Bovine Herpesvirus 4 Modulates Its beta-1,6-N-Acetylglucosaminyltransferase Activity through Alternative Splicing.
Lété, Céline ; ; Machiels, Bénédicte et al
in Journal of virology (2016), 90(4), 2039-51
Carbohydrates play major roles in host-virus interactions. It is therefore not surprising that, during coevolution with their hosts, viruses have developed sophisticated mechanisms to hijack for their ... [more ▼]
Carbohydrates play major roles in host-virus interactions. It is therefore not surprising that, during coevolution with their hosts, viruses have developed sophisticated mechanisms to hijack for their profit different pathways of glycan synthesis. Thus, the Bo17 gene of Bovine herpesvirus 4 (BoHV-4) encodes a homologue of the cellular core 2 protein beta-1,6-N-acetylglucosaminyltransferase-mucin type (C2GnT-M), which is a key player for the synthesis of complex O-glycans. Surprisingly, we show in this study that, as opposed to what is observed for the cellular enzyme, two different mRNAs are encoded by the Bo17 gene of all available BoHV-4 strains. While the first one corresponds to the entire coding sequence of the Bo17 gene, the second results from the splicing of a 138-bp intron encoding critical residues of the enzyme. Antibodies generated against the Bo17 C terminus showed that the two forms of Bo17 are expressed in BoHV-4 infected cells, but enzymatic assays revealed that the spliced form is not active. In order to reveal the function of these two forms, we then generated recombinant strains expressing only the long or the short form of Bo17. Although we did not highlight replication differences between these strains, glycomic analyses and lectin neutralization assays confirmed that the splicing of the Bo17 gene gives the potential to BoHV-4 to fine-tune the global level of core 2 branching activity in the infected cell. Altogether, these results suggest the existence of new mechanisms to regulate the activity of glycosyltransferases from the Golgi apparatus. IMPORTANCE: Viruses are masters of adaptation that hijack cellular pathways to allow their growth. Glycans play a central role in many biological processes, and several studies have highlighted mechanisms by which viruses can affect glycosylation. Glycan synthesis is a nontemplate process regulated by the availability of key glycosyltransferases. Interestingly, bovine herpesvirus 4 encodes one such enzyme which is a key enzyme for the synthesis of complex O-glycans. In this study, we show that, in contrast to cellular homologues, this virus has evolved to alternatively express two proteins from this gene. While the first one is enzymatically active, the second results from the alternative splicing of the region encoding the catalytic site of the enzyme. We postulate that this regulatory mechanism could allow the virus to modulate the synthesis of some particular glycans for function at the location and/or the moment of infection. [less ▲]Detailed reference viewed: 28 (8 ULg)
Deletion of Murid Herpesvirus 4 ORF63 Affects the Trafficking of Incoming Capsids toward the Nucleus.
Latif, Muhammad Bilal ; Machiels, Bénédicte ; Xiao, Xue et al
in Journal of virology (2016), 90(5), 2455-72
Gammaherpesviruses are important human and animal pathogens. Despite the fact that they display the classical architecture of herpesviruses, the function of most of their structural proteins is still ... [more ▼]
Gammaherpesviruses are important human and animal pathogens. Despite the fact that they display the classical architecture of herpesviruses, the function of most of their structural proteins is still poorly defined. This is especially true for tegument proteins. Interestingly, a potential role in immune evasion has recently been proposed for the tegument protein encoded by Kaposi's sarcoma-associated herpesvirus open reading frame 63 (ORF63). To gain insight about the roles of ORF63 in the life cycle of a gammaherpesvirus, we generated null mutations in the ORF63 gene of murid herpesvirus 4 (MuHV-4). We showed that disruption of ORF63 was associated with a severe MuHV-4 growth deficit both in vitro and in vivo. The latter deficit was mainly associated with a defect of replication in the lung but did not affect the establishment of latency in the spleen. From a functional point of view, inhibition of caspase-1 or the inflammasome did not