scholarly journals Two African Swine Fever Virus Proteins Derived from a Common Precursor Exhibit Different Nucleocytoplasmic Transport Activities

2004 ◽  
Vol 78 (18) ◽  
pp. 9731-9739 ◽  
Author(s):  
A. Eulálio ◽  
I. Nunes-Correia ◽  
A. L. Carvalho ◽  
C. Faro ◽  
V. Citovsky ◽  
...  
Keyword(s):  
Nuclear Import ◽  
Mammalian Cells ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Nucleocytoplasmic Transport ◽  
Proteolytic Processing ◽  
Fever Virus ◽  
Hemorrhagic Disease ◽  
Common Precursor ◽  
Green Fluorescent

ABSTRACT African swine fever virus (ASFV), a large icosahedral deoxyvirus, is the causative agent of an economically relevant hemorrhagic disease that affects domestic pigs. The major purpose of the present study was to investigate the nuclear transport activities of the ASFV p37 and p14 proteins, which result from the proteolytic processing of a common precursor. Experiments were performed by using yeast-based nucleocytoplasmic transport assays and by analysis of the subcellular localization of different green fluorescent and Myc fusion proteins in mammalian cells. The results obtained both in yeast and mammalian cells clearly demonstrated that ASFV p14 protein is imported into the nucleus but not exported to the cytoplasm. The ability of p37 protein to be exported from the nucleus to the cytoplasm of both yeast and mammalian cells was also demonstrated, and the results clearly indicate that p37 nuclear export is dependent on the interaction of the protein with the CRM-1 receptor. In addition, p37 was shown to exhibit nuclear import activity in mammalian cells. The p37 protein nuclear import and export abilities described here constitute the first report of a nucleocytoplasmic shuttling protein encoded by the ASFV genome. Overall, the overlapping results obtained for green fluorescent protein fusions and Myc-tagged proteins undoubtedly demonstrate that ASFV p37 and p14 proteins exhibit nucleocytoplasmic transport activities. These findings are significant for understanding the role these proteins play in the replication cycle of ASFV.

scholarly journals Computational Analysis of African Swine Fever Virus Protein Space for the Design of an Epitope-Based Vaccine Ensemble

Pathogens ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 1078 ◽  
Author(s):  
Albert Ros-Lucas ◽  
Florencia Correa-Fiz ◽  
Laia Bosch-Camós ◽  
Fernando Rodriguez ◽  
Julio Alonso-Padilla
Keyword(s):  
T Cell ◽  
Vaccine Development ◽  
De Novo ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Fever Virus ◽  
Economic Losses ◽  
Virus Protein ◽  
Hemorrhagic Disease ◽  
Starting Point

African swine fever virus is the etiological agent of African swine fever, a transmissible severe hemorrhagic disease that affects pigs, causing massive economic losses. There is neither a treatment nor a vaccine available, and the only method to control its spread is by extensive culling of pigs. So far, classical vaccine development approaches have not yielded sufficiently good results in terms of concomitant safety and efficacy. Nowadays, thanks to advances in genomic and proteomic techniques, a reverse vaccinology strategy can be explored to design alternative vaccine formulations. In this study, ASFV protein sequences were analyzed using an in-house pipeline based on publicly available immunoinformatic tools to identify epitopes of interest for a prospective vaccine ensemble. These included experimentally validated sequences from the Immune Epitope Database, as well as de novo predicted sequences. Experimentally validated and predicted epitopes were prioritized following a series of criteria that included evolutionary conservation, presence in the virulent and currently circulating variant Georgia 2007/1, and lack of identity to either the pig proteome or putative proteins from pig gut microbiota. Following this strategy, 29 B-cell, 14 CD4+ T-cell and 6 CD8+ T-cell epitopes were selected, which represent a starting point to investigating the protective capacity of ASFV epitope-based vaccines.

