Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 44
Filtrar
Mais filtros










Base de dados
Tipo de estudo
Intervalo de ano de publicação
2.
J Gen Virol ; 100(6): 911-912, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31021739

RESUMO

Members of the family Filoviridae produce variously shaped, often filamentous, enveloped virions containing linear non-segmented, negative-sense RNA genomes of 15-19 kb. Several filoviruses (e.g., Ebola virus) are pathogenic for humans and are highly virulent. Several filoviruses infect bats (e.g., Marburg virus), whereas the hosts of most other filoviruses are unknown. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on Filoviridae, which is available at www.ictv.global/report/filoviridae.

3.
Viruses ; 11(3)2019 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-30893855

RESUMO

Filoviruses infect a wide range of cell types with the exception of lymphocytes. The intracellular proteins cathepsin B and L, two-pore channel 1 and 2, and bona fide receptor Niemann⁻Pick Disease C1 (NPC1) are essential for the endosomal phase of cell entry. However, earlier steps of filoviral infection remain poorly characterized. Numerous plasma membrane proteins have been implicated in attachment but it is still unclear which ones are sufficient for productive entry. To define a minimal set of host factors required for filoviral glycoprotein-driven cell entry, we screened twelve cell lines and identified the nonlymphocytic cell line SH-SY5Y to be specifically resistant to filovirus infection. Heterokaryons of SH-SY5Y cells fused to susceptible cells were susceptible to filoviruses, indicating that SH-SY5Y cells do not express a restriction factor but lack an enabling factor critical for filovirus entry. However, all tested cell lines expressed functional intracellular factors. Global gene expression profiling of known cell surface entry factors and protein expression levels of analyzed attachment factors did not reveal any correlation between susceptibility and expression of a specific host factor. Using binding assays with recombinant filovirus glycoprotein, we identified cell attachment as the step impaired in filovirus entry in SH-SY5Y cells. Individual overexpression of attachment factors T-cell immunoglobulin and mucin domain 1 (TIM-1), Axl, Mer, or dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) rendered SH-SY5Y cells susceptible to filovirus glycoprotein-driven transduction. Our study reveals that a lack of attachment factors limits filovirus entry and provides direct experimental support for a model of filoviral cell attachment where host factor usage at the cell surface is highly promiscuous.

4.
Arch Virol ; 164(4): 1233-1244, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30663023

RESUMO

In October 2018, the order Mononegavirales was amended by the establishment of three new families and three new genera, abolishment of two genera, and creation of 28 novel species. This article presents the updated taxonomy of the order Mononegavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV).


Assuntos
Mononegavirais/classificação , Mononegavirais/genética , Mononegavirais/isolamento & purificação , Filogenia , Virologia/organização & administração
5.
NPJ Vaccines ; 3: 42, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30323953

RESUMO

The recent Ebola virus outbreak has highlighted the therapeutic potential of antisera and renewed interest in this treatment approach. While human convalescent sera may not be readily available in the early stages of an outbreak, antisera of animal origin can be produced in a short time frame. Here, we compared adjuvanted virus-like particles (VLP) with recombinant modified vaccinia virus Ankara and vesicular stomatitis virus (VSV), both expressing the Ebola virus antigens. The neutralizing antibody titers of rabbits immunized with adjuvanted VLPs were similar to those immunized with the replication-competent VSV, indicating that presentation of the antigen in its native conformation rather than de novo antigen expression is essential for production of functional antibodies. This approach also yielded high-titer antisera against Nipah virus glycoproteins, illustrating that it is transferable to other virus families. Multiple-step immunoglobulin G purification using a two-step 20-40% ammonium sulfate precipitation followed by protein A affinity chromatography resulted in 90% recovery of functionality and sustained in vivo stability. Adjuvanted VLP-based immunization strategies are thus a promising approach for the rapid generation of therapeutic antisera against emerging infections.

