Host-Specific NS5 Ubiquitination Determines Yellow Fever Virus Tropism

ABSTRACT The recent yellow fever virus (YFV) epidemic in Brazil in 2017 and Zika virus (ZIKV) epidemic in 2015 serve to remind us of the importance of flaviviruses as emerging human pathogens. With the current global flavivirus threat, there is an urgent need for antivirals and vaccines to curb the spread of these viruses. However, the lack of suitable animal models limits the research questions that can be answered. A common trait of all flaviviruses studied thus far is their ability to antagonize interferon (IFN) signaling so as to enhance viral replication and dissemination. Previously, we reported that YFV NS5 requires the presence of type I IFN (IFN-α/β) for its engagement with human signal transducer and activator of transcription 2 (hSTAT2). In this manuscript, we report that like the NS5 proteins of ZIKV and dengue virus (DENV), YFV NS5 protein is able to bind hSTAT2 but not murine STAT2 (mSTAT2). Contrary to what has been demonstrated with ZIKV NS5 and DENV NS5, replacing mSTAT2 with hSTAT2 cannot rescue the YFV NS5-STAT2 interaction, as YFV NS5 is also unable to interact with hSTAT2 in murine cells. We show that the IFN-α/β-dependent ubiquitination of YFV NS5 that is required for STAT2 binding in human cells is absent in murine cells. In addition, we demonstrate that mSTAT2 restricts YFV replication in vivo. These data serve as further impetus for the development of an immunocompetent mouse model that can serve as a disease model for multiple flaviviruses. IMPORTANCE Flaviviruses such as yellow fever virus (YFV), Zika virus (ZIKV), and dengue virus (DENV) are important human pathogens. A common flavivirus trait is the antagonism of interferon (IFN) signaling to enhance viral replication and spread. We report that like ZIKV NS5 and DENV NS5, YFV NS5 binds human STAT2 (hSTAT2) but not mouse STAT2 (mSTAT2), a type I IFN (IFN-α/β) pathway component. Additionally, we show that contrary to what has been demonstrated with ZIKV NS5 and DENV NS5, YFV NS5 is unable to interact with hSTAT2 in murine cells. We demonstrate that mSTAT2 restricts YFV replication in mice and that this correlates with a lack of IFN-α/β-induced YFV NS5 ubiquitination in murine cells. The lack of suitable animal models limits flavivirus pathogenesis, vaccine, and drug research. These data serve as further impetus for the development of an immunocompetent mouse model that can serve as a disease model for multiple flaviviruses.

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Dengue and Zika virus 5’-UTRs harbor IRES functions

10.1101/557942 ◽

2019 ◽

Author(s):

Yutong Song ◽

JoAnn Mugavero ◽

Charles B. Stauft ◽

Eckard Wimmer

Keyword(s):

Yellow Fever ◽

Zika Virus ◽

Yellow Fever Virus ◽

Fever Virus ◽

Human Populations ◽

Human Pathogens ◽

West Nile Fever ◽

West Nile ◽

Ribosomal Entry ◽

Dependent Mechanism

AbstractMembers ofFlavivirus, a genus ofFlaviviridae, encompass numerous enveloped plus strand RNA viruses, of which globally dengue virus (DENV) is the leading cause of serious arthropod-borne disease. The genomes of DENV, just as those of yellow fever virus (YFV), West Nile fever virus (WNV), or Zika virus (ZIKV), control their translation by a 5’-terminal capping group. Three other genera of Flaviviridae are remarkable because their viruses use internal ribosomal entry sites (IRESs) to control translation and they are not arthropod transmitted. In 2006 E. Harris’ group published work suggesting that DENV RNA does not stringently need a cap for translation. They proposed that instead DENV translation is controlled by an interplay between 5’ and 3’ termini. Here we present evidence that the DENV or ZIKV 5’-untranslated regions (5’-UTRs) alone have IRES competence. This conclusion is based, first, on the observation that uncapped mono-cistronic mRNAs 5’ terminated with the DENV or ZIKV 5’-UTRs can efficiently direct translation of a reporter gene in BHK and C6/36 cells; second, that either 5’-UTR placed between two reporter genes can efficiently induce expression of the downstream gene in BHK but not in C6/36 cells. These experiments followed observations that uncapped DENV/ZIKV genomic transcripts, 5’ terminated with pppAN… or GpppAN…, can initiate infections of mammalian (BHK) or mosquito (C6/36) cells. IRES competence of the 5’-UTRs of DENV/ZIKV raises many open questions regarding the biology and control, as well as the evolution, of insect-borne flaviviruses.ImportanceMembers of the genusFlavivirusofFlaviviridaeare important human pathogens of great concern because they cause serious diseases, sometimes death, in human populations living in tropical, subtropical (dengue, DENV; Zika, ZIKV; yellow fever virus), or moderate climates (West Nile virus). Flaviviruses are known to control their translation by a cap-dependent mechanism. We have observed, however, that the uncapped genomes of DENV or ZIKV can initiate infection of mammalian and insect cells. We provide evidence that the short 5’ untranslated region (5’-UTR) of DENV or ZIKV genomes can fulfill the function of an internal ribosomal entry site (IRES). This strategy frees these organisms from the cap-dependent mechanism of gene expression at an as yet unknown stage of proliferation. The data raise new questions about the biology and evolution of flaviviruses, possibly leading to new controls of flavivirus disease.