restore the growth of the ORF63-deficient mutant, suggesting that the observed deficit was not associated with the immune evasion mechanism identified previously. Moreover, this growth deficit was also not associated with a defect in virion egress from the infected cells. In contrast, it appeared that MuHV-4 ORF63-deficient mutants failed to address most of their capsids to the nucleus during entry into the host cell, suggesting that ORF63 plays a role in capsid movement. In the future, ORF63 could therefore be considered a target to block gammaherpesvirus infection at a very early stage of the infection. IMPORTANCE: The important diseases caused by gammaherpesviruses in human and animal populations justify a better understanding of their life cycle. In particular, the role of most of their tegument proteins is still largely unknown. In this study, we used murid herpesvirus 4, a gammaherpesvirus infecting mice, to decipher the role of the protein encoded by the viral ORF63 gene. We showed that the absence of this protein is associated with a severe growth deficit both in vitro and in vivo that was mainly due to impaired migration of viral capsids toward the nucleus during entry. Together, our results provide new insights about the life cycle of gammaherpesviruses and could allow the development of new antiviral strategies aimed at blocking gammaherpesvirus infection at the very early stages. [less ▲]Detailed reference viewed: 22 (6 ULg)
An endogenous gibbon ape leukemia virus (GALV) identified in a rodent (Melomys sp.) from Indonesia
; Michaux, Johan ; et al
in Journal of Virology (2016), sous presse
ABSTRACT Gibbon ape leukemia virus (GALV) and koala retrovirus (KoRV) most likely originated from a cross-species transmission of an ancestral retrovirus into koalas and gibbons via one or more ... [more ▼]
ABSTRACT Gibbon ape leukemia virus (GALV) and koala retrovirus (KoRV) most likely originated from a cross-species transmission of an ancestral retrovirus into koalas and gibbons via one or more intermediate as yet unknown hosts. A highly similar virus to GALV has been identified in an Australian rodent (Melomys burtoni) after extensive screening of Australian wildlife. GALV-like viruses have also been discovered in several Southeast Asian species although screening has not been extensive and viruses discovered to date are only distantly related to GALV. We therefore screened 26 Southeast Asian rodent species for KoRV- and GALV-like sequences, using hybridization capture and high-throughput sequencing, in the attempt to identify potential GALV and KoRV hosts. Only one species, an undescribed species of Melomys from Indonesia, was positive yielding an endogenous provirus very closely related to a strain of GALV. The sequence of the critical receptor domain for GALV infection in the Indonesian Melomys sp. was consistent with the susceptibility of the species to GALV infection. The discovery of a GALV in a second Melomys species provides further evidence that Melomys may play a role in the spread of GALV-like viruses, especially since the genus is found in Indonesia, Papua New Guinea and Australia, connecting the home ranges of koalas and gibbons. IMPORTANCE The gibbon ape leukemia virus (GALV) and the koala retrovirus (KoRV) are very closely related, yet their hosts are neither closely related nor overlap geographically. Direct cross-species infection between koalas and gibbons is unlikely. Therfore, GALV and KoRV may have arisen via a cross-species transfer from an intermediate host that overlaps in range with both gibbons and koalas. Using hybridization capture and high-throughput sequencing, we have screened a wide range of rodent candidate hosts from Southeast Asia for KoRV- and GALV-like sequences. Only a Melomys species from Indonesia was positive for GALV. We report the genome sequence of this newly identified GALV, the critical domain for infection of its potential cellular receptor and its phylogenetic relationships with the other previously characterized GALVs. We hypothesize that the genus Melomys may have played a key role in cross-species transmission to other taxa. [less ▲]Detailed reference viewed: 60 (4 ULg)
The genome of a tortoise herpesvirus (testudinid herpesvirus 3) has a novel structure and contains a large region that is not required for replication in vitro or virulence in vivo.
