Access to a main alphaherpesvirus receptor, located basolaterally in the respiratory epithelium, is masked by intercellular junctions

Jolien Van Cleemput,Roger Maes,Kathlyn Laval,Katrien C K Poelaert,Hans J Nauwynck,Gisela S Hussey,Wim Van Den Broeck

doi:10.1038/s41598-017-16804-5

Jolien Van Cleemput, Roger Maes + Show 5 more

Open Access

https://doi.org/10.1038/s41598-017-16804-5

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Abstract

The respiratory epithelium of humans and animals is frequently exposed to alphaherpesviruses, originating from either external exposure or reactivation from latency. To date, the polarity of alphaherpesvirus infection in the respiratory epithelium and the role of respiratory epithelial integrity herein has not been studied. Equine herpesvirus type 1 (EHV1), a well-known member of the alphaherpesvirus family, was used to infect equine respiratory mucosal explants and primary equine respiratory epithelial cells (EREC), grown at the air-liquid interface. EHV1 binding to and infection of mucosal explants was greatly enhanced upon destruction of the respiratory epithelium integrity with EGTA or N-acetylcysteine. EHV1 preferentially bound to and entered EREC at basolateral cell surfaces. Restriction of infection via apical inoculation was overcome by disruption of intercellular junctions. Finally, basolateral but not apical EHV1 infection of EREC was dependent on cellular N-linked glycans. Overall, our findings demonstrate that integrity of the respiratory epithelium is crucial in the host’s innate defence against primary alphaherpesvirus infections. In addition, by targeting a basolaterally located receptor in the respiratory epithelium, alphaherpesviruses have generated a strategy to efficiently escape from host defence mechanisms during reactivation from latency.

Highlights

Alphaherpesviruses replicate in the mucosae of the upper respiratory and/or the genital tract of their specific host, which is essential for their efficient transmission
Tight junctions (TJ) are the most apically located intercellular junctions (ICJ) and function as a size- and ion selective gate for the passage of molecules in between adjacent cells[5,6]. They prevent diffusion of plasma membrane lipids andproteins from the apical to the basolateral surface and vice versa, and of incoming pathogens attached to apical surfaces
None of these methods could be applied on mucosal explants and we set up a new protocol to verify ICJ integrity by examining the intercellular space in haematoxylin-eosin stained paraffin coupes by means of image analysis

Summary

Introduction

Alphaherpesviruses replicate in the mucosae of the upper respiratory and/or the genital tract of their specific host, which is essential for their efficient transmission. Tight junctions (TJ) are the most apically located ICJ and function as a size- and ion selective gate for the passage of molecules in between adjacent cells[5,6] They prevent diffusion of plasma membrane lipids and (glyco)proteins from the apical to the basolateral surface and vice versa, and of incoming pathogens attached to apical surfaces. EHV1 infected leukocytes transfer the virus to local vascular endothelial cells, where viral replication results in vasculitis, thrombosis and edema. These pathologies lead to severe symptoms such as abortion and nervous system disorders. Because vaccines and antivirals are not fully effective, a complete understanding of EHV1 pathogenesis is needed, starting with its primary replication in the respiratory epithelium

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