scholarly journals Vaccinia E5 is a major inhibitor of the DNA sensor cGAS

2021 ◽  
Author(s):  
Ning Yang ◽  
Yi Wang ◽  
Peihong Dai ◽  
Tuo Li ◽  
Christian Zierhut ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Viral Vector ◽  
Type I ◽  
Dna Sensor ◽  
Modified Vaccinia Virus Ankara ◽  
Cytoplasmic Dna ◽  
Infected Cells ◽  
Sting Pathway ◽  
Interferon Gene ◽  
Dc Maturation

The DNA sensor cyclic GMP-AMP synthase (cGAS) is critical in host antiviral immunity. Vaccinia virus (VACV) is a large cytoplasmic DNA virus that belongs to the poxvirus family. How vaccinia virus antagonizes the cGAS-mediated cytosolic DNA-sensing pathway is largely unknown. In this study, we screened 82 vaccinia viral genes to identify potential viral inhibitors of the cGAS/Stimulator of interferon gene (STING) pathway. We discovered that vaccinia E5 is a virulence factor and a major inhibitor of cGAS that elicits proteasome-dependent cGAS degradation. E5 localizes to the cytoplasm and nuclei of infected cells. Cytosolic E5 triggers K48-linked ubiquitination of cGAS and proteasome-dependent degradation via interacting with cGAS. E5 itself also undergoes ubiquitination and degradation. Deleting the E5R gene from the Modified vaccinia virus Ankara (MVA) genome strongly induces type I IFN production by dendritic cells (DCs) and promotes DC maturation, thereby improving the immunogenicity of the viral vector.

scholarly journals Intratumoral delivery of engineered recombinant modified vaccinia virus Ankara express-ing Flt3L and OX40L generates potent antitumor immunity through activating the cGAS/STING pathway and depleting tumor-infiltrating regulatory T cells

2021 ◽  
Author(s):  
Ning Yang ◽  
Yi Wang ◽  
Shuaitong Liu ◽  
Joseph M Luna ◽  
Gregory Mazo ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Vaccinia Virus ◽  
Antitumor Immunity ◽  
Type I ◽  
Innate And Adaptive Immunity ◽  
Modified Vaccinia Virus Ankara ◽  
History Of ◽  
Intratumoral Delivery ◽  
Sting Pathway

Intratumoral (IT) delivery of immune-activating viruses can serve as an important strategy to turn cold tumors into hot tumors, resulting in overcoming resistance to immune checkpoint block-ade (ICB). Modified vaccinia virus Ankara (MVA) is a highly attenuated, non-replicative vaccinia virus that has a long history of human use. Here we report that IT recombinant MVA (rMVA), lacking E5R encoding an inhibitor of the DNA sensor cyclic GMP-AMP synthase (cGAS), ex-pressing a dendritic cell growth factor, Fms-like tyrosine kinase 3 ligand (Flt3L), and a T cell co-stimulator, OX40L, generates strong antitumor immunity, which is dependent on CD8+ T cells, the cGAS/STING-mediated cytosolic DNA-sensing pathway, and STAT1/STAT2-mediated type I IFN signaling. Remarkably, IT rMVA depletes OX40hi regulatory T cells via OX40L/OX40 inter-action and IFNAR signaling. Taken together, our study provides a proof-of-concept for improving MVA-based cancer immunotherapy, through modulation of both innate and adaptive immunity.

scholarly journals Vaccinia Virus F13L Protein with a Conserved Phospholipase Catalytic Motif Induces Colocalization of the B5R Envelope Glycoprotein in Post-Golgi Vesicles

2001 ◽  
Vol 75 (16) ◽  
pp. 7528-7542 ◽  
Author(s):  
Matloob Husain ◽  
Bernard Moss
Keyword(s):  
Vaccinia Virus ◽  
Fluorescent Protein ◽  
Plasmid Vector ◽  
Recombinant Vaccinia Virus ◽  
Open Reading Frames ◽  
Type I ◽  
Golgi Vesicles ◽  
Infected Cells ◽  
Catalytic Motif ◽  
Green Fluorescent

