Influenza A matrix protein M1 is sufficient to induce lipid membrane deformation

Abstract

The matrix protein M1 of the Influenza A virus is considered to mediate viral assembly and budding at the plasma membrane (PM) of infected cells. In order for a new viral particle to form, the PM lipid bilayer has to bend into a vesicle towards the extracellular side. Studies in cellular models have proposed that different viral proteins might be responsible for inducing membrane curvature in this context (including M1), but a clear consensus has not been reached. In this study, we use a combination of fluorescence microscopy, cryogenic transmission electron microscopy (cryo-TEM), cryo-electron tomography (cryo-ET) and scanning fluorescence correlation spectroscopy (sFCS) to investigate M1-induced membrane deformation in biophysical models of the PM. Our results indicate that M1 is indeed capable to cause membrane curvature in lipid bilayers containing negatively-charged lipids, in the absence of other viral components. Furthermore, we prove that simple protein binding is not sufficient to induce membrane restructuring. Rather, it appears that stable M1-M1 interactions and multimer formation are required in order to alter the bilayer threedimensional structure, through the formation of a protein scaffold. Finally, our results suggest that, in a physiological context, M1-induced membrane deformation might be modulated by the initial bilayer curvature and the lateral organization of membrane components (i.e. the presence of lipid domains). FIGURE 4: M1 layer interacting with the membrane is stable even after lipid removal. A-B: LSM confocal image of a typical GUV composed of DOPC:cholesterol:DOPS 50:20:30, after incubation with 5 µM M1-Alexa488 (green channel, panel A). The lipid bilayer is visualized via the addition of 0.05 mol% Rhodamine-DOPE (red channel, panel B). C-D: LSM confocal image of a typical GUV after treatment for 5 min with detergent (e.g. Triton X-100 1.7 mM). Panel C represent the signal from M1-Alexa488. Panel D represents the signal from Rhodamine-DOPE. The excitation laser power used to acquire the image shown in panel D was ~10 times higher than the power used to acquire the image shown in panel B. GUVs contained 150 mM sucrose in their lumen and were suspended in a phosphate buffered protein solution (pH 7.4) with similar osmolarity (see Materials and Methods). Scale bars are 5 µM. Images were acquired at 23°C.

Influenza A virus interactions with macrophages: Lessons from epithelial cells

“…M1 is then recruited to the site of assembly via interaction with the cytoplasmic tails of HA and NA, which then serves as a platform for vRNP binding (Zhang, Pekosz, & Lamb, ). Upon membrane binding, M1 undergoes oligomerization, eventually serving to provide structural rigidity to the envelope of complete virions and also contributing to the membrane deformation required for virus budding (Dahmani, Ludwig, & Chiantia, ; Hilsch et al, ). Additionally, M1 interacts with M2 allowing its recruitment to the periphery of the sites of budding, where the amphipathic helix of M2 can alter the curvature of the membrane, inducing endosomal sorting complexes required for transport (ESCRT) protein‐independent membrane scission (Rossman, Jing, Leser, & Lamb, ).…”