Nucleotide resolution mapping of influenza A virus nucleoprotein-RNA interactions reveals RNA features required for replication

Influenza A virus nucleoprotein (NP) association with viral RNA (vRNA) is essential for packaging, but the pattern of NP binding to vRNA is unclear. Here we applied photoactivatable ribonucleoside enhanced cross-linking and immunoprecipitation (PAR-CLIP) to assess the native-state of NP–vRNA interac...

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Bibliographic Details
Published in:Nature communications Vol. 9; no. 1; pp. 465 - 12
Main Authors: Williams, Graham D., Townsend, Dana, Wylie, Kristine M., Kim, Preston J., Amarasinghe, Gaya K., Kutluay, Sebla B., Boon, Adrianus C. M.
Format: Journal Article
Language:English
Published: London Nature Publishing Group UK 31.01.2018
Nature Publishing Group
Nature Portfolio
Subjects:
14/63
45/70
45/88
45/90
45/91
631/1647/514/2254
631/326/596/1578
Animals
Binding
Conserved Sequence
Crosslinking
Dogs
Gene mapping
Genome, Viral
Genomes
HEK293 Cells
Humanities and Social Sciences
Humans
Immunoprecipitation
Influenza
Influenza A
Influenza A Virus, H1N1 Subtype - genetics
Influenza A Virus, H1N1 Subtype - metabolism
Madin Darby Canine Kidney Cells
multidisciplinary
Mutagenesis
Mutation
Nucleotide Mapping
Nucleotide sequence
Nucleotides
Packaging
Packaging design
Pyridine nucleotide reductase
Ribonucleic acid
RNA
RNA viruses
RNA, Viral - genetics
RNA, Viral - metabolism
RNA-Binding Proteins - genetics
RNA-Binding Proteins - metabolism
Science
Science (multidisciplinary)
Viral Core Proteins - genetics
Viral Core Proteins - metabolism
Virus Replication - genetics
Viruses
ISSN:2041-1723, 2041-1723
Online Access:Get full text
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Summary:Influenza A virus nucleoprotein (NP) association with viral RNA (vRNA) is essential for packaging, but the pattern of NP binding to vRNA is unclear. Here we applied photoactivatable ribonucleoside enhanced cross-linking and immunoprecipitation (PAR-CLIP) to assess the native-state of NP–vRNA interactions in infected human cells. NP binds short fragments of RNA (~12 nucleotides) non-uniformly and without apparent sequence specificity. Moreover, NP binding is reduced at specific locations within the viral genome, including regions previously identified as required for viral genome segment packaging. Synonymous mutations designed to alter the predicted RNA structures in these low-NP-binding regions impact genome packaging and result in virus attenuation, whereas control mutations or mutagenesis of NP-bound regions have no effect. Finally, we demonstrate that the sequence conservation of low-NP-binding regions is required in multiple genome segments for propagation of diverse mammalian and avian IAV in host cells. Influenza A virus packaging depends on interactions between nucleoprotein (NP) and viral RNA (vRNA), but the pattern of NP binding is unclear. Using PAR-CLIP, Williams et al. here show that NP binds vRNA non-uniformly and that RNA structures in low-NP binding regions are important for packaging.
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ISSN:2041-1723
2041-1723
DOI:10.1038/s41467-018-02886-w