ABSTRACT The wrapping of intracellular mature vaccinia virions by modifiedtrans-Golgi or endosomal cisternae to form intracellular enveloped virions is dependent on at least two viral proteins encoded by the B5R and F13L open reading frames. B5R is a type I integral membrane glycoprotein, whereas F13L is an unglycosylated, palmitylated protein with a motif that is conserved in a superfamily of phospholipid-metabolizing enzymes. Microscopic visualization of the F13L protein was achieved by fusing it to the enhanced green fluorescent protein (GFP). F13L-GFP was functional when expressed by a recombinant vaccinia virus in which it replaced the wild-type F13L gene or by transfection of uninfected cells with a plasmid vector followed by infection with an F13L deletion mutant. In uninfected or infected cells, F13L-GFP was associated with Golgi cisternae and post-Golgi vesicles containing the LAMP 2 late endosomal-lysosomal marker. Association of F13L-GFP with vesicles was dependent on an intact phospholipase catalytic motif and sites of palmitylation. The B5R protein was also associated with LAMP2-containing vesicles when F13L-GFP was coexpressed, but was largely restricted to Golgi cisternae in the absence of F13L-GFP or when the F13L moiety was mutated. We suggest that the F13L protein, like its human phospholipase D homolog, regulates vesicle formation and that this process is involved in intracellular enveloped virion membrane formation.

scholarly journals Modified Vaccinia Virus Ankara Triggers Chemotaxis of Monocytes and Early Respiratory Immigration of Leukocytes by Induction of CCL2 Expression

2009 ◽  
Vol 83 (6) ◽  
pp. 2540-2552 ◽  
Author(s):  
Michael H. Lehmann ◽  
Wolfgang Kastenmuller ◽  
Judith D. Kandemir ◽  
Florian Brandt ◽  
Yasemin Suezer ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Viral Vector ◽  
Monocytic Cell ◽  
Monocytic Cell Line ◽  
Human Monocytic Cell Line ◽  
Modified Vaccinia Virus Ankara ◽  
Ccl2 Expression ◽  
Human Monocytic Cell

ABSTRACT Orthopoxviruses commonly enter into humans and animals via the respiratory tract. Herein, we show that immigration of leukocytes into the lung is triggered via intranasal infection of mice with modified vaccinia virus Ankara (MVA) and not with the vaccinia virus (VACV) Elstree, Wyeth, or Western Reserve (WR) strain. Immigrating cells were identified as monocytes, neutrophils, and CD4+ lymphocytes by flow cytometry and could be detected 24 h and 48 h postinfection. Using an in vitro chemotaxis assay, we confirmed that infection with MVA induces the expression of a soluble chemotactic factor for monocytes, identified as CCL2 (monocyte chemotactic protein-1 [MCP-1]). In contrast to infection with several other VACV strains, MVA induced the expression of CCL2, CCL3, CCL4, and CXCL10 in the human monocytic cell line THP-1 as well as in primary human monocytes. Thus, MVA, and not the VACV Elstree, Wyeth, or WR strain, consistently triggered the expression of a panel of chemokines, including CCL2, in the murine lung, correlating considerably with the immigration of leukocytes. Using CCL2-deficient mice, we demonstrate that CCL2 plays a key role in MVA-triggered respiratory immigration of leukocytes. Moreover, UV irradiation of MVA prevented CCL2 expression in vitro and in vivo as well as respiratory immigration of leukocytes, demonstrating the requirement for an activated molecular viral life cycle. We propose that MVA-triggered chemokine expression causes early immigration of leukocytes to the site of infection, a feature that is important for rapid immunization and its safety and efficiency as a viral vector.

scholarly journals Direct Comparison of Antigen Production and Induction of Apoptosis by Canarypox Virus- and Modified Vaccinia Virus Ankara-Human Immunodeficiency Virus Vaccine Vectors

2007 ◽  
Vol 81 (13) ◽  
pp. 7022-7033 ◽  
Author(s):  
Xiugen Zhang ◽  
Farah Cassis-Ghavami ◽  
Mike Eller ◽  
Jeff Currier ◽  
Bonnie M. Slike ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Vaccinia Virus ◽  
Mammalian Cells ◽  
Apoptosis Induction ◽  
Antigen Production ◽  
Modified Vaccinia Virus Ankara ◽  
Immunodeficiency Virus ◽  
Host Cell Interactions ◽  
Infected Cells ◽  
Canarypox Virus

