scholarly journals Activation of Vav by the Gammaherpesvirus M2 Protein Contributes to the Establishment of Viral Latency in B Lymphocytes

2006 ◽  
Vol 80 (12) ◽  
pp. 6123-6135 ◽  
Author(s):  
Lénia Rodrigues ◽  
Marta Pires de Miranda ◽  
María J. Caloca ◽  
Xosé R. Bustelo ◽  
J. Pedro Simas
Keyword(s):  
B Lymphocytes ◽  
Viral Latency ◽  
Murine Gammaherpesvirus ◽  
Host Interaction ◽  
New Type ◽  
Trimolecular Complex ◽  
Gammaherpesvirus 68 ◽  
Novel Strategy ◽  
Murine Gammaherpesvirus 68

ABSTRACT Gammaherpesviruses subvert eukaryotic signaling pathways to favor latent infections in their cellular reservoirs. To this end, they express proteins that regulate or replace functionally specific signaling proteins of eukaryotic cells. Here we describe a new type of such viral-host interaction that is established through M2, a protein encoded by murine gammaherpesvirus 68. M2 associates with Vav proteins, a family of phosphorylation-dependent Rho/Rac exchange factors that play critical roles in lymphocyte signaling. M2 expression leads to Vav1 hyperphosphorylation and to the subsequent stimulation of its exchange activity towards Rac1, a process mediated by the formation of a trimolecular complex with Src kinases. This heteromolecular complex is coordinated by proline-rich and Src family-dependent phosphorylated regions of M2. Infection of Vav-deficient mice with gammaherpesvirus 68 results in increased long-term levels of latency in germinal center B lymphocytes, corroborating the importance of the M2/Vav cross talk in the process of viral latency. These results reveal a novel strategy used by the murine gammaherpesvirus family to subvert the lymphocyte signaling machinery to its own benefit.

scholarly journals Dendritic Cells Loaded with Tumor B Cells Elicit Broad Immunity against Murine Gammaherpesvirus 68 but Fail To Prevent Long-Term Latency

2010 ◽  
Vol 84 (17) ◽  
pp. 8975-8979
Author(s):  
Janet Weslow-Schmidt ◽  
Fang Ye ◽  
Stephanie S. Cush ◽  
Kathleen A. Stuller ◽  
Marcia A. Blackman ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
B Cells ◽  
Acute Infection ◽  
Dendritic Cell Vaccination ◽  
Murine Gammaherpesvirus ◽  
Attenuated Viruses ◽  
Gammaherpesvirus 68 ◽  
Vaccine Approach ◽  
Murine Gammaherpesvirus 68

ABSTRACT It is still unknown whether a noninfectious gammaherpesvirus vaccine is able to prevent or reduce virus persistence. This led us to use dendritic cells loaded with tumor B cells as a vaccine approach for the murine gammaherpesvirus 68 (γHV68) model of infection. Dendritic cells loaded with UV-irradiated latently infected tumor B cells induce broad, strong, and long-lasting immunity against γHV68. Dendritic cell vaccination prevents the enlargement of lymph nodes and severely limits acute infection and early latency but does not prevent γHV68 from establishing long-term latency. Our findings support the concept that attenuated viruses may be the best vaccine option for preventing gammaherpesvirus persistence.

scholarly journals Identification of Infected B-Cell Populations by Using a Recombinant Murine Gammaherpesvirus 68 Expressing a Fluorescent Protein

2009 ◽  
Vol 83 (13) ◽  
pp. 6484-6493 ◽  
Author(s):  
Christopher M. Collins ◽  
Jeremy M. Boss ◽  
Samuel H. Speck
Keyword(s):  
B Cells ◽  
Virus Infection ◽  
Plasma Cells ◽  
Fluorescent Protein ◽  
Viral Latency ◽  
Murine Gammaherpesvirus ◽  
Latently Infected ◽  
Infected Cells ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68

