scholarly journals Ebola Virus Requires Phosphatidylserine Scrambling Activity for Efficient Budding and Optimal Infectivity

Proceedings ◽  
2020 ◽  
Vol 50 (1) ◽  
pp. 35
Author(s):  
Marissa Acciani ◽  
Maria Lay ◽  
Katherine E. Havranek ◽  
Avery Duncan ◽  
Hersha Iyer ◽  
...  
Keyword(s):  
Ebola Virus ◽  
Apoptotic Cell ◽  
Viral Envelope ◽  
Host Cells ◽  
Surface Receptors ◽  
Outer Leaflet ◽  
Virus Like Particle ◽  
Vesicular Stomatitis ◽  
Virion Surface

Ebola virus (EBOV) interacts with cells using multiple categories of cell-surface receptors, including C-type lectins and phosphatidylserine (PS) receptors. PS receptors typically bind to apoptotic cell membrane PS and orchestrate the uptake and clearance of apoptotic bodies. Many viruses coated with PS-containing lipid envelopes, acquired during budding from host cells, can also exploit these receptors for internalization. PS is restricted to the inner leaflet of the plasma membrane in homeostatic cells, an orientation that would be unfavorable for PS receptor-mediated uptake if conserved on the viral envelope. Therefore, it is theorized that viral infection induces host-cell PS externalization to the outer leaflet during replication. Cells have several membrane scramblase enzymes that enrich outer leaflet PS when activated. Here, we investigate the role of two scramblases, TMEM16F and XKR8, as possible mediators of cellular and viral envelope surface PS levels during recombinant vesicular stomatitis virus (VSV) in which the VSV glycoprotein was replaced with the Ebola glycoprotein (rVSV/EBOV-GP) replication and EBOV virus-like particle (VLP) production. We find that rVSV/EBOV-GP and EBOV VLPs produced in XKR8 knockout cells contain two- to threefold less PS in their outer leaflets. Consequently, rVSV/EBOV-GP produced in deltaXKR8 is 70% less efficient at infecting cells through apoptotic mimicry as compared to the viruses produced by parental cells. In addition, the budding efficiency of both recombinant VSV particles and VLPs was significantly reduced in cells lacking XKR8. Our data suggest that virion surface PS acquisition requires XKR8 activity, whereas the deletion of TMEM16F did not affect EBOV-GP-mediated entry of VLP production. Unexpectedly, we observed an additional role of XKR8 in rVSV/G, rVSV/EBOV-GP, and EBOV VLP budding.

scholarly journals Ebola virus requires phosphatidylserine scrambling activity for efficient budding and optimal infectivity

2020 ◽  
Author(s):  
Marissa D. Acciani ◽  
Maria F. Lay-Mendoza ◽  
Katherine E. Havranek ◽  
Avery M. Duncan ◽  
Hersha Iyer ◽  
...  
Keyword(s):  
Host Cell ◽  
Particle Production ◽  
Ebola Virus ◽  
Apoptotic Cell ◽  
Viral Envelope ◽  
Host Cells ◽  
Target Cells ◽  
Surface Receptors ◽  
Envelope Surface ◽  
Outer Leaflet

AbstractEbola virus (EBOV) interacts with cells using two categories of cell surface receptors, C-type lectins and phosphatidylserine (PS) receptors. PS receptors typically bind to apoptotic cell membrane PS and orchestrate the uptake and clearance of apoptotic bodies. Many viruses coated with PS-containing lipid envelopes, acquired during budding from host cells, can also exploit these receptors for internalization. PS is restricted to the inner leaflet of the plasma membrane in homeostatic cells, an orientation that would be unfavorable for PS receptor-mediated uptake if conserved on the viral envelope. Therefore, it is theorized that viral infection induces host cell PS externalization to the outer leaflet during replication. Cells have several membrane scramblase enzymes that enrich outer leaflet PS when activated. Here, we investigate two scramblases, TMEM16F and XKR8, as possible mediators of cellular and viral envelope surface PS levels during recombinant VSV/EBOV-GP replication and EBOV virus-like particle (VLP) production. We found that rVSV/EBOV-GP and EBOV VLPs produced in XKR8 knockout cells contain decreased levels of PS in their outer leaflets. ΔXKR8-made rVSV/EBOV-GP is 70% less efficient at infecting cells through apoptotic mimicry compared to viruses made in parental cells. Our data suggest that virion surface PS acquisition requires XKR8 activity, whereas TMEM16F activity is not essential. Unexpectedly, we observed defective rVSV/G, rVSV/EBOV-GP, and EBOV VLP budding in ΔXKR8 cells, suggesting that phospholipid scrambling via XKR8 enhances both Ebola infectivity and budding efficiency. Overexpression of XKR8 dramatically increased budding activity, suggesting outer leaflet PS is required for both particle production and increased infectivity.ImportanceThe Democratic Republic of the Congo experienced its deadliest Ebola outbreak from 2018 to 2020, with 3,444 confirmed cases and 2,264 deaths (as of March 12, 2020). Owing to the extensive damage that these outbreaks have caused in Africa, as well as its future epidemic potential, Ebola virus (EBOV) ranks among the top eight priority pathogens outlined by the WHO in 2018. A comprehensive understanding of Ebola entry pathways into target cells is critical for antiviral development and outbreak control. Thus far, host-cell scramblases TMEM16F and XKR8 have each been named as the sole mediator of Ebola envelope surface phosphatidylserine (PS). We assessed the contributions of these proteins using CRISPR knockout cells and two EBOV models: rVSV/EBOV-GP and EBOV VLPs. We observed that XKR8 is required for optimal EBOV envelope PS levels, PS receptor engagement, and particle budding across all viral models, whereas TMEM16F did not play a major role.