7.
Arch Virol ; 163(8): 2283-2294, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29637429

RESUMO

In 2018, the order Mononegavirales was expanded by inclusion of 1 new genus and 12 novel species. This article presents the updated taxonomy of the order Mononegavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV) and summarizes additional taxonomic proposals that may affect the order in the near future.


Assuntos
Mononegavirais/classificação , Animais , Humanos , Mononegavirais/genética , Mononegavirais/isolamento & purificação , Infecções por Mononegavirales/veterinária , Infecções por Mononegavirales/virologia , Filogenia
8.
Methods Mol Biol ; 1628: 195-202, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28573621

RESUMO

Observation of molecular processes inside living cells is fundamental to a deeper understanding of virus-host interactions in filoviral-infected cells. These observations can provide spatiotemporal insights into protein synthesis, protein-protein interaction dynamics, and transport processes of these highly pathogenic viruses. Thus, live-cell imaging provides the possibility for antiviral screening in real time and gives mechanistic insights into understanding filovirus assembly steps that are dependent on cellular factors, which then represent potential targets against this highly fatal disease. Here we describe analysis of living filovirus-infected cells under maximum biosafety (i.e., BSL4) conditions using plasmid-driven expression of fluorescently labeled viral and cellular proteins and/or viral genome-encoded expression of fluorescently labeled proteins. Such multiple-color and multidimensional time-lapse live-cell imaging analyses are a powerful method to gain a better understanding of the filovirus infection cycle.


Assuntos
Infecções por Filoviridae/diagnóstico por imagem , Filoviridae/isolamento & purificação , Interações Hospedeiro-Patógeno , Imagem Molecular/métodos , Linhagem Celular , Contenção de Riscos Biológicos , Filoviridae/patogenicidade , Infecções por Filoviridae/virologia , Humanos
9.
Viruses ; 9(5)2017 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-28492506

RESUMO

The mononegaviral family Filoviridae has eight members assigned to three genera and seven species. Until now, genus and species demarcation were based on arbitrarily chosen filovirus genome sequence divergence values (≈50% for genera, ≈30% for species) and arbitrarily chosen phenotypic virus or virion characteristics. Here we report filovirus genome sequence-based taxon demarcation criteria using the publicly accessible PAirwise Sequencing Comparison (PASC) tool of the US National Center for Biotechnology Information (Bethesda, MD, USA). Comparison of all available filovirus genomes in GenBank using PASC revealed optimal genus demarcation at the 55-58% sequence diversity threshold range for genera and at the 23-36% sequence diversity threshold range for species. Because these thresholds do not change the current official filovirus classification, these values are now implemented as filovirus taxon demarcation criteria that may solely be used for filovirus classification in case additional data are absent. A near-complete, coding-complete, or complete filovirus genome sequence will now be required to allow official classification of any novel "filovirus." Classification of filoviruses into existing taxa or determining the need for novel taxa is now straightforward and could even become automated using a presented algorithm/flowchart rooted in RefSeq (type) sequences.


Assuntos
Filoviridae/classificação , Filoviridae/genética , Filogenia , Algoritmos , Sequência de Bases , Bases de Dados de Ácidos Nucleicos , Ebolavirus/classificação , Ebolavirus/genética , Variação Genética , Genoma Viral , Marburgvirus/classificação , Marburgvirus/genética , Mononegavirais/classificação , Mononegavirais/genética , Análise de Sequência de DNA , Desenho de Programas de Computador , Especificidade da Espécie , Sequenciamento Completo do Genoma
10.
Parasit Vectors ; 10(1): 224, 2017 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-28482865