Pathogenesis of African swine fever virus inOrnithodorosticks

2001 ◽  
Vol 2 (2) ◽  
pp. 121-128 ◽  
Author(s):  
Steven B. Kleiboeker ◽  
Glen A. Scoles
Keyword(s):  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Fever Virus ◽  
Sub Saharan Africa ◽  
Hemorrhagic Disease ◽  
Sylvatic Cycle ◽  
Infectious Dose ◽  
Domestic Pigs ◽  
Ornithodoros Porcinus Porcinus ◽  
Sub Saharan

AbstractAfrican swine fever virus (ASFV) is the only known DNA arbovirus and the sole member of the family Asfarviridae. It causes a lethal, hemorrhagic disease in domestic pigs. ASFV is enzootic in sub-Saharan Africa and is maintained in a sylvatic cycle by infecting both wild members of the Suidae (e.g. warthogs) and the argasid tickOrnithodoros porcinus porcinus. The pathogenesis of ASFV inO. porcinus porcinusticks is characterized by a low infectious dose, lifelong infection, efficient transmission to both pigs and ticks, and low mortality until after the first oviposition. ASFV pathogenesis in warthogs is characterized by an inapparent infection with transient, low viremic titers. ThusO. porcinus porcinusticks probably constitute the most important natural vector of ASFV, although both the mammalian and tick hosts are probably required for the maintenance of ASFV in the sylvatic cycle. The mechanism of ASFV transmission from the sylvatic cycle to domestic pigs is probably through infected ticks feeding on pigs. In addition toO. porcinus porcinus, a number of North American, Central American and Caribbean species ofOrnithodoroshave been shown to be potential vectors of ASFV.

scholarly journals African Swine Fever Virus Protein p17 Is Essential for the Progression of Viral Membrane Precursors toward Icosahedral Intermediates

2010 ◽  
Vol 84 (15) ◽  
pp. 7484-7499 ◽  
Author(s):  
Cristina Suárez ◽  
Javier Gutiérrez-Berzal ◽  
Germán Andrés ◽  
María L. Salas ◽  
Javier M. Rodríguez
Keyword(s):  
Early Stage ◽  
African Swine Fever Virus ◽  
Transmembrane Protein ◽  
African Swine Fever ◽  
Integral Membrane Protein ◽  
Proteolytic Processing ◽  
Fever Virus ◽  
Morphological Evidence ◽  
Virus Protein

ABSTRACT The first morphological evidence of African swine fever virus (ASFV) assembly is the appearance of precursor viral membranes, thought to derive from the endoplasmic reticulum, within the assembly sites. We have shown previously that protein p54, a viral structural integral membrane protein, is essential for the generation of the viral precursor membranes. In this report, we study the role of protein p17, an abundant transmembrane protein localized at the viral internal envelope, in these processes. Using an inducible virus for this protein, we show that p17 is essential for virus viability and that its repression blocks the proteolytic processing of polyproteins pp220 and pp62. Electron microscopy analyses demonstrate that when the infection occurs under restrictive conditions, viral morphogenesis is blocked at an early stage, immediately posterior to the formation of the viral precursor membranes, indicating that protein p17 is required to allow their progression toward icosahedral particles. Thus, the absence of this protein leads to an accumulation of these precursors and to the delocalization of the major components of the capsid and core shell domains. The study of ultrathin serial sections from cells infected with BA71V or the inducible virus under permissive conditions revealed the presence of large helicoidal structures from which immature particles are produced, suggesting that these helicoidal structures represent a previously undetected viral intermediate.

scholarly journals African Swine Fever Virus Polyprotein pp62 Is Essential for Viral Core Development

2009 ◽  
Vol 84 (1) ◽  
pp. 176-187 ◽  
Author(s):  
Cristina Suárez ◽  
María L. Salas ◽  
Javier M. Rodríguez
Keyword(s):  
Gene Expression ◽  
Viral Particle ◽  
Virus Assembly ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Proteolytic Processing ◽  
Fever Virus ◽  
Clear Correlation ◽  
Late Gene Expression ◽  
The Core