scholarly journals Ebola virus requires phosphatidylserine scrambling activity for efficient budding and optimal infectivity

2021 ◽  
Author(s):  
Marissa D. Acciani ◽  
Maria F. Lay Mendoza ◽  
Katherine E. Havranek ◽  
Avery M. Duncan ◽  
Hersha Iyer ◽  
...  
Keyword(s):  
Ebola Virus ◽  
Apoptotic Cell ◽  
Transmembrane Protein ◽  
Central Africa ◽  
Viral Envelope ◽  
Cell Entry ◽  
Host Cells ◽  
Target Cells ◽  
Caspase Cleavage ◽  
Envelope Surface

Ebola virus (EBOV) attaches to target cells using two categories of cell surface receptors, C-type lectins and phosphatidylserine (PS) receptors. PS receptors typically bind to apoptotic cell membrane PS and orchestrate the uptake and clearance of apoptotic debris. Many enveloped viruses also contain exposed PS and can therefore exploit these receptors for cell entry. Viral infection can induce PS externalization in host cells, resulting in increased outer PS levels on budding virions. Scramblase enzymes carry out cellular PS externalization, thus, we targeted these proteins in order to manipulate viral envelope PS levels. We investigated two scramblases previously identified to be involved in EBOV PS levels, transmembrane protein 16F and Xk-related protein 8 (XKR8), as possible mediators of cellular and viral envelope surface PS levels during the replication of recombinant vesicular stomatitis virus containing its native glycoprotein (rVSV/G) or the EBOV glycoprotein (rVSV/EBOV-GP). We found that rVSV/G and rVSV/EBOV-GP virions produced in XKR8 knockout cells contain decreased levels of PS on their surfaces, and the PS-deficient rVSV/EBOV-GP virions are 70% less efficient at infecting cells through PS receptors. We also observed reduced rVSV and EBOV virus-like particle (VLP) budding in ΔXKR8 cells. Deleting XKR8 in HAP1 cells reduced rVSV/G and rVSV/EBOV-GP budding by 60% and 65% respectively, and reduced Ebola VLP budding more than 60%. We further demonstrated that caspase cleavage of XKR8 is required to promote budding. This suggests that XKR8, in addition to mediating virion PS levels, may also be critical for enveloped virus budding at the plasma membrane. Importance Within the last decade, countries in western and central Africa have experienced the most widespread and deadly Ebola outbreaks since the virus was identified in 1976. While outbreaks are primarily attributed to zoonotic transfer events, new evidence is emerging that outbreaks may be caused by a combination of spillover events and viral latency or persistence in survivors. The possibility that Ebola can remain dormant then re-emerge in survivors highlights the critical need to prevent the virus from entering and establishing infection in human cells. Thus far, host-cell scramblases TMEM16F and XKR8 have been implicated in Ebola envelope surface phosphatidylserine (PS) and cell entry using PS receptors. We assessed the contributions of these proteins using CRISPR knockout cells and two EBOV models: rVSV/EBOV-GP and EBOV VLPs. We observed that XKR8 is required for optimal EBOV envelope PS levels and infectivity, and particle budding across all viral models.

scholarly journals Viral Hepatitis and Iron Dysregulation: Molecular Pathways and the Role of Lactoferrin

