Browsing by Subject "Phosphatidylserine"

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    Detection of lipid-induced structural changes of the Marburg virus matrix protein VP40 using hydrogen/deuterium exchange-mass spectrometry
    (American Society for Biochemistry and Molecular Biology, 2017-04-14) Wijesinghe, Kaveesha J.; Urata, Sarah; Bhattarai, Nisha; Kooijman, Edgar E.; Gerstman, Bernard S.; Chapagain, Prem P.; Li, Sheng; Stahelin, Robert V.; Biochemistry and Molecular Biology, School of Medicine
    Marburg virus (MARV) is a lipid-enveloped virus from the Filoviridae family containing a negative sense RNA genome. One of the seven MARV genes encodes the matrix protein VP40, which forms a matrix layer beneath the plasma membrane inner leaflet to facilitate budding from the host cell. MARV VP40 (mVP40) has been shown to be a dimeric peripheral protein with a broad and flat basic surface that can associate with anionic phospholipids such as phosphatidylserine. Although a number of mVP40 cationic residues have been shown to facilitate binding to membranes containing anionic lipids, much less is known on how mVP40 assembles to form the matrix layer following membrane binding. Here we have used hydrogen/deuterium exchange (HDX) mass spectrometry to determine the solvent accessibility of mVP40 residues in the absence and presence of phosphatidylserine and phosphatidylinositol 4,5-bisphosphate. HDX analysis demonstrates that two basic loops in the mVP40 C-terminal domain make important contributions to anionic membrane binding and also reveals a potential oligomerization interface in the C-terminal domain as well as a conserved oligomerization interface in the mVP40 N-terminal domain. Lipid binding assays confirm the role of the two basic patches elucidated with HD/X measurements, whereas molecular dynamics simulations and membrane insertion measurements complement these studies to demonstrate that mVP40 does not appreciably insert into the hydrocarbon region of anionic membranes in contrast to the matrix protein from Ebola virus. Taken together, we propose a model by which association of the mVP40 dimer with the anionic plasma membrane facilitates assembly of mVP40 oligomers.
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    Membrane binding and bending in Ebola VP40 assembly and egress
    (Frontiers Media, 2014-06-18) Stahelin, Robert V.; Biochemistry and Molecular Biology, School of Medicine
    Lipid-enveloped viruses contain a lipid bilayer coat that protects their genome and helps to facilitate entry into the host cell. Filoviruses are lipid-enveloped viruses that have up to 90% clinical fatality and include Marbug (MARV) and Ebola (EBOV). These pleomorphic filamentous viruses enter the host cell through their membrane-embedded glycoprotein and then replicate using just seven genes encoded in their negative-sense RNA genome. EBOV budding occurs from the inner leaflet of the plasma membrane (PM) and is driven by the matrix protein VP40, which is the most abundantly expressed protein of the virus. VP40 expressed in mammalian cells alone can trigger budding of filamentous virus-like particles (VLPs) that are nearly indistinguishable from authentic EBOV. VP40, such as matrix proteins from other viruses, has been shown to bind anionic lipid membranes. However, how VP40 selectively interacts with the inner leaflet of the PM and assembles into a filamentous lipid enveloped particle is mostly unknown. This article describes what is known regarding VP40 membrane interactions and what answers will fill the gaps.
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    Phosphatidylserine externalization by apoptotic cells is dispensable for specific recognition leading to innate apoptotic immune responses
    (American Society for Biochemistry and Molecular Biology, 2022) Gomes, Marta T.; Palasiewicz, Karol; Gadiyar, Varsha; Lahey, Kevin; Calianese, David; Birge, Raymond B.; Ucker, David S.; Medicine, School of Medicine
    Surface determinants newly expressed by apoptotic cells that are involved in triggering potent immunosuppressive responses, referred to as "innate apoptotic immunity (IAI)" have not been characterized fully. It is widely assumed, often implicitly, that phosphatidylserine, a phospholipid normally cloistered in the inner leaflet of cells and externalized specifically during apoptosis, is involved in triggering IAI, just as it plays an essential role in the phagocytic recognition of apoptotic cells. It is notable, however, that the triggering of IAI in responder cells is not dependent on the engulfment of apoptotic cells by those responders. Contact between the responder and the apoptotic target, on the other hand, is necessary to elicit IAI. Previously, we demonstrated that exposure of protease-sensitive determinants on the apoptotic cell surface are essential for initiating IAI responses; exposed glycolytic enzyme molecules were implicated in particular. Here, we report our analysis of the involvement of externalized phosphatidylserine in triggering IAI. To analyze the role of phosphatidylserine, we employed a panel of target cells that either externalized phosphatidylserine constitutively, independently of apoptosis, or did not, as well as their WT parental cells that externalized the phospholipid in an apoptosis-dependent manner. We found that the externalization of phosphatidylserine, which can be fully uncoupled from apoptosis, is neither sufficient nor necessary to trigger the profound immunomodulatory effects of IAI. These results reinforce the view that apoptotic immunomodulation and phagocytosis are dissociable and further underscore the significance of protein determinants localized to the cell surface during apoptosis in triggering innate apoptotic immunity.