Gandar, Frederic ; ; et al
in Journal of Virology (2015), 89(22), 11438-11456
Testudinid herpesvirus 3 (TeHV-3) is the causative agent of a lethal disease affecting several tortoise species. The threat that this virus poses to endangered animals is focusing efforts on ... [more ▼]
Testudinid herpesvirus 3 (TeHV-3) is the causative agent of a lethal disease affecting several tortoise species. The threat that this virus poses to endangered animals is focusing efforts on characterizing its properties, in order to enable the development of prophylactic methods. We have sequenced the genomes of the two most studied TeHV-3 strains (1976 and 4295). TeHV-3 strain 1976 has a novel genome structure and is most closely related to a turtle herpesvirus, thus supporting its classification into genus Scutavirus, subfamily Alphaherpesvirinae, family Herpesviridae. The sequence of strain 1976 also revealed viral counterparts of cellular interleukin-10 and semaphorin, which have not been described previously in members of subfamily Alphaherpesvirinae. TeHV-3 strain 4295 is a mixture of three forms (m1, m2, and M), in which, in comparison to strain 1976, the genomes exhibit large, partially overlapping deletions of 12.5 to 22.4 kb. Viral subclones representing these forms were isolated by limiting dilution assays, and each replicated in cell culture comparably to strain 1976. With the goal of testing the potential of the three forms as attenuated vaccine candidates, strain 4295 was inoculated intranasally into Hermann's tortoises (Testudo hermanni). All inoculated subjects died, and PCR analyses demonstrated the ability of the m2 and M forms to spread and invade the brain. In contrast, the m1 form was detected in none of the organs tested, suggesting its potential as the basis of an attenuated vaccine candidate. Our findings represent a major step toward characterizing TeHV-3 and developing prophylactic methods against it. [less ▲]Detailed reference viewed: 53 (9 ULg)
Viral semaphorin inhibits dendritic cell phagocytosis and migration but is not essential for γ-herpesvirus-induced lymphoproliferation in malignant catarrhal fever.
Myster, Françoise ; Gonon Rodrigues Palmeira, Leonor ; Sorel, Océane et al
in Journal of Virology (2015)Detailed reference viewed: 26 (10 ULg)
Mutation of a Single Envelope N-linked Glycosylation Site Enhances the Pathogenicity of Bovine Leukemia Virus
De Brogniez, Alix ; ; Jacques, Jean-Rock et al
in Journal of Virology (2015), 89(17),
Viruses have co-evolved with their host to ensure efficient replication and transmission without inducing excessive pathogenicity that would indirectly impair their persistence. This is exemplified by the ... [more ▼]
Viruses have co-evolved with their host to ensure efficient replication and transmission without inducing excessive pathogenicity that would indirectly impair their persistence. This is exemplified by the bovine leukemia virus (BLV) system in which lymphoproliferative disorders develop in ruminants after latency periods of several years. In principle, the equilibrium reached between the virus and its host could be disrupted by emergence of more pathogenic strains. Intriguingly but fortunately, such a hyperpathogenic BLV strain was never observed in the field nor designed in vitro. In this study, we aimed at understanding the role of envelope N-linked glycosylation with the hypothesis that this posttranslational modification could either favor BLV infection by allowing viral entry or allow immune escape by using glycans as a shield. Using reverse genetics of an infectious molecular provirus, we have identified a N-linked envelope glycosylation site (N230) that limits viral replication and pathogenicity. Indeed, mutation N230E unexpectedly leads to enhanced fusogenicity and protein stability. Occurrence of this mutation may thus represent a potential threat associated with emergence of hyperpathogenic BLV strains and possibly of new variants of the related primate T-lymphotropic viruses. [less ▲]Detailed reference viewed: 29 (14 ULg)
Discovery and characterization of auxiliary proteins encoded by Simian T-cell Lymphotropic Viruses type 3
; ; et al
in Journal of Virology (2014)Detailed reference viewed: 50 (21 ULg)
Deletion of the ORF9p acidic cluster impairs the nuclear egress of Varicella-zoster virus capsids.