ABSTRACT Recombinant poxvirus vectors are undergoing intensive evaluation as vaccine candidates for a variety of infectious pathogens. Avipoxviruses, such as canarypox virus, are replication deficient in mammalian cells by virtue of a poorly understood species-specific restriction. Highly attenuated vaccinia virus strains such as modified vaccinia virus Ankara (MVA) are similarly unable to complete replication in most mammalian cells but have an abortive-late phenotype, in that the block to replication occurs post-virus-specific DNA replication. In this study, an identical expression cassette for human immunodeficiency virus gag, pro, and env coding sequences was placed in canarypox virus and MVA vector backbones in order to directly compare vector-borne expression and to analyze differences in vector-host cell interactions. Antigen production by recombinant MVA was shown to be greater than that from recombinant canarypox virus in the mammalian cell lines and in the primary human cells tested. This observation was primarily due to a longer duration of antigen production in recombinant MVA-infected cells. Apoptosis induction was found to be more profound with the empty canarypox virus vector than with MVA. Remarkably, however, the inclusion of a gag/pro/env expression cassette altered the kinetics of apoptosis induction in recombinant MVA-infected cells to levels equal to those found in canarypox virus-infected cells. Antigen production by MVA was noted to be greater in human dendritic cells and resulted in enhanced T-cell stimulation in an in vitro antigen presentation assay. These results reveal differences in poxvirus vector-host cell interactions that should be relevant to their use as immunization vehicles.

scholarly journals Modified Vaccinia Virus Ankara Triggers Type I IFN Production in Murine Conventional Dendritic Cells via a cGAS/STING-Mediated Cytosolic DNA-Sensing Pathway

2014 ◽  
Vol 10 (4) ◽  
pp. e1003989 ◽  
Author(s):  
Peihong Dai ◽  
Weiyi Wang ◽  
Hua Cao ◽  
Francesca Avogadri ◽  
Lianpan Dai ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Vaccinia Virus ◽  
Type I ◽  
Type I Ifn ◽  
Dna Sensing ◽  
Modified Vaccinia Virus Ankara

scholarly journals Chicken cGAS Senses Fowlpox Virus Infection and Regulates Macrophage Effector Functions

2021 ◽  
Vol 11 ◽  
Author(s):  
Marisa Oliveira ◽  
Damaris Ribeiro Rodrigues ◽  
Vanaique Guillory ◽  
Emmanuel Kut ◽  
Efstathios S. Giotis ◽  
...  
Keyword(s):  
Virus Infection ◽  
Adaptor Protein ◽  
Type I Interferons ◽  
Type I ◽  
Fowlpox Virus ◽  
Effector Functions ◽  
Mhc Ii ◽  
Infected Cells ◽  
Sting Pathway ◽  
Multiple Species

The anti-viral immune response is dependent on the ability of infected cells to sense foreign nucleic acids. In multiple species, the pattern recognition receptor (PRR) cyclic GMP-AMP synthase (cGAS) senses viral DNA as an essential component of the innate response. cGAS initiates a range of signaling outputs that are dependent on generation of the second messenger cGAMP that binds to the adaptor protein stimulator of interferon genes (STING). Here we show that in chicken macrophages, the cGAS/STING pathway is essential not only for the production of type-I interferons in response to intracellular DNA stimulation, but also for regulation of macrophage effector functions including the expression of MHC-II and co-stimulatory molecules. In the context of fowlpox, an avian DNA virus infection, the cGAS/STING pathway was found to be responsible for type-I interferon production and MHC-II transcription. The sensing of fowlpox virus DNA is therefore essential for mounting an anti-viral response in chicken cells and for regulation of a specific set of macrophage effector functions.

scholarly journals Preclinical Studies of a Modified Vaccinia Virus Ankara-Based HIV Candidate Vaccine: Antigen Presentation and Antiviral Effect

