EGFR Interacts with the Fusion Protein of Respiratory Syncytial Virus Strain 2-20 and Mediates Infection and Mucin Expression

Abstract

Respiratory syncytial virus (RSV) is the major cause of viral lower respiratory tract illness in children. In contrast to the RSV prototypic strain A2, clinical isolate RSV 2–20 induces airway mucin expression in mice, a clinically relevant phenotype dependent on the fusion (F) protein of the RSV strain. Epidermal growth factor receptor (EGFR) plays a role in airway mucin expression in other systems; therefore, we hypothesized that the RSV 2–20 F protein stimulates EGFR signaling. Infection of cells with chimeric strains RSV A2-2-20F and A2-2-20GF or over-expression of 2–20 F protein resulted in greater phosphorylation of EGFR than infection with RSV A2 or over-expression of A2 F, respectively. Chemical inhibition of EGFR signaling or knockdown of EGFR resulted in diminished infectivity of RSV A2-2-20F but not RSV A2. Over-expression of EGFR enhanced the fusion activity of 2–20 F protein in trans. EGFR co-immunoprecipitated most efficiently with RSV F proteins derived from “mucogenic” strains. RSV 2–20 F and EGFR co-localized in H292 cells, and A2-2-20GF-induced MUC5AC expression was ablated by EGFR inhibitors in these cells. Treatment of BALB/c mice with the EGFR inhibitor erlotinib significantly reduced the amount of RSV A2-2-20F-induced airway mucin expression. Our results demonstrate that RSV F interacts with EGFR in a strain-specific manner, EGFR is a co-factor for infection, and EGFR plays a role in RSV-induced mucin expression, suggesting EGFR is a potential target for RSV disease.

Author Summary

Respiratory syncytial virus (RSV) is responsible for severe lower respiratory disease in infants and young children. Overabundant airway mucus contributes to airway obstruction in RSV bronchiolitis, and a better understanding of RSV pathogenesis may contribute to needed therapies and vaccines. We reported previously that RSV clinical isolate strain 2–20 induces more airway mucin expression in mice than prototypic RSV strains and that the 2–20 fusion (F) protein mediates mucin induction. Epidermal growth factor receptor (EGFR) has been shown to play a role in lung mucin expression. We identified a functional interaction between 2–20 F and EGFR, in that 2–20 F expression activated EGFR and, reciprocally, EGFR expression increased 2–20 F fusion activity. RSV F and EGFR co-localized in infected cells. EGFR co-immunoprecipitated with RSV F protein from various RSV strains, and the strength of this in vitro interaction correlated with strain-specific airway pathogenicity in mice. EGFR inhibition abrogated 2–20 F-mediated infection in vitro and mucin expression induction in vivo. These data identify EGFR as a novel strain-specific co-factor of RSV infection and suggest EGFR may be a target for ameliorating RSV disease.