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Zika Virus: A Clinical Review

KYAMC Journal ◽

10.3329/kyamcj.v7i1.33766 ◽

2017 ◽

Vol 7 (1) ◽

pp. 719-725

Author(s):

Md Daharul Islam ◽

SM Tajdit Rahman ◽

Khaleda Akhter ◽

Md Azizul Hoque ◽

Anannya Roy ◽

...

Keyword(s):

Yellow Fever ◽

Zika Virus ◽

Yellow Fever Virus ◽

Virus Disease ◽

Specific Treatment ◽

Abrupt Onset ◽

Fever Virus ◽

Control Measures ◽

Disease Treatment ◽

Antibodies Detection

Zika virus is a flavivirus related to Dengue virus, yellow fever virus and West Nile virus. It is considered an emerging arbovirus transmitted by mosquito of the genus Aedes. Its first description took place in 1947 in the Zika Forest in Uganda, isolated on Rhesus monkey used as bait to study the yellow fever virus. Clinical picture is characterized as a 'dengue-like' syndrome, with abrupt onset of fever; and an early onset of evanescent rash, often pruritic. Occasionally the disease has been associated with Guillain-Barré syndrome. The diagnosis can be performed by PCR or by IgG and IgM antibodies detection. No specific treatment or vaccine is available for Zika virus disease. Treatment is generally supportive. Control measures are same for dengue and chikungunya based mostly on health education and vector control.KYAMC Journal Vol. 7, No.-1, Jul 2016, Page 719-725

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Simultaneous detection of Dengue virus, Chikungunya virus, Zika virus, Yellow fever virus and West Nile virus

Journal of Virological Methods ◽

10.1016/j.jviromet.2019.03.014 ◽

2019 ◽

Vol 268 ◽

pp. 53-55 ◽

Cited By ~ 3

Author(s):

José A. Boga ◽

Marta E. Alvarez-Arguelles ◽

Susana Rojo-Alba ◽

Mercedes Rodríguez ◽

María de Oña ◽

...

Keyword(s):

West Nile Virus ◽

Dengue Virus ◽

Yellow Fever ◽

Zika Virus ◽

Chikungunya Virus ◽

Yellow Fever Virus ◽

Simultaneous Detection ◽

Fever Virus ◽

West Nile ◽

Virus Zika

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Yellow Fever Virus/Dengue-2 Virus and Yellow Fever Virus/Dengue-4 Virus Chimeras: Biological Characterization, Immunogenicity, and Protection against Dengue Encephalitis in the Mouse Model

Journal of Virology ◽

10.1128/jvi.77.6.3655-3668.2003 ◽

2003 ◽

Vol 77 (6) ◽

pp. 3655-3668 ◽

Cited By ~ 33

Author(s):

Thomas J. Chambers ◽

Yan Liang ◽

Deborah A. Droll ◽

Jacob J. Schlesinger ◽

Andrew D. Davidson ◽

...

Keyword(s):

Mouse Model ◽

Dengue Virus ◽

Yellow Fever ◽

Yellow Fever Virus ◽

Fever Virus ◽

E Proteins ◽

Dengue Viruses ◽

Intracerebral Inoculation ◽

Intraperitoneal Route ◽

Chimeric Viruses

ABSTRACT Two yellow fever virus (YFV)/dengue virus chimeras which encode the prM and E proteins of either dengue virus serotype 2 (dengue-2 virus) or dengue-4 virus within the genome of the YFV 17D strain (YF5.2iv infectious clone) were constructed and characterized for their properties in cell culture and as experimental vaccines in mice. The prM and E proteins appeared to be properly processed and glycosylated, and in plaque reduction neutralization tests and other assays of antigenic specificity, the E proteins exhibited profiles which resembled those of the homologous dengue virus serotypes. Both chimeric viruses replicated in cell lines of vertebrate and mosquito origin to levels comparable to those of homologous dengue viruses but less efficiently than the YF5.2iv parent. YFV/dengue-4 virus, but not YFV/dengue-2 virus, was neurovirulent for 3-week-old mice by intracerebral inoculation; however, both viruses were attenuated when administered by the intraperitoneal route in mice of that age. Single-dose inoculation of either chimeric virus at a dose of 105 PFU by the intraperitoneal route induced detectable levels of neutralizing antibodies against the homologous dengue virus strains. Mice which had been immunized in this manner were fully protected from challenge with homologous neurovirulent dengue viruses by intracerebral inoculation compared to unimmunized mice. Protection was associated with significant increases in geometric mean titers of neutralizing antibody compared to those for unimmunized mice. These data indicate that YFV/dengue virus chimeras elicit antibodies which represent protective memory responses in the mouse model of dengue encephalitis. The levels of neurovirulence and immunogenicity of the chimeric viruses in mice correlate with the degree of adaptation of the dengue virus strain to mice. This study supports ongoing investigations concerning the use of this technology for development of a live attenuated viral vaccine against dengue viruses.

