Novel #Mutations Evading #Avian #Immunity around the #Receptor Binding Site of the Clade 2.3.2.1c #Hemagglutinin Gene Reduce Viral Thermostability and Mammalian Pathogenicity (Viruses, abstract)

[Source: US National Library of Medicine, full page: (LINK). Abstract, edited.]

Viruses. 2019 Oct 9;11(10). pii: E923. doi: 10.3390/v11100923.

Novel Mutations Evading Avian Immunity around the Receptor Binding Site of the Clade 2.3.2.1c Hemagglutinin Gene Reduce Viral Thermostability and Mammalian Pathogenicity.

An SH1, Lee CY2, Hong SM3, Song CS4, Kim JH5,6, Kwon HJ7,8,9.

Author information: 1 Laboratory of Avian Diseases, College of Veterinary Medicine, Seoul National University, Seoul 08826, Korea. eepdl1201@snu.ac.kr. 2 Department of Microbiology and Immunology, Emory University School of Medicine, 1510 Clifton Road, Atlanta, GA 30322, USA. chung-young.lee@emory.edu. 3 Laboratory of Avian Diseases, College of Veterinary Medicine, Seoul National University, Seoul 08826, Korea. topkin@snu.ac.kr. 4 Laboratory of Avian Diseases, College of Veterinary Medicine, Konkuk University, Seoul 05029, Korea. songcs@konkuk.ac.kr. 5 Laboratory of Avian Diseases, College of Veterinary Medicine, Seoul National University, Seoul 08826, Korea. kimhong@snu.ac.kr. 6 Research Institute for Veterinary Science, College of Veterinary Medicine, Seoul National University, 08826 Seoul, Korea. kimhong@snu.ac.kr. 7 Research Institute for Veterinary Science, College of Veterinary Medicine, Seoul National University, 08826 Seoul, Korea. kwonhj01@snu.ac.kr. 8 Laboratory of Poultry Medicine, Department of Farm Animal Medicine, College of Veterinary Medicine, Seoul National University, Seoul 08826, Korea. kwonhj01@snu.ac.kr. 9 Farm Animal Clinical Training and Research Center (FACTRC), GBST, Seoul National University, Kangwon-do 88026, Korea. kwonhj01@snu.ac.kr.

 

Abstract

Abstract: Since 2007, highly pathogenic clade 2.3.2 H5N1 avian influenza A (A(H5N1)) viruses have evolved to clade 2.3.2.1a, b, and c; currently only 2.3.2.1c A(H5N1) viruses circulate in wild birds and poultry. During antigenic evolution, clade 2.3.2.1a and c A(H5N1) viruses acquired both S144N and V223I mutations around the receptor binding site of hemagglutinin (HA), with S144N generating an N-glycosylation sequon. We introduced single or combined reverse mutations, N144S and/or I223V, into the HA gene of the clade 2.3.2.1c A(H5N1) virus and generated PR8-derived, 2 + 6 recombinant A(H5N1) viruses. When we compared replication efficiency in embryonated chicken eggs, mammalian cells, and mice, the recombinant virus containing both N144S and I223V mutations showed increased replication efficiency in avian and mammalian hosts and pathogenicity in mice. The N144S mutation significantly decreased avian receptor affinity and egg white inhibition, but not all mutations increased mammalian receptor affinity. Interestingly, the combined reverse mutations dramatically increased the thermostability of HA. Therefore, the adaptive mutations possibly acquired to evade avian immunity may decrease viral thermostability as well as mammalian pathogenicity.

KEYWORDS: HA trimer stability; clade 2.3.2.1c H5N1 virus; immunity evasion; mammalian pathogenicity; thermostability

PMID: 31600990 DOI: 10.3390/v11100923

Keywords: Avian Influenza; H5N1; Viral pathogenesis.

——

Advertisements

#Regulation of the #Ebola virus #VP24 protein by #SUMO (J Virol., abstract)

[Source: Journal of Virology, full page: (LINK). Abstract, edited.]