Riva, Laura ; Thiry, Marc ; Lebrun, Marielle et al
in Journal of virology (2014)
The protein encoded by the ORF9 is essential for Varicella-zoster virus (VZV) replication. Previous studies documented its presence in the trans-Golgi network and its involvement in secondary envelopment ... [more ▼]
The protein encoded by the ORF9 is essential for Varicella-zoster virus (VZV) replication. Previous studies documented its presence in the trans-Golgi network and its involvement in secondary envelopment. In this work, we deleted the ORF9p acidic cluster, destroying its interaction with ORF47p and resulting in a nuclear accumulation of both proteins. This phenotype results to an accumulation of primary enveloped capsids in the perinuclear space, reflecting a capsid de-envelopment defect. [less ▲]Detailed reference viewed: 41 (4 ULg)
Tetherin Restricts Herpes Simplex Virus 1 and Is Antagonized by Glycoprotein M
Blondeau, Caroline ; ; et al
in Journal of Virology (2013), 87(24),
Tetherin is a broadly active antiviral effector that works by tethering nascent enveloped virions to a host cell membrane, thus preventing their release. In this study, we demonstrate that herpes simplex ... [more ▼]
Tetherin is a broadly active antiviral effector that works by tethering nascent enveloped virions to a host cell membrane, thus preventing their release. In this study, we demonstrate that herpes simplex virus 1 (HSV-1) is targeted by tetherin. We identify the viral envelope glycoprotein M (gM) as having moderate anti-tetherin activity. We show that gM but not gB or gD efficiently removes tetherin from the plasma membrane and can functionally substitute for the human immunodeficiency virus type 1 (HIV-1) Vpu protein, the prototypic viral tetherin antagonist, in rescuing HIV-1 release from tetherin-expressing cells. Our data emphasize that tetherin is a broadly active antiviral effector and contribute to the emerging hypothesis that viruses must suppress or evade an array of host cell countermeasures in order to establish a productive infection. [less ▲]Detailed reference viewed: 19 (1 ULg)
ORF9p phosphorylation by ORF47p is crucial for the formation and egress of the Varicella-zoster virus (VZV) viral particles.
Riva, Laura ; Thiry, Marc ; BONTEMS, Sébastien et al
in Journal of Virology (2013), 87(5), 2868-2881
The role of the tegument during the herpesvirus lytic cycle is still not clearly established, particularly at the late phase of infection, when the newly produced viral particles need to be fully ... [more ▼]
The role of the tegument during the herpesvirus lytic cycle is still not clearly established, particularly at the late phase of infection, when the newly produced viral particles need to be fully assembled before being released from the infected cell. The Varicella-zoster virus (VZV) protein coded by ORF9 (ORF9p) is an essential tegument protein and, even though its mRNA is the most expressed during the productive infection, little is known about its functions. Using a GalK positive/negative selection technique, we modified a BAC containing the complete VZV genome creating viruses expressing mutant versions of ORF9p.We showed that ORF9p is hyper-phosphorylated during the infection, especially through its interaction with the viral Ser/Thr kinase ORF47p; we identified a consensus site within ORF9p recognized by ORF47p and demonstrated its importance for ORF9p phosphorylation. Strikingly, an ultra-structural analysis revealed that the mutation of this consensus site (Glutamate 85 to Arginine) strongly affects viral assembly and release, reproducing ORF47 kinase dead VZV phenotype. It also slightly diminishes the infectivity towards immature dendritic cells. Taken together, our results identify ORF9p as a new viral substrate of ORF47p and suggest a determinant role of this phosphorylation for viral infectivity, especially during the process of viral particle formation and egress. [less ▲]Detailed reference viewed: 56 (30 ULg)
Glycoprotein B cleavage is important for murid herpesvirus 4 to infect myeloid cells.