RESUMO

BACKGROUND: Although avian trypanosomes are widespread parasites, the knowledge of their vectors is still incomplete. Despite biting midges (Diptera: Ceratopogonidae) are considered as potential vectors of avian trypanosomes, their role in transmission has not been satisfactorily elucidated. Our aim was to clarify the potential of biting midges to sustain the development of avian trypanosomes by testing their susceptibility to different strains of avian trypanosomes experimentally. Moreover, we screened biting midges for natural infections in the wild. RESULTS: Laboratory-bred biting midges Culicoides nubeculosus were highly susceptible to trypanosomes from the Trypanosoma bennetti and T. avium clades. Infection rates reached 100%, heavy infections developed in 55-87% of blood-fed females. Parasite stages from the insect gut were infective for birds. Moreover, midges could be infected after feeding on a trypanosome-positive bird. Avian trypanosomes can thus complete their cycle in birds and biting midges. Furthermore, we succeeded to find infected blood meal-free biting midges in the wild. CONCLUSIONS: Biting midges are probable vectors of avian trypanosomes belonging to T. bennetti group. Midges are highly susceptible to artificial infections, can be infected after feeding on birds, and T. bennetti-infected biting midges (Culicoides spp.) have been found in nature. Moreover, midges can be used as model hosts producing metacyclic avian trypanosome stages infective for avian hosts.


Assuntos
Doenças das Aves/transmissão , Aves/parasitologia , Ceratopogonidae/parasitologia , Insetos Vetores/parasitologia , Tripanossomíase/veterinária , Animais , Doenças das Aves/parasitologia , Canários/parasitologia , Ceratopogonidae/anatomia & histologia , Trato Gastrointestinal/parasitologia , Especificidade de Hospedeiro , Microscopia Eletrônica de Varredura , Reação em Cadeia da Polimerase , Trypanosoma/classificação , Trypanosoma/genética , Trypanosoma/fisiologia , Trypanosoma/ultraestrutura , Tripanossomíase/diagnóstico , Tripanossomíase/parasitologia , Tripanossomíase/transmissão
11.
Arch Virol ; 162(8): 2493-2504, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28389807

RESUMO

In 2017, the order Mononegavirales was expanded by the inclusion of a total of 69 novel species. Five new rhabdovirus genera and one new nyamivirus genus were established to harbor 41 of these species, whereas the remaining new species were assigned to already established genera. Furthermore, non-Latinized binomial species names replaced all paramyxovirus and pneumovirus species names, thereby accomplishing application of binomial species names throughout the entire order. This article presents the updated taxonomy of the order Mononegavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV).


Assuntos
Genoma Viral , Mononegavirais/classificação , Ordem dos Genes , Mononegavirais/genética , Filogenia , Especificidade da Espécie
12.
Syst Biol ; 66(3): 463-473, 2017 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-27798405

RESUMO

Botanical, mycological, zoological, and prokaryotic species names follow the Linnaean format, consisting of an italicized Latinized binomen with a capitalized genus name and a lower case species epithet (e.g., Homo sapiens). Virus species names, however, do not follow a uniform format, and, even when binomial, are not Linnaean in style. In this thought exercise, we attempted to convert all currently official names of species included in the virus family Arenaviridae and the virus order Mononegavirales to Linnaean binomials, and to identify and address associated challenges and concerns. Surprisingly, this endeavor was not as complicated or time-consuming as even the authors of this article expected when conceiving the experiment. [Arenaviridae; binomials; ICTV; International Committee on Taxonomy of Viruses; Mononegavirales; virus nomenclature; virus taxonomy.].


Assuntos
Classificação , Vírus , Terminologia como Assunto
13.
Arch Virol ; 161(8): 2351-60, 2016 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-27216929

RESUMO

In 2016, the order Mononegavirales was emended through the addition of two new families (Mymonaviridae and Sunviridae), the elevation of the paramyxoviral subfamily Pneumovirinae to family status (Pneumoviridae), the addition of five free-floating genera (Anphevirus, Arlivirus, Chengtivirus, Crustavirus, and Wastrivirus), and several other changes at the genus and species levels. This article presents the updated taxonomy of the order Mononegavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV).


