scholarly journals Reporter assays for Ebola virus nucleoprotein oligomerization, virion-like particle budding, and minigenome activity reveal the importance of nucleoprotein amino acid position 111

2018 ◽  
Author(s):  
Aaron E. Lin ◽  
William E. Diehl ◽  
Yingyun Cai ◽  
Courtney L. Finch ◽  
Chidiebere Akusobi ◽  
...  
Keyword(s):  
Ebola Virus ◽  
Virus Disease ◽  
Salt Bridge ◽  
Amino Acid Position ◽  
Live Cells ◽  
Highly Pathogenic ◽  
Pathogenic Viruses ◽  
Reporter Assays ◽  
Neighboring Residue ◽  
Transcription And Replication

AbstractFor highly pathogenic viruses, reporter assays that can be rapidly performed are critically needed to identify potentially functional mutations for further study under maximal containment (e.g., biosafety level 4 [BSL-4]). The Ebola virus nucleoprotein (NP) plays multiple essential roles during the viral life cycle, yet few tools exist to study the protein under BSL-2 or equivalent containment. Therefore, we adapted reporter assays to measure NP oligomerization and virion-like particle (VLP) production in live cells and further measure transcription and replication using established minigenome assays. As a proof-of-concept, we examined the NP-R111C substitution, which emerged during the 2013–2016 Western African Ebola virus disease epidemic and rose to high frequency. NP-R111C slightly increased NP oligomerization and VLP budding but slightly decreased transcription and replication. By contrast, a synthetic charge-reversal mutant, NP-R111E, greatly increased oligomerization but abrogated transcription and replication. These results are intriguing in light of recent structures of NP oligomers, which reveal that the neighboring residue, K110, forms a salt bridge with E349 on adjacent NP molecules. By developing and utilizing multiple reporter assays, we find that the NP-111 position mediates a complex interplay between NP’s roles in protein structure, virion budding, and transcription and replication.

scholarly journals Reporter Assays for Ebola Virus Nucleoprotein Oligomerization, Virion-Like Particle Budding, and Minigenome Activity Reveal the Importance of Nucleoprotein Amino Acid Position 111

Viruses ◽  
2020 ◽  
Vol 12 (1) ◽  
pp. 105
Author(s):  
Aaron E. Lin ◽  
William E. Diehl ◽  
Yingyun Cai ◽  
Courtney L. Finch ◽  
Chidiebere Akusobi ◽  
...  
Keyword(s):  
Ebola Virus ◽  
Virus Disease ◽  
Salt Bridge ◽  
Amino Acid Position ◽  
Live Cells ◽  
Highly Pathogenic ◽  
Pathogenic Viruses ◽  
Reporter Assays ◽  
Neighboring Residue ◽  
Transcription And Replication

For highly pathogenic viruses, reporter assays that can be rapidly performed are critically needed to identify potentially functional mutations for further study under maximal containment (e.g., biosafety level 4 [BSL-4]). The Ebola virus nucleoprotein (NP) plays multiple essential roles during the viral life cycle, yet few tools exist to study the protein under BSL-2 or equivalent containment. Therefore, we adapted reporter assays to measure NP oligomerization and virion-like particle (VLP) production in live cells and further measured transcription and replication using established minigenome assays. As a proof-of-concept, we examined the NP-R111C substitution, which emerged during the 2013–2016 Western African Ebola virus disease epidemic and rose to high frequency. NP-R111C slightly increased NP oligomerization and VLP budding but slightly decreased transcription and replication. By contrast, a synthetic charge-reversal mutant, NP-R111E, greatly increased oligomerization but abrogated transcription and replication. These results are intriguing in light of recent structures of NP oligomers, which reveal that the neighboring residue, K110, forms a salt bridge with E349 on adjacent NP molecules. By developing and utilizing multiple reporter assays, we find that the NP-111 position mediates a complex interplay between NP’s roles in protein structure, virion budding, and transcription and replication.

scholarly journals SARS-CoV-2 Cellular Infection and Therapeutic Opportunities: Lessons Learned from Ebola Virus

