesses a variety of conserved structural elements, in both coding and non-coding regions, that are important for viral replication. These elements are known or predicted to modulate key life cycle events, such as translation and genome replication, some involving conformational changes induced by long-range RNA–RNA interactions. One such element is SLVI, a stem-loop (SL) structure located towards the 5′ end of the core protein-coding region. This element forms an alternative RNA–RNA interaction with complementary sequences in the 5′ untranslated regions that are independently involved in the binding of the cellular microRNA 122 (miR122). The switch between ‘open’ and ‘closed’ structures involving SLVI has previously been proposed to modulate translation, with lower translation efficiency associated with the ‘closed’ conformation. In the current study, we have used selective 2′-hydroxyl acylation analysed by primer extension to validate this RNA–RNA interaction in the absence and presence of miR122. We show that the long-range association (LRA) only forms in the absence of miR122, or otherwise requires the blocking of miR122 binding combined with substantial disruption of SLVI. Using site-directed mutations introduced to promote open or closed conformations of the LRA we demonstrate no correlation between the conformation and the translation phenotype. In addition, we observed no influence on virus replication compared to unmodified genomes. The presence of SLVI is well-documented to suppress translation, but these studies demonstrate that this is not due to its contribution to the LRA. We conclude that, although there are roles for SLVI in translation, the LRA is not a riboswitch regulating the translation and replication phenotypes of the virus.
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spelling pubmed-62258422018-11-09 Structural and functional analysis of the roles of the HCV 5′ NCR miR122-dependent long-range association and SLVI in genome translation and replication Bentley, Kirsten Cook, Jonathan P. Tuplin, Andrew K. Evans, David J. PeerJ Microbiology The hepatitis C virus RNA genome possesses a variety of conserved structural elements, in both coding and non-coding regions, that are important for viral replication. These elements are known or predicted to modulate key life cycle events, such as translation and genome replication, some involving conformational changes induced by long-range RNA–RNA interactions. One such element is SLVI, a stem-loop (SL) structure located towards the 5′ end of the core protein-coding region. This element forms an alternative RNA–RNA interaction with complementary sequences in the 5′ untranslated regions that are independently involved in the binding of the cellular microRNA 122 (miR122). The switch between ‘open’ and ‘closed’ structures involving SLVI has previously been proposed to modulate translation, with lower translation efficiency associated with the ‘closed’ conformation. In the current study, we have used selective 2′-hydroxyl acylation analysed by primer extension to validate this RNA–RNA interaction in the absence and presence of miR122. We show that the long-range association (LRA) only forms in the absence of miR122, or otherwise requires the blocking of miR122 binding combined with substantial disruption of SLVI. Using site-directed mutations introduced to promote open or closed conformations of the LRA we demonstrate no correlation between the conformation and the translation phenotype. In addition, we observed no influence on virus replication compared to unmodified genomes. The presence of SLVI is well-documented to suppress translation, but these studies demonstrate that this is not due to its contribution to the LRA. We conclude that, although there are roles for SLVI in translation, the LRA is not a riboswitch regulating the translation and replication phenotypes of the virus. PeerJ Inc. 2018-11-06 /pmc/articles/PMC6225842/ /pubmed/30416884 http://dx.doi.org/10.7717/peerj.5870 Text en © 2018 Bentley et al. http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited.
spellingShingle Microbiology
Bentley, Kirsten
Cook, Jonathan P.
Tuplin, Andrew K.
Evans, David J.
Structural and functional analysis of the roles of the HCV 5′ NCR miR122-dependent long-range association and SLVI in genome translation and replication
title Structural and functional analysis of the roles of the HCV 5′ NCR miR122-dependent long-range association and SLVI in genome translation and replication
title_full Structural and functional analysis of the roles of the HCV 5′ NCR miR122-dependent long-range association and SLVI in genome translation and replication
title_fullStr Structural and functional analysis of the roles of the HCV 5′ NCR miR122-dependent long-range association and SLVI in genome translation and replication
title_full_unstemmed Structural and functional analysis of the roles of the HCV 5′ NCR miR122-dependent long-range association and SLVI in genome translation and replication
title_short Structural and functional analysis of the roles of the HCV 5′ NCR miR122-dependent long-range association and SLVI in genome translation and replication
title_sort structural and functional analysis of the roles of the hcv 5′ ncr mir122-dependent long-range association and slvi in genome translation and replication
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6225842/
https://www.ncbi.nlm.nih.gov/pubmed/30416884
http://dx.doi.org/10.7717/peerj.5870
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