Co-selection of West Nile virus nucleotides that confer resistance to an antisense oligomer while maintaining long-distance RNA/RNA base pairings

Bo Zhang, Hongping Dong, David A. Stein, Pei-Yong Shi

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

West Nile virus (WNV) genome cyclization is mediated by two pairs of long-distance RNA/RNA interactions: the 5′CS/3′CSI (conserved sequence) and the 5′UAR/3′UAR (upstream AUG region) base pairings. Antisense peptide-conjugated phosphorodiamidate morpholino oligomers (PPMOs), designed to interfere with the 5′CS/3′CSI or 5′UAR/3′UAR base pairings, were previously shown to inhibit WNV. In this study, we selected and characterized WNVs resistant to a PPMO targeting the 3′UAR (3′UAR-PPMO). All resistant viruses accumulated one-nucleotide mutations within the 3′UAR, leading to a single-nucleotide mismatch or a weakened base-pairing interaction with the 3′UAR-PPMO. Remarkably, a one-nucleotide mutation within the 5′UAR was correspondingly co-selected; the 5′UAR mutation restored the base pairing with the 3′UAR mutation. Mutagenesis of WNV demonstrated that the single-nucleotide change within the 3′UAR-PPMO-target site conferred the resistance. RNA binding analysis indicated that the single-nucleotide change reduced the ability of 3′UAR-PPMO to block the RNA/RNA interaction required for genome cyclization. The results suggest a mechanism by which WNV develops resistance to 3′UAR-PPMO, through co-selection of the 5′UAR and 3′UAR, to create a mismatch or a weakened base-pairing interaction with the PPMO, while maintaining the 5′UAR/3′UAR base pairings.

Original languageEnglish (US)
Pages (from-to)98-106
Number of pages9
JournalVirology
Volume382
Issue number1
DOIs
StatePublished - Dec 5 2008
Externally publishedYes

Fingerprint

Morpholinos
West Nile virus
Base Pairing
Nucleotides
RNA
Mutation
Cyclization
Genome
Conserved Sequence
Mutagenesis
Viruses
Peptides

Keywords

  • Antiviral therapy
  • Flavivirus replication
  • Genome cyclization
  • RNA cis elements
  • West Nile virus

ASJC Scopus subject areas

  • Virology

Cite this

Co-selection of West Nile virus nucleotides that confer resistance to an antisense oligomer while maintaining long-distance RNA/RNA base pairings. / Zhang, Bo; Dong, Hongping; Stein, David A.; Shi, Pei-Yong.

In: Virology, Vol. 382, No. 1, 05.12.2008, p. 98-106.

Research output: Contribution to journalArticle

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AB - West Nile virus (WNV) genome cyclization is mediated by two pairs of long-distance RNA/RNA interactions: the 5′CS/3′CSI (conserved sequence) and the 5′UAR/3′UAR (upstream AUG region) base pairings. Antisense peptide-conjugated phosphorodiamidate morpholino oligomers (PPMOs), designed to interfere with the 5′CS/3′CSI or 5′UAR/3′UAR base pairings, were previously shown to inhibit WNV. In this study, we selected and characterized WNVs resistant to a PPMO targeting the 3′UAR (3′UAR-PPMO). All resistant viruses accumulated one-nucleotide mutations within the 3′UAR, leading to a single-nucleotide mismatch or a weakened base-pairing interaction with the 3′UAR-PPMO. Remarkably, a one-nucleotide mutation within the 5′UAR was correspondingly co-selected; the 5′UAR mutation restored the base pairing with the 3′UAR mutation. Mutagenesis of WNV demonstrated that the single-nucleotide change within the 3′UAR-PPMO-target site conferred the resistance. RNA binding analysis indicated that the single-nucleotide change reduced the ability of 3′UAR-PPMO to block the RNA/RNA interaction required for genome cyclization. The results suggest a mechanism by which WNV develops resistance to 3′UAR-PPMO, through co-selection of the 5′UAR and 3′UAR, to create a mismatch or a weakened base-pairing interaction with the PPMO, while maintaining the 5′UAR/3′UAR base pairings.

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