Membranes ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 64
Author(s):  
Jordana Muñoz-Basagoiti ◽  
Daniel Perez-Zsolt ◽  
Jorge Carrillo ◽  
Julià Blanco ◽  
Bonaventura Clotet ◽  
...  
Keyword(s):  
Viral Entry ◽  
Ebola Virus ◽  
Lessons Learned ◽  
Productive Infection ◽  
Target Cells ◽  
Emerging Viruses ◽  
Highly Pathogenic ◽  
Life Threatening ◽  
Pathogenic Viruses ◽  
Cellular Machinery

Viruses rely on the cellular machinery to replicate and propagate within newly infected individuals. Thus, viral entry into the host cell sets up the stage for productive infection and disease progression. Different viruses exploit distinct cellular receptors for viral entry; however, numerous viral internalization mechanisms are shared by very diverse viral families. Such is the case of Ebola virus (EBOV), which belongs to the filoviridae family, and the recently emerged coronavirus SARS-CoV-2. These two highly pathogenic viruses can exploit very similar endocytic routes to productively infect target cells. This convergence has sped up the experimental assessment of clinical therapies against SARS-CoV-2 previously found to be effective for EBOV, and facilitated their expedited clinical testing. Here we review how the viral entry processes and subsequent replication and egress strategies of EBOV and SARS-CoV-2 can overlap, and how our previous knowledge on antivirals, antibodies, and vaccines against EBOV has boosted the search for effective countermeasures against the new coronavirus. As preparedness is key to contain forthcoming pandemics, lessons learned over the years by combating life-threatening viruses should help us to quickly deploy effective tools against novel emerging viruses.

scholarly journals Favipiravir in Therapy of Viral Infections

2021 ◽  
Vol 10 (2) ◽  
pp. 273
Author(s):  
Ryta Łagocka ◽  
Violetta Dziedziejko ◽  
Patrycja Kłos ◽  
Andrzej Pawlik
Keyword(s):  
Ebola Virus ◽  
Viral Infections ◽  
Virus Disease ◽  
Antiviral Drug ◽  
Competitive Inhibitor ◽  
Mechanisms Of Action ◽  
Pharmacokinetic Parameters ◽  
Rna Dependent Rna Polymerase ◽  
Antiviral Therapies ◽  
Transcription And Replication

Favipiravir (FPV) is a novel antiviral drug acting as a competitive inhibitor of RNA-dependent RNA polymerase (RdRp), preventing viral transcription and replication. FPV was approved in Japan in 2014 for therapy of influenza unresponsive to standard antiviral therapies. FPV was also used in the therapy of Ebola virus disease (EVD) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. In this review, we discuss the mechanisms of action, pharmacokinetic parameters, toxicity, and adverse effects of FPV, as well as clinical studies evaluating the use of FPV in the therapy of influenza virus (IV) infection, EVD, and SARS-CoV-2 infection, along with its effectiveness in treating other human RNA infections.

scholarly journals The Cytokine Response Profile of Ebola Virus Disease in a Large Cohort of Rhesus Macaques Treated With Monoclonal Antibodies

2019 ◽  
Vol 6 (3) ◽  
Author(s):  
Logan Banadyga ◽  
Vinayakumar Siragam ◽  
Wenjun Zhu ◽  
Shihua He ◽  
Keding Cheng ◽  
...  
Keyword(s):  
Immune Response ◽  
Ebola Virus ◽  
Virus Disease ◽  
Rhesus Macaques ◽  
Hemorrhagic Fever ◽  
Ebola Virus Disease ◽  
Antibody Treatment ◽  
Highly Pathogenic ◽  
Cytokine Levels ◽  
Response Profile

Abstract Ebola virus (EBOV) is a highly pathogenic filovirus that causes outbreaks of a severe hemorrhagic fever known as EBOV disease (EVD). Ebola virus disease is characterized in part by a dysregulated immune response and massive production of both pro- and anti-inflammatory cytokines. To better understand the immune response elicited by EVD in the context of treatment with experimental anti-EBOV antibody cocktails, we analyzed 29 cytokines in 42 EBOV-infected rhesus macaques. In comparison to the surviving treated animals, which exhibited minimal aberrations in only a few cytokine levels, nonsurviving animals exhibited a dramatically upregulated inflammatory response that was delayed by antibody treatment.

scholarly journals Protection of Nonhuman Primates against Two Species of Ebola Virus Infection with a Single Complex Adenovirus Vector