Molecules ◽  
2020 ◽  
Vol 25 (8) ◽  
pp. 1997 ◽  
Author(s):  
Romina Mancinelli ◽  
Luigi Rosa ◽  
Antimo Cutone ◽  
Maria Stefania Lepanto ◽  
Antonio Franchitto ◽  
...  
Keyword(s):  
Viral Hepatitis ◽  
Viral Entry ◽  
Iron Homeostasis ◽  
Iron Chelation ◽  
Host Cells ◽  
Surface Receptors ◽  
Inflammatory Processes ◽  
Related Proteins ◽  
Binding Glycoprotein

The liver is a frontline immune site specifically designed to check and detect potential pathogens from the bloodstream to maintain a general state of immune hyporesponsiveness. One of the main functions of the liver is the regulation of iron homeostasis. The liver detects changes in systemic iron requirements and can regulate its concentration. Pathological states lead to the dysregulation of iron homeostasis which, in turn, can promote infectious and inflammatory processes. In this context, hepatic viruses deviate hepatocytes’ iron metabolism in order to better replicate. Indeed, some viruses are able to alter the expression of iron-related proteins or exploit host receptors to enter inside host cells. Lactoferrin (Lf), a multifunctional iron-binding glycoprotein belonging to the innate immunity, is endowed with potent antiviral activity, mainly related to its ability to block viral entry into host cells by interacting with viral and/or cell surface receptors. Moreover, Lf can act as an iron scavenger by both direct iron-chelation or the modulation of the main iron-related proteins. In this review, the complex interplay between viral hepatitis, iron homeostasis, and inflammation as well as the role of Lf are outlined.

scholarly journals Apoptotic Killing and Phagocytosis of Host Cells by the Parasite Entamoeba histolytica

2003 ◽  
Vol 71 (2) ◽  
pp. 964-972 ◽  
Author(s):  
Christopher D. Huston ◽  
Douglas R. Boettner ◽  
Vanessa Miller-Sims ◽  
William A. Petri,
Keyword(s):  
Host Cell ◽  
Entamoeba Histolytica ◽  
Caspase 3 ◽  
Apoptotic Cell ◽  
Cell Killing ◽  
Jurkat Cells ◽  
Host Cells ◽  
Apoptotic Cells ◽  
Outer Leaflet ◽  
Phosphatidylserine Exposure

ABSTRACT The ability of Entamoeba histolytica to kill and phagocytose host cells correlates with parasite virulence. This study addressed the role of apoptotic cell killing and host cell phosphatidylserine exposure in the subsequent phagocytosis of Jurkat T cells by E. histolytica. Ingested host cells were apoptotic, as evidenced by the activation of caspase 3 in 88% ± 3% (mean and standard deviation [SD] of the mean) of Jurkat cells engulfed by E. histolytica; ingested cells without detectable active caspase 3 were already disrupted and partially digested. That apoptotic cell killing preceded phagocytosis was supported by the demonstration that a higher percentage of amebae ingested apoptotic cells than ingested healthy cells (62% ± 7% versus 30% ± 9%, respectively [mean and SD]) (P = 0.008). E. histolytica also ingested apoptotic Jurkat cells more rapidly than necrotic control cells (8.5% ± 0.4% versus 3.5% ± 0.7%, respectively [mean and SD]) (P < 0.001). The inhibition of amebic cytotoxicity with d-galactose (which blocks the amebic Gal/GalNAc lectin) blocked the phagocytosis of healthy cells by greater than 80%, providing further evidence that apoptosis preceded engulfment. In contrast, d-galactose blocked the phagocytosis of already apoptotic cells by only 40%, implicating an additional host ligand (besides d-galactose) in amebic engulfment of apoptotic cells. The most characteristic surface change on apoptotic cells is phosphatidylserine exposure. Consistent with a role for host cell phosphatidylserine exposure in amebic ingestion of killed cells, Jurkat cell phosphatidylserine was exposed during incubation with E. histolytica (27% ± 1% [mean and SD] specific increase at 30 min) (the P value versus the control was 0.0003). Approximately 50% more amebae ingested viable Jurkat cells expressing phosphatidylserine on the outer leaflet of the plasma membrane than ingested control cells (30.3% ± 2.2% versus 19.8% ± 1.9%, respectively [mean and SD]) (P = 0.003). By analogy with phagocytic clearance during apoptosis in metazoans, amebic apoptotic host cell killing followed by phagocytosis may limit inflammation and enable amebae to evade the host immune response.

scholarly journals Detection of Cell-Cell Fusion Mediated by Ebola Virus Glycoproteins