Regulation of the Ebola virus VP24 protein by SUMO

Santiago Vidal, Ahmed El Motiam, Rocío Seoane, Viktorija Preitakaite, Yanis Hichem Bouzaher, Sergio Gómez-Medina, Carmen San Martín, Dolores Rodríguez, María Teresa Rejas, Maite Baz-Martínez, Rosa Barrio, James D Sutherland, Manuel S Rodríguez, César Muñoz-Fontela, Carmen Rivas

DOI: 10.1128/JVI.01687-19

 

ABSTRACT

Some viruses take advantage of ubiquitin or ubiquitin-like proteins conjugation to enhance their own replication. One example is Ebola virus, which has evolved strategies to utilize these modification pathways to regulate the viral proteins VP40 and VP35 and to counteract the host defenses. Here we show a novel mechanism by which Ebola virus exploits the ubiquitin and SUMO pathways. Our data expose that the minor matrix protein VP24 of Ebola virus is a bona fide SUMO target. Analysis of a SUMOylation-defective VP24 mutant revealed a reduced capability to block the type I interferon pathway and to inhibit IFN-mediated STAT1-nuclear translocation, exhibiting a weaker interaction with karyopherin 5 and a significantly diminished stability. Using GST-pull down assay we found that VP24 also interacts with SUMO in a non-covalent manner through a SIM domain. Mutation of the SIM domain in VP24 resulted in a complete inability of the protein to down-modulate the IFN pathway and in the monoubiquitination of the protein. We identified the SUMO deubiquitinating enzyme USP7 as an interactor and negative modulator of VP24 ubiquitination. Finally, we show that mutation of one ubiquitination site in VP24 potentiates the IFN modulatory activity of the viral protein and its ability to block IFN-mediated STAT1 nuclear translocation, pointing to the ubiquitination of VP24 as a negative modulator of the VP24 activity. Altogether these results indicate that SUMO interacts with VP24 and promotes its USP7-mediated deubiquitination, playing a key role in the interference with the innate immune response mediated by the viral protein.

 

IMPORTANCE

The Ebola virus VP24 protein plays a critical role in the virus escape from the host innate immune response. Therefore, deciphering the molecular mechanisms modulating VP24 activity may be useful to identify potential targets amenable for therapeutics. Here we identify the cellular proteins USP7, SUMO, and ubiquitin as novel interactors and regulators of VP24. These interactions may represent novel potential targets to design new antivirals with the ability to modulate Ebola virus replication.

Copyright © 2019 American Society for Microbiology. All Rights Reserved.

Keywords: Ebola; Viral pathogenesis.

——

#Hemagglutinin-dependent #tropism of #H5N1 and #H7N9 #influenza viruses to human endothelial cells is determined by reduced stability of the HA which allows the virus to cope with inefficient endosomal acidification and constitutively expressed IFITM3 (J Virol., abstract)

[Source: Journal of Virology, full page: (LINK). Abstract, edited.]

Hemagglutinin-dependent tropism of H5N1 and H7N9 influenza viruses to human endothelial cells is determined by reduced stability of the HA which allows the virus to cope with inefficient endosomal acidification and constitutively expressed IFITM3

Luca Hensen, Tatyana Matrosovich, Katrin Roth, Hans-Dieter Klenk, Mikhail Matrosovich

DOI: 10.1128/JVI.01223-19

 

ABSTRACT

Previous studies revealed that certain avian influenza A viruses (IAVs), including zoonotic H5N1 and H7N9 IAVs, infect cultured human lung microvascular endothelial cells (HULEC) more efficiently than other IAVs and that tropism to HULEC is determined by viral hemagglutinin (HA). To characterize mechanisms of HA-mediated endotheliotropism, we used 2:6 recombinant IAVs harbouring HAs from distinctive avian and human viruses and found that efficient infection of HULEC correlated with low conformational stability of the HA. We next studied effects on viral infectivity of single-point amino acid substitutions in the HA of 2:6 recombinant virus A/Vietnam/1203/2004-PR8 (H5N1). Substitutions H8Q, H103Y, T315I and K582I, which increased stability of the HA, markedly reduced viral infectivity for HULEC, whereas substitutions K189N and K218Q, which altered typical H5N1-virus-like receptor specificity and reduced binding avidity of the HA, led to only marginal reduction of infectivity. None of these substitutions affected virus infection in MDCK cells. We confirmed the previous observation of elevated basal expression of IFITM3 protein in HULEC and found that endosomal acidification is less efficient in HULEC than in MDCK cells. In accord with these findings, counteraction of IFITM3-mediated restriction by amphotericin B and reduction of endosomal pH by moderate acidification of the extracellular medium, enhanced infectivity of viruses with stable HA for HULEC without significant effect on infectivity for MDCK cells. Collectively, our results indicate that relatively high pH optimum of fusion of the HA of zoonotic H5N1 and H7N9 IAVs allows them to overcome antiviral effects of inefficient endosomal acidification and IFITM3 in human endothelial cells.