2010 ◽  
Vol 17 (4) ◽  
pp. 572-581 ◽  
Author(s):  
William D. Pratt ◽  
Danher Wang ◽  
Donald K. Nichols ◽  
Min Luo ◽  
Jan Woraratanadharm ◽  
...  
Keyword(s):  
Nonhuman Primates ◽  
Ebola Virus ◽  
Adenovirus Vector ◽  
Hemorrhagic Fever ◽  
Lethal Challenge ◽  
Highly Pathogenic ◽  
Pathogenic Viruses ◽  
Single Complex ◽  
Zaire Ebolavirus ◽  
Ebola Virus Infection

ABSTRACT Ebola viruses are highly pathogenic viruses that cause outbreaks of hemorrhagic fever in humans and other primates. To meet the need for a vaccine against the several types of Ebola viruses that cause human diseases, we developed a multivalent vaccine candidate (EBO7) that expresses the glycoproteins of Zaire ebolavirus (ZEBOV) and Sudan ebolavirus (SEBOV) in a single complex adenovirus-based vector (CAdVax). We evaluated our vaccine in nonhuman primates against the parenteral and aerosol routes of lethal challenge. EBO7 vaccine provided protection against both Ebola viruses by either route of infection. Significantly, protection against SEBOV given as an aerosol challenge, which has not previously been shown, could be achieved with a boosting vaccination. These results demonstrate the feasibility of creating a robust, multivalent Ebola virus vaccine that would be effective in the event of a natural virus outbreak or biological threat.

scholarly journals A glycoprotein mutation that emerged during the 2013-2016 Ebola virus epidemic alters proteolysis and accelerates membrane fusion

2020 ◽  
Author(s):  
J. Maximilian Fels ◽  
Robert H. Bortz ◽  
Tanwee Alkutkar ◽  
Eva Mittler ◽  
Rohit K. Jangra ◽  
...  
Keyword(s):  
Cell Culture ◽  
Ebola Virus ◽  
Genomic Sequence ◽  
Sequence Data ◽  
Virus Disease ◽  
Live Cells ◽  
Viral Fusion ◽  
Viral Glycoprotein ◽  
Viral Spread

AbstractGenomic surveillance of viral isolates during the 2013-2016 Ebola virus epidemic in Western Africa—the largest and most devastating filovirus outbreak on record—revealed several novel mutations. The responsible strain, named Makona, carries an A to V substitution at position 82 in the glycoprotein (GP), which is associated with enhanced infectivity in vitro. Here, we investigated the mechanistic basis for this enhancement, as well as the interplay between A82V and a T to I substitution at residue 544 of GP, which also modulates infectivity in cell culture. We found that both 82V and 544I destabilize GP with the residue at 544 impacting overall stability, while 82V specifically destabilizes proteolytically cleaved GP. Both residues also promote faster kinetics of lipid mixing of the viral and host membranes in live cells, individually and in tandem, which correlates with faster times to fusion following co-localization with the viral receptor Niemann-Pick C1 (NPC1). Further, GPs bearing 82V are more sensitive to proteolysis by cathepsin L (CatL), a key host factor for viral entry. Intriguingly, CatL processed 82V variant GPs to a novel product of ∼12K size, which we hypothesize corresponds to a form of GP more fully primed for fusion than previously detected. We thus propose a model in which 82V promotes more efficient GP processing by CatL, leading to faster viral fusion kinetics and higher infectivity.ImportanceThe 2013-2016 outbreak of Ebola virus disease in West Africa demonstrated the potential for previously localized outbreaks to turn into regional, or even global, health emergencies. With over 28,000 cases and 11,000 confirmed deaths, this outbreak was over 50 times as large as any previously recorded. This outbreak also afforded the largest ever collection of Ebola virus genomic sequence data, allowing new insights into viral transmission and evolution. Viral mutants arising during the outbreak have attracted attention for their potentially altered patterns of infectivity in cell culture, with potential, if unclear, implications for increased viral spread and/or virulence. Here, we report on the properties of one such mutation in the viral glycoprotein, A82V, and its interplay with a previously described polymorphism at position 544. We show that mutations at both residues promote infection and fusion activation in cells, but that A82V additionally leads to increased infectivity under cathepsin-limited conditions, and the generation of a novel glycoprotein cleavage product.