2010 ◽  
Vol 84 (10) ◽  
pp. 5314-5328 ◽  
Author(s):  
Samantha Brandler ◽  
Alice Lepelley ◽  
Marion Desdouits ◽  
Florence Guivel-Benhassine ◽  
Pierre-Emmanuel Ceccaldi ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Antiviral Effect ◽  
Vaccine Antigen ◽  
Type I ◽  
Hiv Replication ◽  
National Agency ◽  
Class I Mhc ◽  
Mhc I ◽  
Cross Presentation ◽  
Modified Vaccinia Virus Ankara

ABSTRACT Poxvirus-based human immunodeficiency virus (HIV) vaccine candidates are currently under evaluation in preclinical and clinical trials. Modified vaccinia virus Ankara (MVA) vectors have excellent safety and immunogenicity records, but their behavior in human cell cultures remains only partly characterized. We studied here various virological and immunological aspects of the interactions of MVA-HIV, a vaccine candidate developed by the French National Agency for AIDS Research (ANRS), with primary human cells. We report that MVA-HIV infects and drives Gag expression in primary macrophages, dendritic cells (DCs), and epithelial and muscle cells. MVA-HIV-infected DCs matured, efficiently presented Gag, Pol, and Nef antigens, and activated HIV-specific cytotoxic T lymphocytes (CTLs). As expected with this type of vector, infection was cytopathic and led to DC apoptosis. Coculture of MVA-HIV-infected epithelial cells or myotubes with DCs promoted efficient Gag antigen major histocompatibility complex class I (MHC-I) cross-presentation without inducing direct infection and death of DCs. Antigen-presenting cells (APCs) infected with MVA-HIV also activated HIV-specific CD4+ T cells. Moreover, exposure of DCs to MVA-HIV or to MVA-HIV-infected myotubes induced type I interferon (IFN) production and inhibited subsequent HIV replication and transfer to lymphocytes. Altogether, these results show that MVA-HIV promotes efficient MHC-I and MHC-II presentation of HIV antigens by APCs without facilitating HIV replication. Deciphering the immune responses to MVA in culture experiments will help in the design of innovative vaccine strategies.

scholarly journals Novel Modified Vaccinia Virus Ankara Vector Expressing Anti-apoptotic GeneB13RDelays Apoptosis and Enhances Humoral Responses

2018 ◽  
Vol 93 (5) ◽  
Author(s):  
Lynette S. Chea ◽  
Linda S. Wyatt ◽  
Sailaja Gangadhara ◽  
Bernard Moss ◽  
Rama R. Amara
Keyword(s):  
Vaccinia Virus ◽  
Vaccine Development ◽  
Modified Vaccinia Virus Ankara ◽  
Immunodeficiency Virus ◽  
Antibody Titers ◽  
Infected Cells ◽  
Induced Apoptosis ◽  
Humoral Responses ◽  
Antigen Presenting

ABSTRACTModified vaccinia virus Ankara (MVA), an attenuated poxvirus, has been developed as a potential vaccine vector for use against cancer and multiple infectious diseases, including human immunodeficiency virus (HIV). MVA is highly immunogenic and elicits strong cellular and humoral responses in preclinical models and humans. However, there is potential to further enhance the immunogenicity of MVA, as MVA-infected cells undergo rapid apoptosis, leading to faster clearance of recombinant antigens and potentially blunting a greater response. Here, we generated MVA-B13Rby replacing the fragmented181R/182Rgenes of MVA with a functional anti-apoptotic gene,B13R, and confirmed its anti-apoptotic function against chemically induced apoptosisin vitro. In addition, MVA-B13Rshowed a significant delay in induction of apoptosis in muscle cells derived from mice and humans, as well as in plasmacytoid dendritic cells (pDCs) and CD141+DCs from rhesus macaques, compared to the induction of apoptosis in MVA-infected cells. MVA-B13Rexpressing simian immunodeficiency virus (SIV) Gag and Pol and HIV envelope (SHIV) (MVA-B13R/SHIV) produced higher levels of envelope in the supernatants than MVA/SHIV-infected DF-1 cellsin vitro. Immunization of BALB/c mice showed induction of higher levels of envelope-specific antibody-secreting cells and memory B cells, higher IgG antibody titers, and better persistence of antibody titers with MVA-B13R/SHIV than with MVA/SHIV. Gene set enrichment analysis of draining lymph node cells from day 1 after immunization showed negative enrichment for interferon responses in MVA-B13R/SHIV-immunized mice compared to the responses in MVA/SHIV-immunized mice. Taken together, these results demonstrate that restoringB13Rfunctionality in MVA significantly delays MVA-induced apoptosis in muscle and antigen-presenting cellsin vitroand augments vaccine-induced humoral immunity in mice.IMPORTANCEMVA is an attractive viral vector for vaccine development due to its safety and immunogenicity in multiple species and humans even under conditions of immunodeficiency. Here, to further improve the immunogenicity of MVA, we developed a novel vector, MVA-B13R, by replacing the fragmented anti-apoptotic genes181R/182Rwith a functional version derived from vaccinia virus,B13R. Our results show that MVA-B13Rsignificantly delays apoptosis in antigen-presenting cells and muscle cellsin vitroand augments vaccine-induced humoral immunity in mice, leading to the development of a novel vector for vaccine development against infectious diseases and cancer.

