Investigation of the Lipid Binding Properties of the Marburg Virus Matrix Protein VP40

Abstract

Marburg virus (MARV), which belongs to the virus family Filoviridae, causes hemorrhagic fever in humans and nonhuman primates that is often fatal. MARV is a lipid-enveloped virus that during the replication process extracts its lipid coat from the plasma membrane of the host cell it infects. MARV carries seven genes, one of which encodes its matrix protein VP40 (mVP40), which regulates the assembly and budding of the virions. Currently, little information is available on mVP40 lipid binding properties. Here, we have investigated the in vitro and cellular mechanisms by which mVP40 associates with lipid membranes. mVP40 associates with anionic membranes in a nonspecific manner that is dependent upon the anionic charge density of the membrane. These results are consistent with recent structural determination of mVP40, which elucidated an mVP40 dimer with a flat and extensive cationic lipid binding interface. IMPORTANCEMarburg virus (MARV) is a lipid-enveloped filamentous virus from the family Filoviridae. MARV was discovered in 1967, and yet little is known about how its seven genes are used to assemble and form a new viral particle in the host cell it infects. The MARV matrix protein VP40 (mVP40) underlies the inner leaflet of the virus and regulates budding from the host cell plasma membrane. In vitro and cellular assays in this study investigated the mechanism by which mVP40 associates with lipids. The results demonstrate that mVP40 interactions with lipid vesicles or the inner leaflet of the plasma membrane are electrostatic but nonspecific in nature and are dependent on the anionic charge density of the membrane surface. Small molecules that can disrupt lipid trafficking or reduce the anionic charge of the plasma membrane interface may be useful in inhibiting assembly and budding of MARV. Marburg virus (MARV) and Ebola virus (EBOV) are members of the virus family Filoviridae that are characterized by their filamentous lipid -enveloped morphology (1). Electron microscopy studies revealed that MARV particles have a uniform diameter of approximately 80 nm, with an average particle length of 740 nm (2, 3). MARV harbors a negative-sense RNA genome 19.1 kb in length that includes seven genes (4). The glycoprotein (GP) is a transmembrane protein present in the viral envelope, and it mediates viral entry into the host cell. VP40 is the major matrix protein (mVP40) and completely underlies the lipid envelope of the virus. VP40 alone is sufficient to produce virus-like particles (VLPs) from mammalian cells that are nearly indistinguishable from virions (5-8). In addition to interacting with the host cell membrane and mediating budding, VP40 interacts with the nucleocapsid, which consists of nucleoprotein, VP24, VP30, VP35, and the L protein (9). VP40 also plays other essential roles, such as immunosuppression (10), regulation of viral transcription, and/or replication (11,12), and is also able to interact with GP, coexpression of which leads to GP enrichment at sites of mVP40 budding (13). mVP40 is a peripheral ...

Mutation of Hydrophobic Residues in the C-Terminal Domain of the Marburg Virus Matrix Protein VP40 Disrupts Trafficking to the Plasma Membrane

“…mVP40 protein and L201A harboring a 6x-His Tag were expressed and purified as described in Wijesinghe K. J., et al (2016) [10].…”

“…A multilamellar vesicle (MLV) sedimentation assay was performed as previously described in in Wijesinghe K. J., et al (2016) [10] and Oda S., et al (2016) [8].…”

“…Expression of either eVP40 or mVP40 in mammalian cells, in the absence of other filovirus proteins, led to the formation of filamentous virus-like particles (VLPs) that resemble the authentic virions [4][5][6][7]. eVP40 and mVP40 form dimers [8,9] using an amino-terminal domain (NTD) -helical interface, where the dimers are thought to be building blocks for large VP40 oligomers that form at the plasma membrane and are necessary for budding [8][9][10][11][12][13][14]. Notably, mutation of the dimer interface of eVP40 or mVP40 abrogated VLP formation and significantly reduced VP40 plasma membrane localization [8,9,14].…”

“…Previous studies have demonstrated that interaction of mVP40 with the plasma membrane depends on electrostatic interactions between mVP40 basic residues and the anionic charge of lipids such as phosphatidylserine (PS) and PI (4,5)P2. mVP40 interacts with a broad range of anionic phospholipids functioning as an anionic charge sensor [8,10,12]. In contrast, eVP40 primarily interacts with the plasma membrane displaying selectivity for phosphatidylserine (PS) [9,10,14,17]and phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2) [18].…”

“…mVP40 interacts with a broad range of anionic phospholipids functioning as an anionic charge sensor [8,10,12]. In contrast, eVP40 primarily interacts with the plasma membrane displaying selectivity for phosphatidylserine (PS) [9,10,14,17]and phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2) [18]. Further, eVP40 was also shown to penetrate membranes containing PS using a hydrophobic loop region at the (CTD) supplementing electrostatic interactions during membrane association [19,20].…”