The role of glycosylation in the N-terminus of the hemagglutinin of a unique H4N2 with a natural polybasic cleavage site in virus fitness in vitro and in vivo

Marcel Gischke, Ola Bagato, Angele Breithaupt, David Scheibner, Claudia Blaurock, Melina Vallbracht, Axel Karger, Beate Crossley, Jutta Veits, Eva Böttcher-Friebertshäuser, Thomas C. Mettenleiter, Elsayed M. Abdelwhab

Research output: Contribution to journalArticlepeer-review

Abstract

To date, only low pathogenic (LP) H5 and H7 avian influenza viruses (AIV) have been observed to naturally shift to a highly pathogenic (HP) phenotype after mutation of the monobasic hemagglutinin (HA) cleavage site (HACS) to polybasic motifs. The LPAIV monobasic HACS is activated by tissue-restricted trypsin-like enzymes, while the HPAIV polybasic HACS is activated by ubiquitous furin-like enzymes. However, glycosylation near the HACS can affect proteolytic activation and reduced virulence of some HPAIV in chickens. In 2012, a unique H4N2 virus with a polybasic HACS was isolated from quails but was LP in chickens. Whether glycosylation sites (GS) near the HACS hinder the evolution of HPAIV H4N2 remains unclear. Here, we analyzed the prevalence of potential GS in the N-terminus of HA1, 2NYT4 and 18NGT20, in all AIV sequences and studied their impact on H4N2 virus fitness. Although the two motifs are conserved, some non-H5/H7 subtypes lack one or both GS. Both sites were glycosylated in this H4N2 virus. Deglycosylation increased trypsin-independent replication in cell culture, cell-to-cell spread and syncytium formation at low-acidic pH, but negatively affected the thermostability and receptor-binding affinity. Alteration of 2NYT4 with or without 18NGT20 enabled systemic spread of the virus to different organs including the brain of chicken embryos. However, all intranasally inoculated chickens did not show clinical signs. Together, although the conserved GS near the HACS are important for HA stability and receptor binding, deglycosylation increased the H4N2 HA-activation, replication and tissue tropism suggesting a potential role for virus adaptation in poultry.

Original languageEnglish (US)
Pages (from-to)666-678
Number of pages13
JournalVirulence
Volume12
Issue number1
DOIs
StatePublished - 2021

Keywords

  • avian influenza virus
  • glycosylation
  • h4n2
  • hemagglutinin
  • Highly pathogenic
  • low pathogenic
  • non-H5/H7
  • proteolytic activation
  • transmission
  • virulence

ASJC Scopus subject areas

  • Parasitology
  • Microbiology
  • Immunology
  • Microbiology (medical)
  • Infectious Diseases

Fingerprint

Dive into the research topics of 'The role of glycosylation in the N-terminus of the hemagglutinin of a unique H4N2 with a natural polybasic cleavage site in virus fitness in vitro and in vivo'. Together they form a unique fingerprint.

Cite this