The Ebola virus matrix protein clusters phosphatidylserine, a critical step in viral budding

Abstract

Phosphatidylserine (PS) has been shown to be a critical lipid factor in the assembly and spread of numerous lipid enveloped viruses. Here, we describe the ability of the Ebola virus (EBOV) matrix protein eVP40 to induce clustering of PS and promote viral budding in vitro, as well as the ability of an FDA approved drug, fendiline, to reduce PS clustering subsequently reducing virus budding and entry. To gain mechanistic insight into fendiline inhibition of EBOV replication, multiple in vitro assays were employed including imaging, viral budding and viral entry assays. Fendiline reduced the PS content in mammalian cells and PS in the plasma membrane, reducing the ability of VP40 to form new virus particles. Further, particles that do form from fendiline treated cells have altered particle morphology and decreased infectivity capacity. These complementary studies reveal the mechanism by which filovirus matrix proteins cluster PS to enhance viral assembly, budding, and spread from the host cell while also laying the groundwork for fundamental drug targeting strategies.

Mechanisms of phosphatidylserine influence on viral production: A computational model of Ebola virus matrix protein assembly

“…Our initial analysis aims to determine if the known impact of PS on VP40 membrane association is sufficient to produce the observed changes in VP40 oligomer levels and VLP production ( 34 , 35 ). Our “preliminary” model ( Table 1 ) therefore only includes the known influence of PS level on the dissociation constant (K D2 ) of VP40 dimer binding to cell membrane ( 32 , 34 ).…”

“…All of our models assume that VP40 hexamers are the building units of VP40 assembly ( 27 , 28 , 34 , 35 , 49 , 51 , 52 , 53 ). However, recent work indicates that the building block for VP40 VLPs can be dimers instead of hexamers ( 33 ).…”

“…Membrane PS levels have been shown to affect VP40 membrane binding ( 34 ), as well as impact the relative number of VP40 oligomers and VLPs produced over 48 h ( 34 , 35 ). However, it remains unclear if the impact of PS on VP40 membrane association is able to account for the observed impacts on VP40 oligomer levels and VLP production or if additional PS-dependent mechanisms exist ( Fig.…”

“…Our model incorporates VP40 dynamics spanning VP40 monomer production to VLP budding. PS is known to affect VP40 membrane association (k 2 , black dashed line ), but it remains unclear if this is enough to explain experimentally observed differences in VLP filament growth profiles and VLP production under different PS concentrations ( 34 , 35 ). In this work, we test if PS impacts on VP40 membrane association are sufficient to explain differences in filament growth and VLP production using our “preliminary” model.…”

“…In contrast, subcellular models for EBOV assembly are not currently available. However, emerging experimental data and a more detailed understanding of the EBOV replication cycle ( 34 , 35 ) create an opportunity for a quantitative, computational characterization of EBOV assembly.…”