; ; et al
in Journal of Virology (2013)
Glycoprotein B (gB) is a conserved herpesvirus virion component implicated in membrane fusion. As with many - but not all - herpesviruses, the gB of murid herpesvirus 4 (MuHV-4) is cleaved into disulfide ... [more ▼]
Glycoprotein B (gB) is a conserved herpesvirus virion component implicated in membrane fusion. As with many - but not all - herpesviruses, the gB of murid herpesvirus 4 (MuHV-4) is cleaved into disulfide-linked subunits, apparently by furin. Preventing gB cleavage for some herpesviruses causes minor infection deficits in vitro, but what the cleavage contributes to host colonization has been unclear. To address this we mutated the furin cleavage site (R-R-K-R) of the MuHV-4 gB. Abolishing gB cleavage did not affect its expression levels, glycosylation or antigenic conformation. In vitro, mutant viruses entered fibroblasts and epithelial cells normally, but had a significant entry deficit in myeloid cells such as macrophages and bone marrow-derived dendritic cells. The deficit in myeloid cells was not due to reduced virion binding or endocytosis, suggesting that gB cleavage promotes infection at a post-endocytic entry step, presumably viral membrane fusion. In vivo, viruses lacking gB cleavage showed reduced lytic spread in the lungs. Alveolar epithelial cell infection was normal, but alveolar macrophage infection was significantly reduced. Normal long-term latency in lymphoid tissue was established nonetheless. [less ▲]Detailed reference viewed: 31 (4 ULg)
Viral expression directs the fate of B cells in BLV-infected sheep.
Florins, Arnaud-Francois ; De Brogniez, Alix ; et al
in Journal of Virology (2012), 86(1), 621-624
The host immune response is believed to tightly control viral replication of deltaretroviruses such as human T-lymphotropic virus type 1 (HTLV-1) and bovine leukemia virus (BLV). However, this assumption ... [more ▼]
The host immune response is believed to tightly control viral replication of deltaretroviruses such as human T-lymphotropic virus type 1 (HTLV-1) and bovine leukemia virus (BLV). However, this assumption has not been definitely proven in vivo. In order to further evaluate the importance of immune response in the BLV model, we studied the fate of cells in which viral expression was transiently induced. Using a dual fluorochrome labeling approach, we show that ex vivo induction of viral expression induces higher death rates of B cells in vivo. Furthermore, cyclosporine treatment of these animals indicated that an efficient immune response is required to control virus expressing cells. [less ▲]Detailed reference viewed: 39 (9 ULg)
Proteomic characterization of bovine herpesvirus 4 extracellular virions.
Lété, Céline ; Palmeira, Leonor ; et al
in Journal of Virology (2012), 86(21), 11567-80
Gammaherpesviruses are important pathogens in human and animal populations. During early events of infection, these viruses manipulate preexisting host cell signaling pathways to allow successful ... [more ▼]
Gammaherpesviruses are important pathogens in human and animal populations. During early events of infection, these viruses manipulate preexisting host cell signaling pathways to allow successful infection. The different proteins that compose viral particles are therefore likely to have critical functions not only in viral structures and in entry into target cell but also in evasion of the host's antiviral response. In this study, we analyzed the protein composition of bovine herpesvirus 4 (BoHV-4), a close relative of the human Kaposi's sarcoma-associated herpesvirus. Using mass spectrometry-based approaches, we identified 37 viral proteins associated with extracellular virions, among which 24 were resistant to proteinase K treatment of intact virions. Analysis of proteins associated with purified capsid-tegument preparations allowed us to define protein localization. In parallel, in order to identify some previously undefined open reading frames, we mapped peptides detected in whole virion lysates onto the six frames of the BoHV-4 genome to generate a proteogenomic map of BoHV-4 virions. Furthermore, we detected important glycosylation of three envelope proteins: gB, gH, and gp180. Finally, we identified 38 host proteins associated with BoHV-4 virions; 15 of these proteins were resistant to proteinase K treatment of intact virions. Many of these have important functions in different cellular pathways involved in virus infection. This study extends our knowledge of gammaherpesvirus virions composition and provides new insights for understanding the life cycle of these viruses. [less ▲]Detailed reference viewed: 54 (9 ULg)
Persistent infection of thymic epithelial cells with coxsackievirus B4 results in a decreased expression of insulin-like growth factor 2