 

IMPORTANCE

Receptor specificity of the HA of IAVs is known to be a critical determinant of viral cell tropism. Here we show that fusion properties of the HA may also play a key role in the tropism. Thus, we demonstrate that IAVs having relatively low pH optimum of fusion, cannot efficiently infect human endothelial cells owing to their relatively high endosomal pH and increased expression of fusion-inhibiting IFITM3 protein. These restrictions can be overcome by IAVs with elevated pH of fusion, such as zoonotic H5N1 and H7N9. Our results illustrate that infectivity of IAVs depends on an interplay between HA conformational stability, endosomal acidification and IFITM3 expression in target cells and extracellular pH. Given significant variation of HA stability among animal, human and zoonotic IAVs, our findings prompt further studies on the fusion-dependent tropism of IAVs to different cell types in humans and its role in viral host range and pathogenicity.

Copyright © 2019 American Society for Microbiology. All Rights Reserved.

Keywords: Influenza A; Avian Influenza; H5N1; H7N9; Viral pathogenesis.

——

Expression of #microRNA in #human #retinal pigment #epithelial cells following #infection with #Zaire #ebolavirus (BMC Res Notes, abstract)

[Source: US National Library of Medicine, full page: (LINK). Abstract, edited.]

BMC Res Notes. 2019 Oct 1;12(1):639. doi: 10.1186/s13104-019-4671-8.

Expression of microRNA in human retinal pigment epithelial cells following infection with Zaire ebolavirus.

Oliver GF1, Orang AV1, Appukuttan B1, Marri S1, Michael MZ1, Marsh GA2, Smith JR3.

Author information: 1 Flinders University College of Medicine and Public Health, Flinders Medical Centre Room 4E-431, Flinders Drive, Bedford Park, SA, 5042, Australia. 2 Health and Biosecurity, Commonwealth Scientific and Industrial Research Organisation, 5 Portarlington Rd, Newcomb, VIC, 3219, Australia. 3 Flinders University College of Medicine and Public Health, Flinders Medical Centre Room 4E-431, Flinders Drive, Bedford Park, SA, 5042, Australia. justine.smith@flinders.edu.au.

 

Abstract

OBJECTIVE:

Survivors of Ebola virus disease (EVD) are at risk of developing blinding intraocular inflammation-or uveitis-which is associated with retinal pigment epithelial (RPE) scarring and persistence of live Zaire ebolavirus (EBOV) within the eye. As part of a large research project aimed at defining the human RPE cell response to being infected with EBOV, this work focused on the microRNAs (miRNAs) associated with the infection.

RESULTS:

Using RNA-sequencing, we detected 13 highly induced and 2 highly repressed human miRNAs in human ARPE-19 RPE cells infected with EBOV, including hsa-miR-1307-5p, hsa-miR-29b-3p and hsa-miR-33a-5p (up-regulated), and hsa-miR-3074-3p and hsa-miR-27b-5p (down-regulated). EBOV-miR-1-5p was also found in infected RPE cells. Through computational identification of putative miRNA targets, we predicted a broad range of regulatory activities, including effects on innate and adaptive immune responses, cellular metabolism, cell cycle progression, apoptosis and autophagy. The most highly-connected molecule in the miR-target network was leucine-rich repeat kinase 2, which is involved in neuroinflammation and lysosomal processing. Our findings should stimulate new studies on the impact of miRNA changes in EBOV-infected RPE cells to further understanding of intraocular viral persistence and the pathogenesis of uveitis in EVD survivors.

KEYWORDS: Ebola; Filovirus; Retina; Retinal pigment epithelium; Uveitis; Zaire ebolavirus; microRNA

PMID: 31570108 DOI: 10.1186/s13104-019-4671-8

Keywords: Ebola; ZEBOV; Viral pathogenesis.

——

#Dendritic Cells / #Macrophages-Targeting Feature of #Ebola #Glycoprotein and its Potential as Immunological Facilitator for Antiviral #Vaccine Approach (Microorganisms, abstract)

[Source: US National Library of Medicine, full page: (LINK). Abstract, edited.]

Microorganisms. 2019 Sep 29;7(10). pii: E402. doi: 10.3390/microorganisms7100402.

Dendritic Cells/Macrophages-Targeting Feature of Ebola Glycoprotein and its Potential as Immunological Facilitator for Antiviral Vaccine Approach.

Olukitibi TA1, Ao Z2, Mahmoudi M3, Kobinger GA4, Yao X5.