2006 ◽  
Vol 80 (6) ◽  
pp. 2815-2822 ◽  
Author(s):  
Séverine Bär ◽  
Ayato Takada ◽  
Yoshihiro Kawaoka ◽  
Marc Alizon
Keyword(s):  
Membrane Fusion ◽  
Viral Entry ◽  
Ebola Virus ◽  
Low Ph ◽  
Viral Envelope ◽  
Target Cells ◽  
Direct Role ◽  
Influenza Virus Hemagglutinin ◽  
Ph Dependent

ABSTRACT Ebola viruses (EboV) are enveloped RNA viruses infecting cells by a pH-dependent process mediated by viral glycoproteins (GP) involving endocytosis of virions and their routing into acidic endosomes. As with well-characterized pH-dependent viral entry proteins, in particular influenza virus hemagglutinin, it is thought that EboV GP require activation by low pH in order to mediate fusion of the viral envelope with the membrane of endosomes. However, it has not yet been possible to confirm the direct role of EboV GP in membrane fusion and the requirement for low-pH activation. It was in particular not possible to induce formation of syncytia by exposing cells expressing EboV GP to acidic medium. Here, we have used an assay based on the induction of a β-galactosidase (lacZ) reporter gene in target cells to detect cytoplasmic exchanges, indicating membrane fusion, with cells expressing EboV GP (Zaire species). Acidic activation of GP-expressing cells was required for efficient fusion with target cells. The direct role of EboV GP in this process is indicated by its inhibition by anti-GP antibodies and by the lack of activity of mutant GP normally expressed at the cell surface but defective for virus entry. Fusion was not observed when target cells underwent acidic treatment, for example, when they were placed in coculture with GP-expressing cells before the activation step. This unexpected feature, possibly related to the nature of the EboV receptor, could explain the impossibility of inducing formation of syncytia among GP-expressing cells.

The variations of virus shape and size in different preparations

1990 ◽  
Vol 48 (3) ◽  
pp. 580-581
Author(s):  
Ding Mingxiao ◽  
Jiao Renjie ◽  
Liang Fengxia ◽  
Zhai Zhonghe
Keyword(s):  
Sindbis Virus ◽  
Viral Envelope ◽  
Host Cells ◽  
Enveloped Viruses ◽  
Viral Attachment ◽  
Cytopathic Effects ◽  
Surface Replica ◽  
Freeze Etching ◽  
Vesicular Stomatitis ◽  
Important Structure

Envelope is a very important structure for viral attachment and entry into the host cell, but it is also a morphologically variable portion of enveloped viruses. Studying the fine structure of enveloped viruses, we noticed that different sample preparations of viruses resulted in the change of viral size and shape to some extent, which we believe was caused by the variation of the viral envelope. Four typical enveloped viruses: IBRV (Infectious Bovine Rhinotracheitis Virus), GPV (Goat Pox Virus), SbV (Sindbis Virus) and VSV (Vesicular Stomatitis Virus) were investigated in our experiments.Host cells infected with IBRV, GPV, SbV and VSV respectively were fixed with 1-5% glutaraldehyde in Hank's buffer when the cytopathic effects appeared in 50-70% of the cells, then the specimens were treated respectively with different conventional methods of EM sample preparation: 1) ultrathin sectioning, 2) negative staining,3) freeze etching, 4) surface replica, 5) whole mount or SEM observations. All the samples were examined under JEM-200CX TEM or JSM-35CF SEM.

scholarly journals Inflammation, Extracellular Matrix Remodeling, and Proteostasis in Tumor Microenvironment

2021 ◽  
Vol 22 (15) ◽  
pp. 8102
Author(s):  
Marina Marozzi ◽  
Arianna Parnigoni ◽  
Aide Negri ◽  
Manuela Viola ◽  
Davide Vigetti ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Tumor Microenvironment ◽  
Signaling Pathways ◽  
Intracellular Signaling ◽  
Host Cells ◽  
Inflammatory Factors ◽  
Matrix Remodeling ◽  
Surface Receptors ◽  
Degrading Enzymes

Cancer is a multifaceted and complex pathology characterized by uncontrolled cell proliferation and decreased apoptosis. Most cancers are recognized by an inflammatory environment rich in a myriad of factors produced by immune infiltrate cells that induce host cells to differentiate and to produce a matrix that is more favorable to tumor cells’ survival and metastasis. As a result, the extracellular matrix (ECM) is changed in terms of macromolecules content, degrading enzymes, and proteins. Altered ECM components, derived from remodeling processes, interact with a variety of surface receptors triggering intracellular signaling that, in turn, cancer cells exploit to their own benefit. This review aims to present the role of different aspects of ECM components in the tumor microenvironment. Particularly, we highlight the effect of pro- and inflammatory factors on ECM degrading enzymes, such as metalloproteases, and in a more detailed manner on hyaluronan metabolism and the signaling pathways triggered by the binding of hyaluronan with its receptors. In addition, we sought to explore the role of extracellular chaperones, especially of clusterin which is one of the most prominent in the extracellular space, in proteostasis and signaling transduction in the tumor microenvironment. Although the described tumor microenvironment components have different biological roles, they may engage common signaling pathways that favor tumor growth and metastasis.

