Generation of replication-proficient influenza virus NS1 point mutants with interferon-hyperinducer phenotype

Maite Pérez-Cidoncha, Marian J Killip, Víctor J Asensio, Yolanda Fernández, José A Bengoechea, Richard E Randall, Juan Ortín

Research output: Contribution to journalArticlepeer-review

2 Citations (Scopus)
210 Downloads (Pure)

Abstract

The NS1 protein of influenza A viruses is the dedicated viral interferon (IFN)-antagonist. Viruses lacking NS1 protein expression cannot multiply in normal cells but are viable in cells deficient in their ability to produce or respond to IFN. Here we report an unbiased mutagenesis approach to identify positions in the influenza A NS1 protein that modulate the IFN response upon infection. A random library of virus ribonucleoproteins containing circa 40 000 point mutants in NS1 were transferred to infectious virus and amplified in MDCK cells unable to respond to interferon. Viruses that activated the interferon (IFN) response were subsequently selected by their ability to induce expression of green-fluorescent protein (GFP) following infection of A549 cells bearing an IFN promoter-dependent GFP gene. Using this approach we isolated individual mutant viruses that replicate to high titers in IFN-compromised cells but, compared to wild type viruses, induced higher levels of IFN in IFN-competent cells and had a reduced capacity to counteract exogenous IFN. Most of these viruses contained not previously reported NS1 mutations within either the RNA-binding domain, the effector domain or the linker region between them. These results indicate that subtle alterations in NS1 can reduce its effectiveness as an IFN antagonist without affecting the intrinsic capacity of the virus to multiply. The general approach reported here may facilitate the generation of replication-proficient, IFN-inducing virus mutants, that potentially could be developed as attenuated vaccines against a variety of viruses.

Original languageEnglish
Article numbere98668
Number of pages10
JournalPloS one
Volume9
Issue number6
DOIs
Publication statusPublished - 02 Jun 2014

Keywords

  • Animals
  • Base Sequence
  • Cell Line
  • DNA Primers
  • Dogs
  • Green Fluorescent Proteins
  • Humans
  • Immunity, Innate
  • Influenza A virus
  • Interferons
  • Point Mutation
  • Promoter Regions, Genetic
  • Viral Nonstructural Proteins
  • Virus Replication

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