Author information: 1 Laboratory of Molecular Human Retrovirology, Department of Medical Microbiology, Max Rady College of Medicine, Rady Faculty of Health Sciences, University of Manitoba, Winnipeg, MB R3E 0J9, Canada. olukitit@myumanitoba.ca. 2 Laboratory of Molecular Human Retrovirology, Department of Medical Microbiology, Max Rady College of Medicine, Rady Faculty of Health Sciences, University of Manitoba, Winnipeg, MB R3E 0J9, Canada. zhujun.ao@umanitoba.ca. 3 Laboratory of Molecular Human Retrovirology, Department of Medical Microbiology, Max Rady College of Medicine, Rady Faculty of Health Sciences, University of Manitoba, Winnipeg, MB R3E 0J9, Canada. mahmoud1@myumanitoba.ca. 4 Centre de Recherche en Infectiologie de l’ Université Laval/Centre Hospitalier de l’ Université Laval (CHUL), Québec, QC G1V 4G2, Canada. Gary.Kobinger@crchudequebec.ulaval.ca. 5 Laboratory of Molecular Human Retrovirology, Department of Medical Microbiology, Max Rady College of Medicine, Rady Faculty of Health Sciences, University of Manitoba, Winnipeg, MB R3E 0J9, Canada. xiao-jian.yao@umanitoba.ca.

 

Abstract

In the prevention of epidemic and pandemic viral infection, the use of the antiviral vaccine has been the most successful biotechnological and biomedical approach. In recent times, vaccine development studies have focused on recruiting and targeting immunogens to dendritic cells (DCs) and macrophages to induce innate and adaptive immune responses. Interestingly, Ebola virus (EBOV) glycoprotein (GP) has a strong binding affinity with DCs and macrophages. Shreds of evidence have also shown that the interaction between EBOV GP with DCs and macrophages leads to massive recruitment of DCs and macrophages capable of regulating innate and adaptive immune responses. Therefore, studies for the development of vaccine can utilize the affinity between EBOV GP and DCs/macrophages as a novel immunological approach to induce both innate and acquired immune responses. In this review, we will discuss the unique features of EBOV GP to target the DC, and its potential to elicit strong immune responses while targeting DCs/macrophages. This review hopes to suggest and stimulate thoughts of developing a stronger and effective DC-targeting vaccine for diverse virus infection using EBOV GP.

KEYWORDS: Ebola glycoprotein; antiviral vaccine; dendritic cells and macrophages; immune response

PMID: 31569539 DOI: 10.3390/microorganisms7100402

Keywords: Ebola; Viral pathogenesis.

——

Activation of C-type lectin receptor and (#RIG)-I-like #receptors contributes to #proinflammatory response in #MERS #coronavirus infected #macrophages (J Infect Dis., abstract)

[Source: Journal of Infectious Diseases, full page: (LINK). Abstract, edited.]

Activation of C-type lectin receptor and (RIG)-I-like receptors contributes to proinflammatory response in MERS coronavirus infected macrophages

Xiaoyu Zhao, Hin Chu, Bosco Ho-Yin Wong, Man Chun Chiu, Dong Wang, Cun Li, Xiaojuan Liu, Dong Yang, Vincent Kwok-Man Poon, Jianpiao Cai, Jasper Fuk-Woo Chan, Kelvin Kai-Wang To, Jie Zhou, Kwok-Yung Yuen

The Journal of Infectious Diseases, jiz483, https://doi.org/10.1093/infdis/jiz483

Published: 28 September 2019

 

Abstract

Background

Human infection with Middle East respiratory syndrome coronavirus (MERS-CoV) poses an ongoing threat to public health worldwide. The studies of MERS patients with severe disease and experimentally-infected animals showed that robust viral replication and intensive proinflammatory response in lung tissues contribute to high pathogenicity of MERS-CoV. We sought to identify pattern recognition receptor (PRR) signaling pathway(s) that mediates the inflammatory cascade in human macrophages upon MERS-CoV infection.

Methods

The potential signaling pathways were manipulated individually by pharmacological inhibition, siRNA depletion and antibody blocking. MERS-CoV-induced proinflammatory response was evaluated by measuring the expression levels of key cytokines/chemokines. RT-qPCR assay, flow cytometry analysis and Western blotting were applied to evaluate the activation of related PRRs and engagement of adaptors.

Results

MERS-CoV replication significantly upregulated C-type lectin receptor (CLR) Mincle. The role of Mincle for MERS-CoV-triggered cytokine/chemokine induction was established based on the results of antibody blockage, siRNA depletion of Mincle and its adaptor Syk, and Syk pharmacological inhibition. The cytokine/chemokine induction was significantly attenuated by siRNA depletion of RIG-I-like receptors (RLR) or adaptor, indicating RLR signaling also contributed to MERS-CoV-induced proinflammatory response.

Conclusion

CLR and RLR pathways are activated and contribute to the proinflammatory response in MERS-CoV-infected macrophages.