scholarly journals The role of phosphatidylserine on the membrane in immunity and blood coagulation

2022 ◽  
Vol 10 (1) ◽  
Author(s):  
Jiao Wang ◽  
Changxin Yu ◽  
Junyi Zhuang ◽  
Wenxin Qi ◽  
Jiawen Jiang ◽  
...  
Keyword(s):  
Blood Coagulation ◽  
Lupus Erythematosus ◽  
Ebola Virus ◽  
Virus Disease ◽  
Interaction Mechanism ◽  
Coagulation Factors ◽  
Host Cells ◽  
Outer Leaflet ◽  
Enhanced Production ◽  
Immunodeficiency Virus

AbstractThe negatively charged aminophospholipid, phosphatidylserine (PtdSer), is located in the inner leaflet of the plasma membrane in normal cells, and may be exposed to the outer leaflet under some immune and blood coagulation processes. Meanwhile, Ptdser exposed to apoptotic cells can be recognized and eliminated by various immune cells, whereas on the surface of activated platelets Ptdser interacts with coagulation factors prompting enhanced production of thrombin which significantly facilitates blood coagulation. In the case where PtdSer fails in exposure or mistakenly occurs, there are occurrences of certain immunological and haematological diseases, such as the Scott syndrome and Systemic lupus erythematosus. Besides, viruses (e.g., Human Immunodeficiency Virus (HIV), Ebola virus (EBOV)) can invade host cells through binding the exposed PtdSer. Most recently, the Corona Virus Disease 2019 (COVID-19) has been similarly linked to PtdSer or its receptors. Therefore, it is essential to comprehensively understand PtdSer and its functional characteristics. Therefore, this review summarizes Ptdser, its eversion mechanism; interaction mechanism, particularly with its immune receptors and coagulation factors; recognition sites; and its function in immune and blood processes. This review illustrates the potential aspects for the underlying pathogenic mechanism of PtdSer-related diseases, and the discovery of new therapeutic strategies as well.

scholarly journals Enterohemorrhagic Escherichia coliInduces Apoptosis Which Augments Bacterial Binding and Phosphatidylethanolamine Exposure on the Plasma Membrane Outer Leaflet

2000 ◽  
Vol 68 (6) ◽  
pp. 3108-3115 ◽  
Author(s):  
Debora Barnett Foster ◽  
Maan Abul-Milh ◽  
Mario Huesca ◽  
Clifford A. Lingwood
Keyword(s):  
Cell Death ◽  
Apoptotic Cell ◽  
Morphological Changes ◽  
Bacterial Attachment ◽  
Host Cells ◽  
Watery Diarrhea ◽  
Similar Treatment ◽  
Outer Leaflet ◽  
Gastrointestinal Pathogen ◽  
Induction Of Apoptosis

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) is a gastrointestinal pathogen that causes watery diarrhea and hemorrhagic colitis and can lead to serious and even fatal complications such as hemolytic uremic syndrome. We investigated the ability of EHEC to kill host cells using three human epithelial cell lines. Analysis of phosphatidylserine expression, internucleosomal cleavage of host cell DNA and morphological changes detected by electron microscopy changes revealed evidence of apoptotic cell death. The rates and extents of cell death were similar for both verotoxin-producing and nonproducing strains of EHEC as well as for a related gastrointestinal pathogen, enteropathogenic E. coli (EPEC). The induction of apoptosis by bacterial attachment was independent of verotoxin production and greater than that produced by a similar treatment with verotoxin alone. Expression of phosphatidylethanolamine, previously reported to bind EHEC and EPEC, was also increased on apoptotic cells but with little correlation to phosphatidylserine expression. Phosphatidylethanolamine levels but not phosphatidylserine levels on dying cells correlated with EHEC binding. Cells treated with phosphatidylethanolamine-containing liposomes also showed increased EHEC binding. These results suggest that bacterial induction of apoptosis offers an advantage for bacterial attachment by augmenting outer leaflet levels of the phosphatidylethanolamine receptor.

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