Assuntos
Genoma Viral , Mononegavirais/classificação , Mononegavirais/genética , Filogenia
14.
J Infect Dis ; 212 Suppl 2: S226-33, 2015 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-26138826

RESUMO

Synthesis of the surface glycoprotein GP of Ebola virus (EBOV) is dependent on transcriptional RNA editing, whereas direct expression of the GP gene results in synthesis of nonstructural secreted glycoprotein sGP. In this study, we investigate the role of RNA editing in the pathogenicity of EBOV using a guinea pig model and recombinant guinea pig-adapted EBOV containing mutations at the editing site, allowing expression of surface GP without the need for RNA editing, and also preventing synthesis of sGP. We demonstrate that the elimination of the editing site leads to EBOV attenuation in vivo, explained by lower virus spread caused by the higher virus cytotoxicity and, most likely, by an increased ability of the host defense systems to recognize and eliminate virus-infected cells. We also demonstrate that expression of sGP does not affect pathogenicity of EBOV in guinea pigs. In conclusion, data obtained indicate that downregulation of the level of surface GP expression through a mechanism of GP gene RNA editing plays an important role in the high pathogenicity of EBOV.


Assuntos
Ebolavirus/genética , Genes Virais/genética , Doença pelo Vírus Ebola/virologia , Edição de RNA/genética , Proteínas do Envelope Viral/genética , Proteínas Virais/genética , Fatores de Virulência/genética , Animais , Linhagem Celular , Regulação para Baixo/genética , Ebolavirus/patogenicidade , Regulação Viral da Expressão Gênica/genética , Cobaias , Glicoproteínas de Membrana/genética , Mutação/genética
15.
J Infect Dis ; 212 Suppl 2: S322-8, 2015 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-26092855

RESUMO

The surface glycoprotein (GP) is responsible for Ebola virus (EBOV) attachment and membrane fusion during virus entry. Surface expression of highly glycosylated GP causes marked cytotoxicity via masking of a wide range of cellular surface molecules, including integrins. Considerable amounts of surface GP are shed from virus-infected cells in a soluble truncated form by tumor necrosis factor α-converting enzyme. In this study, the role of GP shedding was investigated using a reverse genetics approach by comparing recombinant viruses possessing amino acid substitutions at the GP shedding site. Virus with an L635V substitution showed a substantial decrease in shedding, whereas a D637V substitution resulted in a striking increase in the release of shed GP. Variations in shedding efficacy correlated with observed differences in the amounts of shed GP in the medium, GP present in virus-infected cells, and GP present on virions. An increase in shedding appeared to be associated with a reduction in viral cytotoxicity, and, vice versa, the virus that shed less was more cytotoxic. An increase in shedding also resulted in a reduction in viral infectivity, whereas a decrease in shedding efficacy enhanced viral growth characteristics in vitro. Differences in shedding efficacy and, as a result, differences in the amount of mature GP available for incorporation into budding virions did not equate to differences in overall release of viral particles. Likewise, data suggest that the resulting differences in the amount of mature GP on the cell surface led to variations in the GP content of released particles and, as a consequence, in infectivity. In conclusion, fine-tuning of the levels of EBOV GP expressed at the surface of virus-infected cells via GP shedding plays an important role in EBOV replication by orchestrating the balance between optimal virion GP content and cytotoxicity caused by GP.


Assuntos
Ebolavirus/metabolismo , Ebolavirus/patogenicidade , Doença pelo Vírus Ebola/virologia , Glicoproteínas de Membrana/metabolismo , Substituição de Aminoácidos/genética , Animais , Linhagem Celular , Cercopithecus aethiops , Ebolavirus/genética , Glicoproteínas de Membrana/genética , Células Vero , Proteínas Virais/genética , Proteínas Virais/metabolismo , Vírion/genética , Vírion/metabolismo , Vírion/patogenicidade , Virulência/genética , Internalização do Vírus , Replicação Viral/genética
16.
J Infect Dis ; 212 Suppl 2: S160-6, 2015 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-26038396