MERS-CoV, CLR, RLR, proinflammatory response, Mincle

Topic:  cytokine – chemokines – macrophages – antibodies – pharmacology – signal pathway – signal transduction pathways – rna, small interfering – middle east respiratory syndrome coronavirus – middle east respiratory syndrome – c-type lectins

Issue Section: Major Article

This content is only available as a PDF.

© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

This article is published and distributed under the terms of the Oxford University Press, Standard Journals Publication Model (https://academic.oup.com/journals/pages/open_access/funder_policies/chorus/standard_publication_model)

Keywords: MERS-CoV; Viral pathogenesis.

——

#Zika virus enhances #monocyte adhesion and transmigration favoring viral #dissemination to #neural cells (Nat Commun., abstract)

[Source: US National Library of Medicine, full page: (LINK). Abstract, edited.]

Nat Commun. 2019 Sep 27;10(1):4430. doi: 10.1038/s41467-019-12408-x.

Zika virus enhances monocyte adhesion and transmigration favoring viral dissemination to neural cells.

Ayala-Nunez NV1,2, Follain G3, Delalande F4, Hirschler A4, Partiot E1, Hale GL5, Bollweg BC5, Roels J6, Chazal M7, Bakoa F7, Carocci M8, Bourdoulous S9, Faklaris O10, Zaki SR5, Eckly A8, Uring-Lambert B11, Doussau F12, Cianferani S4, Carapito C4, Jacobs FMJ6, Jouvenet N7, Goetz JG3, Gaudin R13,14.

Author information: 1 Institut de Recherche en Infectiologie de Montpellier (IRIM), CNRS, Université de Montpellier, 34293, Montpellier, France. 2 Université de Strasbourg, INSERM, 67000, Strasbourg, France. 3 INSERM U1109 and FMTS, 67000, Strasbourg, France. 4 Laboratoire de Spectrométrie de Masse Bio-Organique, IPHC, UMR 7178, CNRS-Université de Strasbourg, ECPM, 67087, Strasbourg, France. 5 Infectious Diseases Pathology Branch, Division of High-Consequence Pathogens and Pathology, National Center for Emerging and Zoonotic Infectious Diseases (NCEZID), Centers for Disease Control and Prevention, 1600 Clifton Rd NE, MS: G32, Atlanta, GA, 30329-4027, USA. 6 University of Amsterdam, Swammerdam Institute for Life Sciences, Science Park 904, 1098XH, Amsterdam, The Netherlands. 7 Viral Genomics and Vaccination Unit, UMR3569 CNRS, Virology Department, Institut Pasteur, 75015, Paris, France. 8 Université de Strasbourg, INSERM, EFS Grand Est, BPPS UMR-S1255, FMTS, 67000, Strasbourg, France. 9 INSERM U1016, Institut Cochin, CNRS UMR8104, Université Paris Descartes, Paris, France. 10 MRI Core facility, Biocampus, CNRS UMS 3426, 34293, Montpellier, France. 11 Hôpitaux universitaires de Strasbourg, laboratoire central d’immunologie, 67000, Strasbourg, France. 12 Institut des Neurosciences Cellulaires et Intégratives, CNRS, Université de Strasbourg, 67000, Strasbourg, France. 13 Institut de Recherche en Infectiologie de Montpellier (IRIM), CNRS, Université de Montpellier, 34293, Montpellier, France. raphael.gaudin@irim.cnrs.fr. 14 Université de Strasbourg, INSERM, 67000, Strasbourg, France. raphael.gaudin@irim.cnrs.fr.

 

Abstract

Zika virus (ZIKV) invades and persists in the central nervous system (CNS), causing severe neurological diseases. However the virus journey, from the bloodstream to tissues through a mature endothelium, remains unclear. Here, we show that ZIKV-infected monocytes represent suitable carriers for viral dissemination to the CNS using human primary monocytes, cerebral organoids derived from embryonic stem cells, organotypic mouse cerebellar slices, a xenotypic human-zebrafish model, and human fetus brain samples. We find that ZIKV-exposed monocytes exhibit higher expression of adhesion molecules, and higher abilities to attach onto the vessel wall and transmigrate across endothelia. This phenotype is associated to enhanced monocyte-mediated ZIKV dissemination to neural cells. Together, our data show that ZIKV manipulates the monocyte adhesive properties and enhances monocyte transmigration and viral dissemination to neural cells. Monocyte transmigration may represent an important mechanism required for viral tissue invasion and persistence that could be specifically targeted for therapeutic intervention.

PMID: 31562326 DOI: 10.1038/s41467-019-12408-x

Keywords: Zika Virus; Neuroinvasion; Viral pathogenesis.

——