Rewriting nature’s assembly manual for a ssRNA virus

Reidun Twarock, Nikesh Patel, Wroblewski Emma, German Leonov, Simon E. V. Phillips, Roman Tuma, Peter G. Stockley

Research output: Contribution to journalArticlepeer-review


Satellite tobacco necrosis virus (STNV) is one of the smallest viruses known. Its genome encodes only its coat protein (CP) subunit, relying on the polymerase of its helper virus TNV for replication. The genome has been shown to contain a cryptic set of dispersed assembly signals in the form of stem-loops that each present a minimal CP-binding motif AXXA in the loops. The genomic fragment encompassing nucleotides 1–127 is predicted to contain five such packaging signals (PSs). We have used mutagenesis to determine the critical assembly features in this region. These include the CP-binding motif, the relative placement of PS stem-loops, their number, and their folding propensity. CP binding has an electrostatic contribution, but assembly nucleation is dominated by the recognition of the folded PSs in the RNA fragment. Mutation to remove all AXXA motifs in PSs throughout the genome yields an RNA that is unable to assemble efficiently. In contrast, when a synthetic 127-nt fragment encompassing improved PSs is swapped onto the RNA otherwise lacking CP recognition motifs, assembly is partially restored, although the virus-like particles created are incomplete, implying that PSs outside this region are required for correct assembly. Swapping this improved region into the wild-type STNV1 sequence results in a better assembly substrate than the viral RNA, producing complete capsids and outcompeting the wild-type genome in head-to-head competition. These data confirm details of the PS-mediated assembly mechanism for STNV and identify an efficient approach for production of stable virus-like particles encapsidating nonnative RNAs or other cargoes.

Original languageEnglish
Article number1706951114
Pages (from-to)12255-12260
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number46
Publication statusPublished - 14 Nov 2017

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  • Satellite tobacco necrosis virus
  • packaging signals
  • viral assembly
  • synthetic virology
  • Viral assembly
  • Packaging signals
  • Synthetic virology

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