; ; et al
in Journal of Virology (2012), 86
It has been hypothesized that a disturbance of central self-tolerance to islet β-cell may play a role in the enteroviral pathogenesis of type 1 diabetes. Whether enteroviruses can induce an impaired ... [more ▼]
It has been hypothesized that a disturbance of central self-tolerance to islet β-cell may play a role in the enteroviral pathogenesis of type 1 diabetes. Whether enteroviruses can induce an impaired expression of β-cell self-antigens in thymic epithelial cells has been investigated in a murine thymic epithelial (MTE) cell line. This cell line was permissive to the diabetogenic strain CV-B4 E2 and spontaneously expressed type 2 insulin-like growth factor (Igf2), the dominant self-antigen of the insulin family. In this model, a persistent replication of CV-B4 E2 was obtained as attested by the prolonged detection of intracellular positive and negative-strand viral RNA by RT-PCR, and capsid protein VP1 by IF and by the release of infectious particles in culture supernatant fluids. The chronic stage of the infection was characterized by a low proportion of VP1-positive cells (1-2%) whereas many cells harbored enteroviral RNA as displayed by RT-PCR without extraction applied directly on a few cells. Igf2 mRNA and IGF-2 protein were dramatically decreased in CV-B4 E2-infected MTE cultures compared with mock-infected cultures, whereas housekeeping and Il6 genes expression were maintained and Igf1 mRNA was decreased but at a lower extent. Inoculation of CV-B3-, CV-B4 JVB- or Echovirus 1 resulted in a low level of IGF-2 in culture supernatant fluids as well, whereas HSV-1 stimulated the production of the protein. Thus, a persistent infection of a thymic epithelial cell line with enteroviruses, like CV-B4 E2 can result in a disturbed production of IGF-2, a protein involved in central self-tolerance towards islet β-cells. [less ▲]Detailed reference viewed: 60 (11 ULg)
Bovine Herpesvirus Type 4 Glycoprotein L Is Nonessential for Infectivity but Triggers Virion Endocytosis during Entry
Lété, Céline ; Machiels, Bénédicte ; et al
in Journal of Virology (2012)
The core entry machinery of mammalian herpesviruses comprises glycoproteins B, H and L (gB, gH and gL). gH and gL form a heterodimer with a central role in viral membrane fusion. When archetypal alpha- or ... [more ▼]
The core entry machinery of mammalian herpesviruses comprises glycoproteins B, H and L (gB, gH and gL). gH and gL form a heterodimer with a central role in viral membrane fusion. When archetypal alpha- or beta-herpesviruses lack gL, gH misfolds and progeny virions are non-infectious. However, the gL of the rhadinovirus Murid herpesvirus 4 (MuHV-4) is non-essential for infection. In order to define more generally what role gL plays in rhadinovirus infections, we disrupted its coding sequence in Bovine herpesvirus-4 (BoHV-4). BoHV-4 lacking gL showed altered gH glycosylation and incorporated somewhat less gH into virions but remained infectious. However, gL- virions showed poor growth associated with an entry deficit. Moreover a major part of their entry defect appeared to reflect impaired endocytosis, which occurs upstream of membrane fusion itself. Thus, the rhadinovirus gL may be more important for driving virion endocytosis than for incorporating gH into virions, and is non-essential for membrane fusion. [less ▲]Detailed reference viewed: 95 (21 ULg)
Complete genome sequence of a novel bovine norovirus: Evidence for slow genetic evolution in genogroup III genotype 2 noroviruses
Mauroy, Axel ; ; et al
in Journal of Virology (2012), 86(22), 12449-12450
A new genogroup III genotype 2 bovine norovirus, B309/2003/BE, was entirely sequenced and genetically compared to the original Newbury2/1976/UK strain and to Dumfries/1994/UK, detected in 1976 and 1994 ... [more ▼]
A new genogroup III genotype 2 bovine norovirus, B309/2003/BE, was entirely sequenced and genetically compared to the original Newbury2/1976/UK strain and to Dumfries/1994/UK, detected in 1976 and 1994, respectively. Interestingly, except in welldefined coding regions (N-terminal protein, 3A-like protease, hypervariable region of the capsid protein, and C-terminal part of the minor structural protein), very low genetic differences were noted between the entire genomes of these three strains along a 30-year-long period. It allowed some hypotheses of hotspots of genetic evolution through a low genetic evolution background in genotype 2 genogroup III bovine noroviruses. © 2012, American Society for Microbiology. [less ▲]Detailed reference viewed: 40 (8 ULg)
Ex vivo bioluminescent detection of Alcelaphine herpesvirus 1 infection during malignant catarrhal fever.
