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Functional analysis reveals G/U pairs critical for replication and trafficking of an infectious non-coding viroid RNA

While G/U pairs are present in many RNAs, the lack of molecular studies to characterize the roles of multiple G/U pairs within a single RNA limits our understanding of their biological significance. From known RNA 3D structures, we observed that the probability a G/U will form a Watson–Crick (WC) ba...

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Autores principales: Wu, Jian, Zhou, Cuiji, Li, James, Li, Chun, Tao, Xiaorong, Leontis, Neocles B, Zirbel, Craig L, Bisaro, David M, Ding, Biao
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7102988/
https://www.ncbi.nlm.nih.gov/pubmed/32083649
http://dx.doi.org/10.1093/nar/gkaa100
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author Wu, Jian
Zhou, Cuiji
Li, James
Li, Chun
Tao, Xiaorong
Leontis, Neocles B
Zirbel, Craig L
Bisaro, David M
Ding, Biao
author_facet Wu, Jian
Zhou, Cuiji
Li, James
Li, Chun
Tao, Xiaorong
Leontis, Neocles B
Zirbel, Craig L
Bisaro, David M
Ding, Biao
author_sort Wu, Jian
collection PubMed
description While G/U pairs are present in many RNAs, the lack of molecular studies to characterize the roles of multiple G/U pairs within a single RNA limits our understanding of their biological significance. From known RNA 3D structures, we observed that the probability a G/U will form a Watson–Crick (WC) base pair depends on sequence context. We analyzed 17 G/U pairs in the 359-nucleotide genome of Potato spindle tuber viroid (PSTVd), a circular non-coding RNA that replicates and spreads systemically in host plants. Most putative G/U base pairs were experimentally supported by selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE). Deep sequencing PSTVd genomes from plants inoculated with a cloned master sequence revealed naturally occurring variants, and showed that G/U pairs are maintained to the same extent as canonical WC base pairs. Comprehensive mutational analysis demonstrated that nearly all G/U pairs are critical for replication and/or systemic spread. Two selected G/U pairs were found to be required for PSTVd entry into, but not for exit from, the host vascular system. This study identifies critical roles for G/U pairs in the survival of an infectious RNA, and increases understanding of structure-based regulation of replication and trafficking of pathogen and cellular RNAs.
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spelling pubmed-71029882020-04-02 Functional analysis reveals G/U pairs critical for replication and trafficking of an infectious non-coding viroid RNA Wu, Jian Zhou, Cuiji Li, James Li, Chun Tao, Xiaorong Leontis, Neocles B Zirbel, Craig L Bisaro, David M Ding, Biao Nucleic Acids Res Molecular Biology While G/U pairs are present in many RNAs, the lack of molecular studies to characterize the roles of multiple G/U pairs within a single RNA limits our understanding of their biological significance. From known RNA 3D structures, we observed that the probability a G/U will form a Watson–Crick (WC) base pair depends on sequence context. We analyzed 17 G/U pairs in the 359-nucleotide genome of Potato spindle tuber viroid (PSTVd), a circular non-coding RNA that replicates and spreads systemically in host plants. Most putative G/U base pairs were experimentally supported by selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE). Deep sequencing PSTVd genomes from plants inoculated with a cloned master sequence revealed naturally occurring variants, and showed that G/U pairs are maintained to the same extent as canonical WC base pairs. Comprehensive mutational analysis demonstrated that nearly all G/U pairs are critical for replication and/or systemic spread. Two selected G/U pairs were found to be required for PSTVd entry into, but not for exit from, the host vascular system. This study identifies critical roles for G/U pairs in the survival of an infectious RNA, and increases understanding of structure-based regulation of replication and trafficking of pathogen and cellular RNAs. Oxford University Press 2020-02-21 /pmc/articles/PMC7102988/ /pubmed/32083649 http://dx.doi.org/10.1093/nar/gkaa100 Text en & copy; The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research. 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 reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Molecular Biology
Wu, Jian
Zhou, Cuiji
Li, James
Li, Chun
Tao, Xiaorong
Leontis, Neocles B
Zirbel, Craig L
Bisaro, David M
Ding, Biao
Functional analysis reveals G/U pairs critical for replication and trafficking of an infectious non-coding viroid RNA
title Functional analysis reveals G/U pairs critical for replication and trafficking of an infectious non-coding viroid RNA
title_full Functional analysis reveals G/U pairs critical for replication and trafficking of an infectious non-coding viroid RNA
title_fullStr Functional analysis reveals G/U pairs critical for replication and trafficking of an infectious non-coding viroid RNA
title_full_unstemmed Functional analysis reveals G/U pairs critical for replication and trafficking of an infectious non-coding viroid RNA
title_short Functional analysis reveals G/U pairs critical for replication and trafficking of an infectious non-coding viroid RNA
title_sort functional analysis reveals g/u pairs critical for replication and trafficking of an infectious non-coding viroid rna
topic Molecular Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7102988/
https://www.ncbi.nlm.nih.gov/pubmed/32083649
http://dx.doi.org/10.1093/nar/gkaa100
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