ABSTRACT Infection of inbred mice with murine gammaherpesvirus 68 (MHV68) has proven to be a powerful tool to study gammaherpesvirus pathogenesis. However, one of the limitations of this system has been the inability to directly detect infected cells harvested from infected animals. To address this issue, we generated a transgenic virus that expresses the enhanced yellow fluorescent protein (YFP), driven by the human cytomegalovirus immediate-early promoter and enhancer, from a neutral locus within the viral genome. This virus, MHV68-YFP, replicated and established latency as efficiently as did the wild-type virus. During the early phase of viral latency, MHV68-YFP efficiently marked latently infected cells in the spleen after intranasal inoculation. Staining splenocytes for expression of various surface markers demonstrated the presence of MHV68 in distinct populations of splenic B cells harboring MHV68. Notably, these analyses also revealed that markers used to discriminate between newly formed, follicular and marginal zone B cells may not be reliable for phenotyping B cells harboring MHV68 since virus infection appears to modulate cell surface expression levels of CD21 and CD23. However, as expected, we observed that the overwhelming majority of latently infected B cells at the peak of latency exhibited a germinal center phenotype. These analyses also demonstrated that a significant percentage of MHV68-infected splenocytes at the peak of viral latency are plasma cells (ca. 15% at day 14 and ca. 8% at day 18). Notably, the frequency of virus-infected plasma cells correlated well with the frequency of splenocytes that spontaneously reactivate virus upon explant. Finally, we observed that the efficiency of marking latently infected B cells with the MHV68-YFP recombinant virus declined at later times postinfection, likely due to shut down of transgene expression, and indicating that the utility of this marking strategy is currently limited to the early stages of virus infection.

scholarly journals Analysis of a Novel Strain of Murine Gammaherpesvirus Reveals a Genomic Locus Important for Acute Pathogenesis

2001 ◽  
Vol 75 (11) ◽  
pp. 5315-5327 ◽  
Author(s):  
Alastair I. Macrae ◽  
Bernadette M. Dutia ◽  
Steven Milligan ◽  
David G. Brownstein ◽  
Deborah J. Allen ◽  
...  
Keyword(s):  
Small Animal ◽  
Genomic Locus ◽  
Unique Region ◽  
Murine Gammaherpesvirus ◽  
Growth Properties ◽  
Infected Cells ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68 ◽  
Yellow Necked Mouse

ABSTRACT Infection of mice by murine gammaherpesvirus 68 (MHV-68) is an excellent small-animal model of gammaherpesvirus pathogenesis in a natural host. We have carried out comparative studies of another herpesvirus, murine herpesvirus 76 (MHV-76), which was isolated at the same time as MHV-68 but from a different murid host, the yellow-necked mouse (Apodemus flavicollis). Molecular analyses revealed that the MHV-76 genome is essentially identical to that of MHV-68, except for deletion of 9,538 bp at the left end of the unique region. MHV-76 is therefore a deletion mutant that lacks four genes unique to MHV-68 (M1, M2,M3, and M4) as well as the eight viral tRNA-like genes. Replication of MHV-76 in cell culture was identical to that of MHV-68. However, following infection of mice, MHV-76 was cleared more rapidly from the lungs. In line with this, there was an increased inflammatory response in lungs with MHV-76. Splenomegaly was also significantly reduced following MHV-76 infection, and much less latent MHV-76 was detected in the spleen. Nevertheless, MHV-76 maintained long-term latency in the lungs and spleen. We utilized a cosmid containing the left end of the MHV-68 genome to reinsert the deleted sequence into MHV-76 by recombination in infected cells, and we isolated a rescuant virus designated MHV-76(cA8+)4 which was ostensibly genetically identical to MHV-68. The growth properties of the rescuant in infected mice were identical to those of MHV-68. These results demonstrate that genetic elements at the left end of the unique region of the MHV-68 genome play vital roles in host evasion and are critical to the development of splenic pathology.

scholarly journals Analysis of Virus-Specific CD4+ T Cells during Long-Term Gammaherpesvirus Infection

2001 ◽  
Vol 75 (16) ◽  
pp. 7744-7748 ◽  
Author(s):  
Emilio Flaño ◽  
David L. Woodland ◽  
Marcia A. Blackman ◽  
Peter C. Doherty
Keyword(s):  
T Cells ◽  
Major Histocompatibility Complex ◽  
Complex Class ◽  
Mediastinal Lymph Nodes ◽  
Total Response ◽  
Murine Gammaherpesvirus ◽  
Histocompatibility Complex ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68

ABSTRACT Major histocompatibility complex class II-mediated antigen presentation after intranasal infection with murine gammaherpesvirus 68 differs in mediastinal lymph nodes and spleen. Evidence that virus-specific CD4+ T cells were being stimulated was found as late as 6 to 8 months after infection, and cells specific for the viral gp15067–83 and ORF11168–180 peptides were maintained as a fairly stable proportion of the total response.

scholarly journals Effective Vaccination against Long-Term Gammaherpesvirus Latency

2003 ◽  
Vol 77 (4) ◽  
pp. 2522-2529 ◽  
Author(s):  
Scott A. Tibbetts ◽  
J. Scott McClellan ◽  
Shivaprakash Gangappa ◽  
Samuel H. Speck ◽  
Herbert W. Virgin
Keyword(s):  
Latent Infection ◽  
Limit Of Detection ◽  
Challenge Virus ◽  
Murine Gammaherpesvirus ◽  
Relevant Situation ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68 ◽  
Memory Immune Response ◽  
Sterilizing Immunity