Dewals, Benjamin G ; Myster, Françoise ; Palmeira, Leonor et al
in Journal of Virology (2011), 85(14), 6941-54
Alcelaphine herpesvirus 1 (AlHV-1), carried by wildebeest asymptomatically, causes malignant catarrhal fever (WD-MCF) when cross-species transmitted to a variety of susceptible species of the Artiodactyla ... [more ▼]
Alcelaphine herpesvirus 1 (AlHV-1), carried by wildebeest asymptomatically, causes malignant catarrhal fever (WD-MCF) when cross-species transmitted to a variety of susceptible species of the Artiodactyla order. Experimentally, WD-MCF can be reproduced in rabbits. WD-MCF is described as a combination of lymphoproliferation and degenerative lesions in virtually all organs and caused by unknown mechanisms. Recently, we demonstrated that WD-MCF is associated with the proliferation of CD8(+) cells supporting a latent type of infection in lymphoid tissues. Here, we investigated the macroscopic distribution of AlHV-1 infection using ex vivo bioluminescence imaging in rabbit to determine whether it correlates with the distribution of lesions in lymphoid and non-lymphoid organs. To reach that goal, a recombinant AlHV-1 strain was produced by insertion of a luciferase expression cassette (luc) in an intergenic region. In vitro, the reconstituted AlHV-1 luc(+) strain replicated comparably to the parental strain and luciferase activity was detected by bioluminescence imaging. In vivo, rabbits infected with the AlHV-1 luc(+) strain developed WD-MCF comparably to the parental wild-type strain with hyperthermia and increase of both CD8(+) T cells frequencies and viral genomic charge over time in peripheral blood mononuclear cells and in lymph nodes at time of euthanasia. Bioluminescent imaging revealed that AlHV-1 infection could be detected ex vivo in lymphoid organs but also in lung, liver and kidney during WD-MCF, demonstrating that AlHV-1 infection is prevalent in tissue lesions. Finally, we show that the infiltrating mononuclear leukocytes in non-lymphoid organs are mainly CD8(+) T cells and that latency is predominant during WD-MCF. [less ▲]Detailed reference viewed: 90 (32 ULg)
Bovine Herpesvirus 4 Bo10 gene encodes a nonessential viral envelope protein that regulates viral tropism through both positive and negative effects.