RESUMO

BACKGROUND: Transport of ebolavirus (EBOV) nucleocapsids from perinuclear viral inclusions, where they are formed, to the site of budding at the plasma membrane represents an obligatory step of virus assembly. Until now, no live-cell studies on EBOV nucleocapsid transport have been performed, and participation of host cellular factors in this process, as well as the trajectories and speed of nucleocapsid transport, remain unknown. METHODS: Live-cell imaging of EBOV-infected cells treated with different inhibitors of cellular cytoskeleton was used for the identification of cellular proteins involved in the nucleocapsid transport. EBOV nucleocapsids were visualized by expression of green fluorescent protein (GFP)-labeled nucleocapsid viral protein 30 (VP30) in EBOV-infected cells. RESULTS: Incorporation of the fusion protein VP30-GFP into EBOV nucleocapsids was confirmed by Western blot and indirect immunofluorescence analyses. Importantly, VP30-GFP fluorescence was readily detectable in the densely packed nucleocapsids inside perinuclear viral inclusions and in the dispersed rod-like nucleocapsids located outside of viral inclusions. Live-cell imaging of EBOV-infected cells revealed exit of single nucleocapsids from the viral inclusions and their intricate transport within the cytoplasm before budding at the plasma membrane. Nucleocapsid transport was arrested upon depolymerization of actin filaments (F-actin) and inhibition of the actin-nucleating Arp2/3 complex, and it was not altered upon depolymerization of microtubules or inhibition of N-WASP. Actin comet tails were often detected at the rear end of nucleocapsids. Marginally located nucleocapsids entered filopodia, moved inside, and budded from the tip of these thin cellular protrusions. CONCLUSIONS: Live-cell imaging of EBOV-infected cells revealed actin-dependent long-distance transport of EBOV nucleocapsids before budding at the cell surface. These findings provide useful insights into EBOV assembly and have potential application in the development of antivirals.


Assuntos
Actinas/metabolismo , Transporte Biológico/fisiologia , Ebolavirus/metabolismo , Nucleocapsídeo/metabolismo , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Citoplasma/metabolismo , Citoesqueleto/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Humanos , Corpos de Inclusão/metabolismo , Pseudópodes/metabolismo , Fatores de Transcrição/metabolismo , Proteínas Virais/metabolismo
17.
Eur J Cell Biol ; 94(7-9): 323-31, 2015 Jul-Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26070789

RESUMO

Filovirus infection of target cells leads to the formation of virally induced cytoplasmic inclusions that contain viral nucleocapsids at different stages of maturation. While the role of the inclusions has been unclear since the identification of Marburg and Ebola viruses, it recently became clear that the inclusions are the sites of viral replication, nucleocapsid formation and maturation. Live cell imaging analyses revealed that mature nucleocapsids are transported from inclusions to the filopodia, which represent the major budding sites. Moreover, inclusions recruit cellular proteins that have been shown to support the transport of nucleocapsids. For example, the tumor susceptibility gene 101 protein (Tsg101) interacts with a late domain motif in the nucleocapsid protein NP and recruits the actin-nucleation factor IQGAP1. Complexes of nucleocapsids together with Tsg101 and IQGAP1 are then co-transported along actin filaments. We detected additional proteins (Alix, Nedd4 and the AAA-type ATPase VPS4) of the endosomal sorting complex required for transport (ESCRT) that are recruited into inclusions. Together, the results suggest that nucleocapsids recruit the machinery that enhances viral budding at the plasma membrane. Furthermore, we identified Lamp1 as a marker of the late endosomal compartment in inclusions, while ER, Golgi, TGN and early endosomal markers were absent. In addition, we observed that LC3, a marker of autophagosomal membranes, was present in inclusions. The 3D structures of inclusions show an intricate structure that seems to accommodate an intimate cooperation between cellular and viral components with the intention to support viral transport and budding.


