A cationic, C-terminal patch and structural rearrangements in Ebola virus matrix VP40 protein control its interactions with phosphatidylserine

Abstract

Ebola virus (EBOV) is a filamentous lipid-enveloped virus that causeshemorrhagic fever with a high fatality rate. Viral protein 40 (VP40) is the major EBOV matrix protein and regulates viral budding from the plasma membrane. VP40 is a transformer/morpheein that can structurally rearrange its native homodimer into either a hexameric filament that facilitates viral budding or a RNA-binding octameric ring that regulates viral transcription. VP40 associates with plasma-membrane lipids such as phosphatidylserine (PS), and this association is critical to budding from the host cell. However, it is poorly understood how different VP40 structures interact with PS, what essential residues are involved in this association, and whether VP40 has true selectivity for PS among different glycerophospholipid headgroups. In this study, we used lipid-binding assays, MD simulations, and cellular imaging to investigate the molecular basis of VP40-PS interactions and to determine whether different VP40 structures (i.e. monomer, dimer, and octamer) can interact with PScontaining membranes. Results from quantitative analysis indicated that VP40 associates with PS vesicles via a cationic patch in the C-terminal domain (Lys-224, 225 and Lys-274, 275). Substitutions of these residues with alanine reduced PSvesicle binding by >40-fold and abrogated VP40 localization to the plasma membrane. Dimeric VP40 had 2-fold greater affinity for PS-containing membranes than the monomer, whereas binding of the VP40 octameric ring was reduced by nearly 10-fold. Taken together, these results suggest http://www.jbc.org/cgi

Ebola Virus VP40 Modulates Cell Cycle and Biogenesis of Extracellular Vesicles

“…Previous studies have shown VP40 to be transiently present in the nucleus of infected cells at early time points [72][73][74]. Here, we have shown that VP40 is present at relatively high levels within the nucleus of stably transfected cells, and that VP40 is found on a large proportion of cyclin D1 promoters Figure 2C and D).…”

“…Based upon our observations of accelerated growth and increased levels of cyclin D1 in cells producing VP40, we next wished to investigate potential factors contributing to cyclin D1 transcription. Previous studies have shown the intriguing presence of EBOV VP40 within the nucleus, especially at early time points in infection [72][73][74]. To confirm VP40 presence in the nuclear compartment of our VP40 clones, cytoplasm and nuclear fractions of 293T and V2CH cells were isolated by NE-PER Nuclear and Cytoplasmic Extraction kit.…”

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

“…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].…”

“…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]. Though in general, eVP40 and mVP40 are thought to assemble at the plasma membrane inner leaflet in a similar capacity, fundamental differences in their membrane binding properties [8][9][10][11][12]14] as well as trafficking pathways to the plasma membrane [15,16] have been identified.…”

“…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].…”