ABSTRACT One of the most characteristic features of African swine fever virus gene expression is its use of two polyproteins, pp220 and pp62, to produce several structural proteins that account for approximately 32% of the total protein virion mass. Equimolecular amounts of these proteins are the major components of the core shell, a thick protein layer that lies beneath the inner envelope, surrounding the viral nucleoid. Polyprotein pp220, which is located immediately underneath the internal envelope, is essential for the encapsidation of the core of the viral particle. In its absence, the infection produces essentially coreless particles. In this study we analyzed, by means of an IPTG (isopropyl-β-d-thiogalactopyranoside)-inducible virus, the role of polyprotein pp62 in virus assembly. Polyprotein pp62 is indispensable for viral replication. The repression of polyprotein pp62 expression does not alter late gene expression or the proteolytic processing of the polyprotein pp220. However, it has a profound impact on the subcellular localization of polyprotein pp220. Electron microscopy studies revealed that polyprotein pp62 is necessary for the correct assembly and maturation of the core of the viral particle. Its repression leads to the appearance of a significant fraction of empty particles, to an increase in the number of immature-like particles, and to the accumulation of defective particles. Immunoelectron microscopy analysis showed a clear correlation between the amount of polyprotein pp62, the quantity of polyprotein pp220, and the state of development of the core, suggesting that the complete absence of polyprotein pp62 during morphogenesis would produce a homogenous population of empty particles.

scholarly journals African swine fever virus A238L inhibitor of NF-κB and of calcineurin phosphatase is imported actively into the nucleus and exported by a CRM1-mediated pathway

2007 ◽  
Vol 88 (2) ◽  
pp. 411-419 ◽  
Author(s):  
Rhiannon N. Silk ◽  
Gavin C. Bowick ◽  
Charles C. Abrams ◽  
Linda K. Dixon
Keyword(s):  
Molecular Mass ◽  
Nuclear Import ◽  
Nuclear Export ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Fever Virus ◽  
Infected Cells ◽  
Calcineurin Phosphatase

This study examined nuclear and cytoplasmic shuttling of the African swine fever virus (ASFV) A238L protein, which is an inhibitor of NF-κB and of calcineurin phosphatase. The results showed that the protein was present in both the nucleus and the cytoplasm in ASFV-infected cells and that the higher molecular mass 32 kDa form of the A238L protein was the predominant nuclear form, which accumulated later in infection. In contrast, both the 28 and 32 kDa forms of the A238L protein were present in the cytoplasm. The A238L protein was actively imported into the nucleus and exported by a CRM1-mediated pathway, although a pool of the protein remained in the cytoplasm and did not enter the nucleus. By using a recombinant ASFV from which the A238L gene had been deleted, it was shown that expression of A238L did not inhibit nuclear import of the NF-κB p50 or p65 subunit and did not inhibit nuclear export of p65 by a CRM1-mediated pathway. The results were consistent with a model in which A238L functions within both the nucleus and the cytoplasm.

scholarly journals African Swine Fever Virus Proteinase Is Essential for Core Maturation and Infectivity

2003 ◽  
Vol 77 (10) ◽  
pp. 5571-5577 ◽  
Author(s):  
Alí Alejo ◽  
Germán Andrés ◽  
María L. Salas
Keyword(s):  
African Swine Fever Virus ◽  
Sequence Similarity ◽  
Cysteine Proteinase ◽  
African Swine Fever ◽  
Virus Production ◽  
Electron Microscopic Examination ◽  
Proteolytic Processing ◽  
Fever Virus ◽  
Core Shell ◽  
Electron Microscopic

ABSTRACT African swine fever virus (ASFV) encodes two polyprotein precursors named pp220 and pp62 that are sequentially processed during viral infection, giving rise to six major structural proteins. These reside at the core shell, a matrix domain located between the endoplasmic reticulum-derived inner envelope and the DNA-containing nucleoid. Proteolytic processing of the polyprotein precursors is catalyzed by the viral proteinase pS273R, a cysteine proteinase that shares sequence similarity with the SUMO1-processing peptidases. We describe here the construction and characterization of an ASFV recombinant, vS273Ri, that inducibly expresses the ASFV proteinase. Using vS273Ri, we show that repression of proteinase expression inhibits polyprotein processing and strongly impairs infective virus production. Electron microscopic examination of vS273Ri-infected cells showed that inhibition of proteolytic processing leads to the assembly of defective icosahedral particles containing a noncentered electron-dense nucleoid surrounded by an abnormal core shell of irregular thickness. The analysis of purified extracellular defective particles revealed that they contain the unprocessed pp220 and pp62 precursors, as well as the major DNA-binding nucleoid proteins p10 and pA104R. Altogether, these results indicate that the proteolytic processing of the polyproteins is not required for their incorporation into the assembling particles nor for the incorporation of the DNA-containing nucleoid. Instead, the ASFV proteinase is involved in a late maturational step that is essential for proper core assembly and infectivity.