Assuntos
Compartimento Celular/fisiologia , Endossomos/metabolismo , Corpos de Inclusão Viral/metabolismo , Marburgvirus/crescimento & desenvolvimento , Corpos Multivesiculares/fisiologia , Animais , Linhagem Celular , Proteínas de Ligação a DNA/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Humanos , Glicoproteínas de Membrana Associadas ao Lisossomo/metabolismo , Macrófagos/virologia , Doença do Vírus de Marburg/virologia , Nucleocapsídeo/biossíntese , Nucleocapsídeo/metabolismo , Nucleoproteínas/metabolismo , Transporte Proteico , Pseudópodes/metabolismo , Fatores de Transcrição/metabolismo , Liberação de Vírus/fisiologia , Replicação Viral/fisiologia , Proteínas Ativadoras de ras GTPase/metabolismo
18.
J Virol ; 89(7): 3859-69, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25609809

RESUMO

UNLABELLED: The Middle East respiratory syndrome coronavirus (MERS-CoV) emerged in 2012 as the causative agent of a severe respiratory disease with a fatality rate of approximately 30%. The high virulence and mortality rate prompted us to analyze aspects of MERS-CoV pathogenesis, especially its interaction with innate immune cells such as antigen-presenting cells (APCs). Particularly, we analyzed secretion of type I and type III interferons (IFNs) by APCs, i.e., B cells, macrophages, monocyte-derived/myeloid dendritic cells (MDDCs/mDCs), and by plasmacytoid dendritic cells (pDCs) of human and murine origin after inoculation with MERS-CoV. Production of large amounts of type I and III IFNs was induced exclusively in human pDCs, which were significantly higher than IFN induction by severe acute respiratory syndrome (SARS)-CoV. Of note, IFNs were secreted in the absence of productive replication. However, receptor binding, endosomal uptake, and probably signaling via Toll-like receptor 7 (TLR7) were critical for sensing of MERS-CoV by pDCs. Furthermore, active transcription of MERS-CoV N RNA and subsequent N protein expression were evident in infected pDCs, indicating abortive infection. Taken together, our results point toward dipeptidyl peptidase 4 (DPP4)-dependent endosomal uptake and subsequent infection of human pDCs by MERS-CoV. However, the replication cycle is stopped after early gene expression. In parallel, human pDCs are potent IFN-producing cells upon MERS-CoV infection. Knowledge of such IFN responses supports our understanding of MERS-CoV pathogenesis and is critical for the choice of treatment options. IMPORTANCE: MERS-CoV causes a severe respiratory disease with high fatality rates in human patients. Recently, confirmed human cases have increased dramatically in both number and geographic distribution. Understanding the pathogenesis of this highly pathogenic CoV is crucial for developing successful treatment strategies. This study elucidates the interaction of MERS-CoV with APCs and pDCs, particularly the induction of type I and III IFN secretion. Human pDCs are the immune cell population sensing MERS-CoV but secrete significantly larger amounts of IFNs, especially IFN-α, than in response to SARS-CoV. A model for molecular virus-host interactions is presented outlining IFN induction in pDCs. The massive IFN secretion upon contact suggests a critical role of this mechanism for the high degree of immune activation observed during MERS-CoV infection.


Assuntos
Células Dendríticas/imunologia , Células Dendríticas/virologia , Interferons/metabolismo , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Animais , Dipeptidil Peptidase 4/metabolismo , Endocitose , Endossomos/metabolismo , Endossomos/virologia , Humanos , Camundongos Endogâmicos C57BL , Internalização do Vírus
19.
PLoS Pathog ; 10(11): e1004509, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25412102