Machiels, Bénédicte ; Lété, Céline ; et al
in Journal of Virology (2011), 85(2), 1011-1024
All gammaherpesviruses encode a glycoprotein positionally homologous to the Epstein-Barr virus gp350 and the Kaposi's Sarcoma associated herpesvirus (KSHV) K8.1. In this study, we characterized that of ... [more ▼]
All gammaherpesviruses encode a glycoprotein positionally homologous to the Epstein-Barr virus gp350 and the Kaposi's Sarcoma associated herpesvirus (KSHV) K8.1. In this study, we characterized that of Bovine Herpesvirus-4 (BoHV-4), encoded by the Bo10 gene. We identified a 180 kDa gene product, gp180, which was incorporated into the virion envelope. A Bo10 deletion virus was viable, but showed a growth deficit associated with reduced binding to epithelial cells. This seemed to reflect an interaction of gp180 with glycosaminoglycans (GAGs), since the Bo10 mutant was both less infectious for GAG(+) cells than the wild-type and more infectious for GAG(-) cells. However, we could not identify a direct interaction between gp180 and GAGs, implying that any direct interaction must be of low affinity. This function of gp180 was very similar to that previously identified for the Murid Herpesvirus 4 gp150, and also to the Epstein-Barr virus gp350 that promotes CD21(+) cell infection and inhibits CD21(-) cell infection. We propose that such proteins generally regulate virion attachment both by binding to cells and by covering another receptor-binding protein until they are displaced. Thus they regulate viral tropism both positively and negatively depending upon the presence or absence of their receptor. [less ▲]Detailed reference viewed: 114 (48 ULg)
Co-infection with two closely related alphaherpesviruses results in a highly diversified recombination mosaic displaying negative genetic interference
Muylkens, Benoît ; Farnir, Frédéric ; et al
in Journal of Virology (2009), 83(7), 3127-3137
Phylogenetic studies of the emergence and spread of natural recombinants in herpesviruses infecting humans and animals have been reported recently. However, despite an ever-increasing amount of evidence ... [more ▼]
Phylogenetic studies of the emergence and spread of natural recombinants in herpesviruses infecting humans and animals have been reported recently. However, despite an ever-increasing amount of evidence of recombination in herpesvirus history, the recombination process and the consequences on the genetic diversity of the progeny remain poorly characterized. We addressed this issue by using multiple single-nucleotide polymorphisms (SNPs) differentiating the two subtypes of an alphaherpesvirus, bovine herpesvirus 1 (BoHV-1). Analysis of a large sample of progeny virions obtained in a single growth cycle of coinfected BoHV-1 strains provided a prospective investigation of the recombination dynamics by using SNPs as recombination markers. We found that the simultaneous infection with two closely related herpesviruses results in a highly diversified recombination mosaic. From the analysis of multiple recombinants arising in the progeny, we provide the first evidence of genetic interference influencing the recombination process in herpesviruses. In addition, we report striking differences in the levels of recombination frequency observed along the BoHV-1 genome. With particular emphasis on the genetic structure of a progeny virus population rising in vitro, our data show to which extent recombination participates to the genetic diversification of herpesviruses [less ▲]Detailed reference viewed: 85 (28 ULg)
The major portal of entry of koi herpesvirus in cyprinus carpio is the skin.
Costes, Bérénice ; ; Michel, Benjamin et al
in Journal of Virology (2009)
Koi herpesvirus (KHV), recently designated in the species Cyprinid Herpesvirus 3, is the causative agent of a lethal disease in koi and common carp. In the present study, we investigated the portal of ... [more ▼]
Koi herpesvirus (KHV), recently designated in the species Cyprinid Herpesvirus 3, is the causative agent of a lethal disease in koi and common carp. In the present study, we investigated the portal of entry of KHV in carp using bioluminescence imaging. Taking profit of the recent cloning of the KHV genome as a bacterial artificial chromosome (BAC), we produced a recombinant plasmid encoding a firefly luciferase (LUC) expression cassette inserted in the intergenic region between ORF 136 and ORF 137. Two viral strains were then reconstituted from the modified plasmid: the FL BAC 136 LUC excised strain and the FL BAC 136 LUC TK revertant strain encoding a disrupted and a wild-type thymidine kinase (TK) locus, respectively. In vitro, the two recombinant strains replicated comparably to the parental FL strain. The FL BAC 136 LUC TK revertant strain was shown in vitro to induce a bioluminescent signal allowing the detection of single positive cells as early as 24 hours post-infection; while in vivo, it induced KHV infection in carp that was indistinguishable from that induced by the parental FL strain. To identify the KHV portal of entry, carp were analyzed by bioluminescence imaging at different time post-infection with the FL BAC 136 LUC TK revertant strain. These analyses demonstrated that the skin of the fish, covering the fins and also the body, is the major portal of entry of KHV in carp. Finally, to further demonstrate the role of the skin as the KHV portal of entry, we constructed an original system nicknamed "U-tube" to perform per-cutaneous infection restricted to the posterior part of the fish. All the data obtained in the present study demonstrate that the skin and not the gills is the major portal of entry of KHV in carp. [less ▲]Detailed reference viewed: 86 (15 ULg)