scholarly journals RNA species generated in vaccinia virus infected cells activate cell type-specific MDA5 or RIG-I dependent interferon gene transcription and PKR dependent apoptosis

Virology ◽  
2011 ◽  
Vol 413 (2) ◽  
pp. 183-193 ◽  
Author(s):  
Chad Myskiw ◽  
Janilyn Arsenio ◽  
Evan P. Booy ◽  
Craig Hammett ◽  
Yvon Deschambault ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Gene Transcription ◽  
Cell Type ◽  
Infected Cells ◽  
Cell Type Specific ◽  
Activate Cell ◽  
Interferon Gene

scholarly journals Vaccinia Virus-Mediated Inhibition of Type I Interferon Responses Is a Multifactorial Process Involving the Soluble Type I Interferon Receptor B18 and Intracellular Components

2008 ◽  
Vol 83 (4) ◽  
pp. 1563-1571 ◽  
Author(s):  
Zoe Waibler ◽  
Martina Anzaghe ◽  
Theresa Frenz ◽  
Astrid Schwantes ◽  
Christopher Pöhlmann ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Type I Interferon ◽  
Poly I:C ◽  
Host Responses ◽  
Type I ◽  
Immune Modulators ◽  
Modified Vaccinia Virus Ankara ◽  
Multistep Process ◽  
Interferon Responses

ABSTRACT Poxviruses such as virulent vaccinia virus (VACV) strain Western Reserve encode a broad range of immune modulators that interfere with host responses to infection. Upon more than 570 in vitro passages in chicken embryo fibroblasts (CEF), chorioallantois VACV Ankara (CVA) accumulated mutations that resulted in highly attenuated modified vaccinia virus Ankara (MVA). MVA infection of mice and of dendritic cells (DC) induced significant type I interferon (IFN) responses, whereas infection with VACV alone or in combination with MVA did not. These results implied that VACV expressed an IFN inhibitor(s) that was functionally deleted in MVA. To further characterize the IFN inhibitor(s), infection experiments were carried out with CVA strains isolated after 152 (CVA152) and 386 CEF passages (CVA386). Interestingly, neither CVA152 nor CVA386 induced IFN-α, whereas the latter variant did induce IFN-β. This pattern suggested a consecutive loss of inhibitors during MVA attenuation. Similar to supernatants of VACV- and CVA152-infected DC cultures, recombinantly expressed soluble IFN decoy receptor B18, which is encoded in the VACV genome, inhibited MVA-induced IFN-α but not IFN-β. In the same direction, a B18R-deficient VACV variant triggered only IFN-α, confirming B18 as the soluble IFN-α inhibitor. Interestingly, VACV infection inhibited IFN responses induced by a multitude of different stimuli, including oligodeoxynucleotides containing CpG motifs, poly(I:C), and vesicular stomatitis virus. Collectively, the data presented show that VACV-mediated IFN inhibition is a multistep process involving secreted factors such as B18 plus intracellular components that cooperate to efficiently shut off systemic IFN-α and IFN-β responses.

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