RESUMO

During Ebola virus (EBOV) infection a significant amount of surface glycoprotein GP is shed from infected cells in a soluble form due to cleavage by cellular metalloprotease TACE. Shed GP and non-structural secreted glycoprotein sGP, both expressed from the same GP gene, have been detected in the blood of human patients and experimentally infected animals. In this study we demonstrate that shed GP could play a particular role during EBOV infection. In effect it binds and activates non-infected dendritic cells and macrophages inducing the secretion of pro- and anti-inflammatory cytokines (TNFα, IL1ß, IL6, IL8, IL12p40, and IL1-RA, IL10). Activation of these cells by shed GP correlates with the increase in surface expression of co-stimulatory molecules CD40, CD80, CD83 and CD86. Contrary to shed GP, secreted sGP activates neither DC nor macrophages while it could bind DCs. In this study, we show that shed GP activity is likely mediated through cellular toll-like receptor 4 (TLR4) and is dependent on GP glycosylation. Treatment of cells with anti-TLR4 antibody completely abolishes shed GP-induced activation of cells. We also demonstrate that shed GP activity is negated upon addition of mannose-binding sera lectin MBL, a molecule known to interact with sugar arrays present on the surface of different microorganisms. Furthermore, we highlight the ability of shed GP to affect endothelial cell function both directly and indirectly, demonstrating the interplay between shed GP, systemic cytokine release and increased vascular permeability. In conclusion, shed GP released from virus-infected cells could activate non-infected DCs and macrophages causing the massive release of pro- and anti-inflammatory cytokines and effect vascular permeability. These activities could be at the heart of the excessive and dysregulated inflammatory host reactions to infection and thus contribute to high virus pathogenicity.


Assuntos
Células Dendríticas/imunologia , Ebolavirus/imunologia , Doença pelo Vírus Ebola/imunologia , Células Endoteliais da Veia Umbilical Humana/imunologia , Macrófagos/imunologia , Glicoproteínas de Membrana/imunologia , Proteínas Virais/imunologia , Animais , Antígenos CD/imunologia , Citocinas/imunologia , Células Dendríticas/patologia , Células Dendríticas/virologia , Cobaias , Células Endoteliais da Veia Umbilical Humana/patologia , Células Endoteliais da Veia Umbilical Humana/virologia , Humanos , Macrófagos/patologia , Macrófagos/virologia , Receptor 4 Toll-Like/imunologia
20.
PLoS Pathog ; 10(10): e1004463, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25330247

RESUMO

Endosomal sorting complex required for transport (ESCRT) machinery supports the efficient budding of Marburg virus (MARV) and many other enveloped viruses. Interaction between components of the ESCRT machinery and viral proteins is predominantly mediated by short tetrapeptide motifs, known as late domains. MARV contains late domain motifs in the matrix protein VP40 and in the genome-encapsidating nucleoprotein (NP). The PSAP late domain motif of NP recruits the ESCRT-I protein tumor susceptibility gene 101 (Tsg101). Here, we generated a recombinant MARV encoding NP with a mutated PSAP late domain (rMARV(PSAPmut)). rMARV(PSAPmut) was attenuated by up to one log compared with recombinant wild-type MARV (rMARV(wt)), formed smaller plaques and exhibited delayed virus release. Nucleocapsids in rMARV(PSAPmut)-infected cells were more densely packed inside viral inclusions and more abundant in the cytoplasm than in rMARV(wt)-infected cells. A similar phenotype was detected when MARV-infected cells were depleted of Tsg101. Live-cell imaging analyses revealed that Tsg101 accumulated in inclusions of rMARV(wt)-infected cells and was co-transported together with nucleocapsids. In contrast, rMARV(PSAPmut) nucleocapsids did not display co-localization with Tsg101, had significantly shorter transport trajectories, and migration close to the plasma membrane was severely impaired, resulting in reduced recruitment into filopodia, the major budding sites of MARV. We further show that the Tsg101 interacting protein IQGAP1, an actin cytoskeleton regulator, was recruited into inclusions and to individual nucleocapsids together with Tsg101. Moreover, IQGAP1 was detected in a contrail-like structure at the rear end of migrating nucleocapsids. Down regulation of IQGAP1 impaired release of MARV. These results indicate that the PSAP motif in NP, which enables binding to Tsg101, is important for the efficient actin-dependent transport of nucleocapsids to the sites of budding. Thus, the interaction between NP and Tsg101 supports several steps of MARV assembly before virus fission.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Marburgvirus , Nucleocapsídeo/metabolismo , Ribonucleoproteínas/metabolismo , Fatores de Transcrição/metabolismo , Proteínas Virais/metabolismo , Citoesqueleto de Actina/metabolismo , Animais , Humanos , Transporte Proteico/fisiologia , Liberação de Vírus/fisiologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA