Host Lipid Rafts Play a Major Role in Binding and Endocytosis of Influenza A Virus.

Dileep Verma,Dinesh Gupta,Sunil Lal

doi:10.3390/v10110650

Abstract

Influenza still remains one of the most challenging diseases, posing a significant threat to public health. Host lipid rafts play a critical role in influenza A virus (IAV) assembly and budding, however, their role in polyvalent IAV host binding and endocytosis had remained elusive until now. In the present study, we observed co-localization of IAV with a lipid raft marker ganglioside, GM1, on the host surface. Further, we isolated the lipid raft micro-domains from IAV infected cells and detected IAV protein in the raft fraction. Finally, raft disruption using Methyl-β-Cyclodextrin revealed significant reduction in IAV host binding, suggesting utilization of host rafts for polyvalent binding on the host cell surface. In addition to this, cyclodextrin mediated inhibition of raft-dependent endocytosis showed significantly reduced IAV internalization. Interestingly, exposure of cells to cyclodextrin two hours post-IAV binding showed no such reduction in IAV entry, indicating use of raft-dependent endocytosis for host entry. In summary, this study demonstrates that host lipid rafts are selected by IAV as a host attachment factors for multivalent binding, and IAV utilizes these micro-domains to exploit raft-dependent endocytosis for host internalization, a virus entry route previously unknown for IAV.

Highlights

Flu caused by influenza A virus (IAV) affects millions of people by seasonal and rare pandemic outbreaks [1]
To investigate whether IAV utilizes the host membrane rafts during binding, seeded A549 cells were either incubated with IAV at 4 ◦ C to arrest the attached virus on the host cell surface or left uninfected
These cells were subjected to cholera toxin-GM1 based lipid raft labeling (Red) at 4 ◦ C to prevent internalization of GM1-Cholera Toxin-B (CTB) complex and to visualize plasma membrane (PM) rafts in the presence and absence of ligand (IAV) binding

Summary

Introduction

Flu caused by influenza A virus (IAV) affects millions of people by seasonal and rare pandemic outbreaks [1]. IAV belongs to the Orthomyxoviridae family of viruses containing a negative-sense, single-stranded, segmented RNA genome distributed along eight segments encoding 11 major viral and some accessory proteins. Each virus is structurally divided into three major components: a central core of RNA genome (vRNA) with associated proteins surrounded by matrix proteins (M1) and the outermost host derived lipid bilayer with two key viral proteins: hemagglutinin (HA) and neuraminidase (NA) [2]. Host rafts have been reported to serve as a platform for virus attachment and entry, virion assembly, and they are a preferred site for viral budding in the case of enveloped viruses [5,6]. Clustering of host lipid rafts induces a signaling platform and activates receptor tyrosine kinases RTKs upon multivalent binding for virus uptake [7]. Cyclodextrins are Viruses 2018, 10, 650; doi:10.3390/v10110650 www.mdpi.com/journal/viruses

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