Obtaining and Characterisitcs of p17 and p54 Proteins of African Swine Fever Virus in Mammalian Cells

2017 ◽  
pp. 41-47
Author(s):  
K.A. Mima ◽  
◽  
G.S. Burmakina ◽  
I.A. Titov ◽  
M.V. Shkalikova ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Fever Virus

scholarly journals Swift and Reliable “Easy Lab” Methods for the Sensitive Molecular Detection of African Swine Fever Virus

2021 ◽  
Vol 22 (5) ◽  
pp. 2307
Author(s):  
Ahmed Elnagar ◽  
Jutta Pikalo ◽  
Martin Beer ◽  
Sandra Blome ◽  
Bernd Hoffmann
Keyword(s):  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Fever Virus ◽  
Molecular Diagnostic ◽  
Economic Losses ◽  
Hemorrhagic Disease ◽  
Tissue Samples ◽  
Wild Boars ◽  
Field Samples ◽  
Edta Blood

African swine fever (ASF) is a contagious viral hemorrhagic disease of domestic pigs and wild boars. The disease is notifiable to the World Organisation for Animal Health (OIE) and is responsible for high mortality and serious economic losses. PCR and real-time PCR (qPCR) are the OIE-recommended standard methods for the direct detection of African swine fever virus (ASFV) DNA. The aim of our work was the simplification and standardization of the molecular diagnostic workflow in the lab. For validation of this “easy lab” workflow, different sample materials from animal trials were collected and analyzed (EDTA blood, serum, oral swabs, chewing ropes, and tissue samples) to identify the optimal sample material for diagnostics in live animals. Based on our data, the EDTA blood samples or bloody tissue samples represent the best specimens for ASFV detection in the early and late phases of infection. The application of prefilled ready-to-use reagents for nucleic acid extraction or the use of a Tissue Lysis Reagent (TLR) delivers simple and reliable alternatives for the release of the ASFV nucleic acids. For the qPCR detection of ASFV, different published and commercial kits were compared. Here, a lyophilized commercial kit shows the best results mainly based on the increased template input. The good results of the “easy lab” strategy could be confirmed by the ASFV detection in field samples from wild boars collected from the 2020 ASFV outbreak in Germany. Appropriate internal control systems for extraction and PCR are key features of the “easy lab” concept and reduce the risk of false-negative and false-positive results. In addition, the use of easy-to-handle machines and software reduces training efforts and the misinterpretation of results. The PCR diagnostics based on the “easy lab” strategy can realize a high sensitivity and specificity comparable to the standard PCR methods and should be especially usable for labs with limited experiences and resources.

scholarly journals African Swine Fever Virus A528R Inhibits TLR8 Mediated NF-κB Activity by Targeting p65 Activation and Nuclear Translocation

Viruses ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 2046
Author(s):  
Xueliang Liu ◽  
Da Ao ◽  
Sen Jiang ◽  
Nengwen Xia ◽  
Yulin Xu ◽  
...  
Keyword(s):  
Promoter Activity ◽  
Nuclear Translocation ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Viral Proteins ◽  
Fever Virus ◽  
Economic Losses ◽  
Effective Vaccine ◽  
Hemorrhagic Disease ◽  
Host Innate Immunity

African swine fever (ASF) is mainly an acute hemorrhagic disease which is highly contagious and lethal to domestic pigs and wild boars. The global pig industry has suffered significant economic losses due to the lack of an effective vaccine and treatment. The African swine fever virus (ASFV) has a large genome of 170–190 kb, encoding more than 150 proteins. During infection, ASFV evades host innate immunity via multiple viral proteins. A528R is a very important member of the polygene family of ASFV, which was shown to inhibit IFN-β production by targeting NF-κB, but its mechanism is not clear. This study has shown that A528R can suppress the TLR8-NF-κB signaling pathway, including the inhibition of downstream promoter activity, NF-κB p65 phosphorylation and nuclear translocation, and the antiviral and antibacterial activity. Further, we found the cellular co-localization and interaction between A528R and p65, and ANK repeat domains of A528R and RHD of p65 are involved in their interaction and the inhibition of p65 activity. Therefore, we conclude that A528R inhibits TLR8-NF-κB signaling by targeting p65 activation and nuclear translocation.

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