ABSTRACT The fundamental question of whether a primed immune system is capable of preventing latent gammaherpesvirus infection remains unanswered. Recent studies showing that vaccination can reduce acute replication and short-term latency but cannot alter long-term latency further call into question the possibility of achieving sterilizing immunity against gammaherpesviruses. Using the murine gammaherpesvirus 68 (γHV68) system, we demonstrate that it is possible to effectively vaccinate against long-term latency. By immunizing mice with a γHV68 mutant virus that is deficient in its ability to reactivate from latency, we reduced latent infection of wild-type challenge virus to a level below the limit of detection. Establishment of latency was inhibited by vaccination regardless of whether mice were challenged intraperitoneally or intranasally. Passive transfer of antibody from vaccinated mice could partially reconstitute the effect, demonstrating that antibody is an important component of vaccination. These results demonstrate the potential of a memory immune response against gammaherpesviruses to alter long-term latency and suggest that limiting long-term latent infection in a clinically relevant situation is an attainable goal.

scholarly journals Cloning and Mutagenesis of the Murine Gammaherpesvirus 68 Genome as an Infectious Bacterial Artificial Chromosome

2000 ◽  
Vol 74 (15) ◽  
pp. 6964-6974 ◽  
Author(s):  
Heiko Adler ◽  
Martin Messerle ◽  
Markus Wagner ◽  
Ulrich H. Koszinowski
Keyword(s):  
Bacterial Artificial Chromosome ◽  
Fluorescent Protein ◽  
Artificial Chromosome ◽  
Mouse Strains ◽  
Viral Gene ◽  
Wild Type ◽  
Murine Gammaherpesvirus ◽  
Host Interaction ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68

ABSTRACT Gammaherpesviruses cause important infections of humans, in particular in immunocompromised patients. Recently, murine gammaherpesvirus 68 (MHV-68) infection of mice has been developed as a small animal model of gammaherpesvirus pathogenesis. Efficient generation of mutants of MHV-68 would significantly contribute to the understanding of viral gene functions in virus-host interaction, thereby further enhancing the potential of this model. To this end, we cloned the MHV-68 genome as a bacterial artificial chromosome (BAC) inEscherichia coli. During propagation in E. coli, spontaneous recombination events within the internal and terminal repeats of the cloned MHV-68 genome, affecting the copy number of the repeats, were occasionally observed. The gene for the green fluorescent protein was incorporated into the cloned BAC for identification of infected cells. BAC vector sequences were flanked byloxP sites to allow the excision of these sequences using recombinase Cre and to allow the generation of recombinant viruses with wild-type genome properties. Infectious virus was reconstituted from the BAC-cloned MHV-68. Growth of the BAC-derived virus in cell culture was indistinguishable from that of wild-type MHV-68. To assess the feasibility of mutagenesis of the cloned MHV-68 genome, a mutant virus with a deletion of open reading frame 4 was generated. Genetically modified MHV-68 can now be analyzed in functionally modified mouse strains to assess the role of gammaherpesvirus genes in virus-host interaction and pathogenesis.

scholarly journals Primary Clearance of Murine Gammaherpesvirus 68 by PKCθ−/− CD8 T Cells Is Compromised in the Absence of Help from CD4 T Cells

2008 ◽  
Vol 82 (23) ◽  
pp. 11970-11975 ◽  
Author(s):  
Peter Dias ◽  
Ashley L. Shea ◽  
Chandra Inglis ◽  
Francesca Giannoni ◽  
Lian Ni Lee ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell Activation ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Viral Control ◽  
Murine Gammaherpesvirus ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68

ABSTRACT CD4 T cells are dispensable for acute control of murine gammaherpesvirus 68 (MHV-68) but are necessary for effective long-term control of the virus by CD8 T cells. In contrast, protein kinase C θ (PKCθ) is not essential for either acute or long-term viral control. However, we found that while either CD4 or CD8 T cells could mediate the clearance of MHV-68 from the lungs of PKCθ+/+ mice, PKCθ−/− mice depleted of either subset failed to clear the virus. These data suggest that there are two alternative pathways for MHV-68 clearance, one dependent on CD4 T cells and the other on PKCθ. Protection mediated by the latter appears to be short-lived. These observations may help to explain the differential requirement for PKCθ in various models of CD8 T-cell activation and differences in the costimulatory requirements for acute and long-term viral control.

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