scholarly journals A Glycoprotein Mutation That Emerged during the 2013–2016 Ebola Virus Epidemic Alters Proteolysis and Accelerates Membrane Fusion

mBio ◽  
2021 ◽  
Vol 12 (1) ◽  
Author(s):  
J. Maximilian Fels ◽  
Robert H. Bortz ◽  
Tanwee Alkutkar ◽  
Eva Mittler ◽  
Rohit K. Jangra ◽  
...  
Keyword(s):  
Cell Culture ◽  
Ebola Virus ◽  
Genomic Sequence ◽  
Sequence Data ◽  
Virus Disease ◽  
Live Cells ◽  
Viral Fusion ◽  
Viral Glycoprotein ◽  
Viral Spread

ABSTRACT Genomic surveillance of viral isolates during the 2013–2016 Ebola virus epidemic in Western Africa, the largest and most devastating filovirus outbreak on record, revealed several novel mutations. The responsible strain, named Makona, carries an A-to-V substitution at position 82 (A82V) in the glycoprotein (GP), which is associated with enhanced infectivity in vitro. Here, we investigated the mechanistic basis for this enhancement as well as the interplay between A82V and a T-to-I substitution at residue 544 of GP, which also modulates infectivity in cell culture. We found that both 82V and 544I destabilize GP, with the residue at position 544 impacting overall stability, while 82V specifically destabilizes proteolytically cleaved GP. Both residues also promote faster kinetics of lipid mixing of the viral and host membranes in live cells, individually and in tandem, which correlates with faster times to fusion following colocalization with the viral receptor Niemann-Pick C1 (NPC1). Furthermore, GPs bearing 82V are more sensitive to proteolysis by cathepsin L (CatL), a key host factor for viral entry. Intriguingly, CatL processed 82V variant GPs to a novel product with a molecular weight of approximately 12,000 (12K), which we hypothesize corresponds to a form of GP that is pre-triggered for fusion. We thus propose a model in which 82V promotes more efficient GP processing by CatL, leading to faster viral fusion kinetics and higher levels of infectivity. IMPORTANCE The 2013–2016 outbreak of Ebola virus disease in West Africa demonstrated the potential for previously localized outbreaks to turn into regional, or even global, health emergencies. With over 28,000 cases and 11,000 confirmed deaths, this outbreak was over 50 times as large as any previously recorded. This outbreak also afforded the largest-ever collection of Ebola virus genomic sequence data, allowing new insights into viral transmission and evolution. Viral mutants arising during the outbreak have attracted attention for their potentially altered patterns of infectivity in cell culture, with potential, if unclear, implications for increased viral spread and/or virulence. Here, we report the properties of one such mutation in the viral glycoprotein, A82V, and its interplay with a previously described polymorphism at position 544. We show that mutations at both residues promote infection and fusion activation in cells but that A82V additionally leads to increased infectivity under cathepsin-limited conditions and the generation of a novel glycoprotein cleavage product.

Treatment of ebola and other infectious diseases: melatonin “goes viral”

2020 ◽  
Vol 3 (1) ◽  
pp. 43-57 ◽  
Author(s):  
Russel J Reiter ◽  
Qiang Ma ◽  
Ramaswamy Sharma
Keyword(s):  
Mortality Rate ◽  
Infectious Diseases ◽  
Hemorrhagic Shock ◽  
Safety Profile ◽  
Ebola Virus ◽  
Viral Infections ◽  
Virus Disease ◽  
Attractive Potential ◽  
Potential Treatment

This review summarizes published reports on the utility of melatonin as a treatment for virus-mediated diseases. Of special note are the data related to the role of melatonin in influencing Ebola virus disease. This infection and deadly condition has no effective treatment and the published works documenting the ability of melatonin to attenuate the severity of viral infections generally and Ebola infection specifically are considered. The capacity of melatonin to prevent one of the major complications of an Ebola infection, i.e., the hemorrhagic shock syndrome, which often contributes to the high mortality rate, is noteworthy. Considering the high safety profile of melatonin, the fact that it is easily produced, inexpensive and can be self-administered makes it an attractive potential treatment for Ebola virus pathology.  

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