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T Cell-Mediated Immunity towards Yellow Fever Virus and Useful Animal Models

Viruses ◽

10.3390/v9040077 ◽

2017 ◽

Vol 9 (4) ◽

pp. 77 ◽

Cited By ~ 13

Author(s):

Alan Watson ◽

William Klimstra

Keyword(s):

T Cell ◽

Animal Models ◽

Yellow Fever ◽

Yellow Fever Virus ◽

Fever Virus ◽

Cell Mediated Immunity

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A Mouse Model for Studying Viscerotropic Disease Caused by Yellow Fever Virus Infection

PLoS Pathogens ◽

10.1371/journal.ppat.1000614 ◽

2009 ◽

Vol 5 (10) ◽

pp. e1000614 ◽

Cited By ~ 82

Author(s):

Kathryn C. Meier ◽

Christina L. Gardner ◽

Mikhail V. Khoretonenko ◽

William B. Klimstra ◽

Kate D. Ryman

Keyword(s):

Virus Infection ◽

Mouse Model ◽

Yellow Fever ◽

Yellow Fever Virus ◽

Fever Virus

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A Yellow Fever Virus 17D Infection and Disease Mouse Model Used to Evaluate a Chimeric Binjari-Yellow Fever Virus Vaccine

Vaccines ◽

10.3390/vaccines8030368 ◽

2020 ◽

Vol 8 (3) ◽

pp. 368 ◽

Cited By ~ 3

Author(s):

Kexin Yan ◽

Laura J. Vet ◽

Bing Tang ◽

Jody Hobson-Peters ◽

Daniel J. Rawle ◽

...

Keyword(s):

Weight Loss ◽

Mouse Model ◽

Yellow Fever ◽

Virus Vaccine ◽

Neutralizing Antibodies ◽

Yellow Fever Virus ◽

Fever Virus ◽

Liver Pathology ◽

Vaccine Production ◽

Mosquito Cells

Despite the availability of an effective, live attenuated yellow fever virus (YFV) vaccine (YFV 17D), this flavivirus still causes up to ≈60,000 deaths annually. A number of new approaches are seeking to address vaccine supply issues and improve safety for the immunocompromised vaccine recipients. Herein we describe an adult female IFNAR-/- mouse model of YFV 17D infection and disease that recapitulates many features of infection and disease in humans. We used this model to evaluate a new YFV vaccine that is based on a recently described chimeric Binjari virus (BinJV) vaccine technology. BinJV is an insect-specific flavivirus and the chimeric YFV vaccine (BinJ/YFV-prME) was generated by replacing the prME genes of BinJV with the prME genes of YFV 17D. Such BinJV chimeras retain their ability to replicate to high titers in C6/36 mosquito cells (allowing vaccine production), but are unable to replicate in vertebrate cells. Vaccination with adjuvanted BinJ/YFV-prME induced neutralizing antibodies and protected mice against infection, weight loss and liver pathology after YFV 17D challenge.

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E Protein Domain III Determinants of Yellow Fever Virus 17D Vaccine Strain Enhance Binding to Glycosaminoglycans, Impede Virus Spread, and Attenuate Virulence

Journal of Virology ◽

10.1128/jvi.02509-07 ◽

2008 ◽

Vol 82 (12) ◽

pp. 6024-6033 ◽

Cited By ~ 62

Author(s):

Eva Lee ◽

Mario Lobigs

Keyword(s):

Yellow Fever ◽

Yellow Fever Virus ◽

Fever Virus ◽

Protein Domain ◽

Type I ◽

Wild Type ◽

E Protein ◽

Virulence Attenuation ◽

Domain Iii

ABSTRACT The yellow fever virus (YFV) 17D strain is one of the most effective live vaccines for human use, but the in vivo mechanisms for virulence attenuation of the vaccine and the corresponding molecular determinants remain elusive. The vaccine differs phenotypically from wild-type YFV by the loss of viscerotropism, despite replicative fitness in cell culture, and genetically by 20 amino acid changes predominantly located in the envelope (E) protein. We show that three residues in E protein domain III inhibit spread of 17D in extraneural tissues and attenuate virulence in type I/II interferon-deficient mice. One of these residues (Arg380) is a dominant glycosaminoglycan-binding determinant, which mainly accounts for more rapid in vivo clearance of 17D from the bloodstream in comparison to 17D-derived variants with wild-type-like E protein. While other mutations will account for loss of neurotropism and phenotypic stability, the described impact of E protein domain III changes on virus dissemination and virulence is the first rational explanation for the safety of the 17D vaccine in humans.

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