The Matrix protein M1 from influenza C virus induces tubular membrane invaginations in an in vitro cell membrane model

David Saletti,Marta Bally,Daniel Midtvedt,Patricia Bassereau,Jens Radzimanowski,Stéphanie Mangenot,Winfried Weissenhorn,Gregory Effantin

doi:10.1038/srep40801

Abstract

Matrix proteins from enveloped viruses play an important role in budding and stabilizing virus particles. In order to assess the role of the matrix protein M1 from influenza C virus (M1-C) in plasma membrane deformation, we have combined structural and in vitro reconstitution experiments with model membranes. We present the crystal structure of the N-terminal domain of M1-C and show by Small Angle X-Ray Scattering analysis that full-length M1-C folds into an elongated structure that associates laterally into ring-like or filamentous polymers. Using negatively charged giant unilamellar vesicles (GUVs), we demonstrate that M1-C full-length binds to and induces inward budding of membrane tubules with diameters that resemble the diameter of viruses. Membrane tubule formation requires the C-terminal domain of M1-C, corroborating its essential role for M1-C polymerization. Our results indicate that M1-C assembly on membranes constitutes the driving force for budding and suggest that M1-C plays a key role in facilitating viral egress.

Highlights

To the plasma membrane by HA and/or NA through interactions with their cytoplasmic tails[15,16] or by the M2 protein[17]
In addition to the fraction of M1 from influenza C virus (M1-C) eluting as a monomer from the size exclusion chromatography (SEC) column, ringlike and/or spiral structures were present in the void volume fraction of the SEC column at low pH conditions
We confirm that full length M1-C forms an elongated structure at low pH with an extended possibly flexible C-terminal domain, as predicted for M1-A by small angle x-ray scattering (SAXS) analysis[39,40]

Summary

Introduction

To the plasma membrane by HA and/or NA through interactions with their cytoplasmic tails[15,16] or by the M2 protein[17]. The N-terminal domain is followed by a short linker that was proposed to contain a zinc binding site[36] and the C-terminal domain The latter dimerizes in vitro[37], is essential for M1 polymerization[38] and provides a link to the ribonucleoprotein (RNP)[32]. We show that the N-terminal domain of M1-C adopts a similar fold as M1-A, despite the low sequence homology of both proteins, but coordinates two Mg2+ ions which increases the positive surface charge and may facilitate electrostatic interactions with negatively charged membranes. M1-C interaction with GUVs containing negatively charged lipids leads to inward membrane tubulation, topologically similar to virus budding. Our study confirms that M1-C assembly on membranes constitutes the driving force for influenza C virus bud formation

Methods

Results

Conclusion