scholarly journals CD4 T Cells Mediate Killing during Persistent Gammaherpesvirus 68 Infection

2009 ◽  
Vol 83 (9) ◽  
pp. 4700-4703 ◽  
Author(s):  
Kathleen A. Stuller ◽  
Emilio Flaño
Keyword(s):  
T Cells ◽  
Cd4 T Cells ◽  
Persistently Infected ◽  
Murine Gammaherpesvirus ◽  
Cytotoxicity Assays ◽  
Gammaherpesvirus 68 ◽  
In Vivo Cytotoxicity ◽  
Murine Gammaherpesvirus 68

ABSTRACT CD4 T cells are critical for the control of gammaherpesvirus persistence, but their direct effector mechanisms of virus control in vivo are still poorly understood. In this study, we use murine gammaherpesvirus 68 (γHV68) in in vitro and in vivo cytotoxicity assays to show CD4-dependent killing of γHV68-loaded cells in mice persistently infected with γHV68. Our results underscore the cytotoxic capacity of CD4 T cells during γHV68 persistence.

scholarly journals Characterization of a Spontaneous 9.5-Kilobase-Deletion Mutant of Murine Gammaherpesvirus 68 Reveals Tissue-Specific Genetic Requirements for Latency

2002 ◽  
Vol 76 (13) ◽  
pp. 6532-6544 ◽  
Author(s):  
Eric T. Clambey ◽  
Herbert W. Virgin ◽  
Samuel H. Speck
Keyword(s):  
Deletion Mutant ◽  
Latent Infection ◽  
Loss Of Function ◽  
Murine Gammaherpesvirus ◽  
Peritoneal Cells ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68

ABSTRACT Murine gammaherpesvirus 68 (γHV68 [also known as MHV-68]) establishes a latent infection in mice, providing a small-animal model with which to identify host and viral factors that regulate gammaherpesvirus latency. While γHV68 establishes a latent infection in multiple tissues, including splenocytes and peritoneal cells, the requirements for latent infection within these tissues are poorly defined. Here we report the characterization of a spontaneous 9.5-kb-deletion mutant of γHV68 that lacks the M1, M2, M3, and M4 genes and eight viral tRNA-like genes. Previously, this locus has been shown to contain the latency-associated M2, M3, and viral tRNA-like genes. Through characterization of this mutant, we found that the M1, M2, M3, M4 genes and the viral tRNA-like genes are dispensable for (i) in vitro replication and (ii) the establishment and maintenance of latency in vivo and reactivation from latency following intraperitoneal infection. In contrast, following intranasal infection with this mutant, there was a defect in splenic latency at both early and late times, a phenotype not observed in peritoneal cells. These results indicate (i) that there are different genetic requirements for the establishment of latency in different latent reservoirs and (ii) that the genetic requirements for latency depend on the route of infection. While some of these phenotypes have been observed with specific mutations in the M1 and M2 genes, other phenotypes have never been observed with the available γHV68 mutants. These studies highlight the importance of loss-of-function mutations in defining the genetic requirements for the establishment and maintenance of herpesvirus latency.

scholarly journals Disruption of CCL21-Induced Chemotaxis In Vitro and In Vivo by M3, a Chemokine-Binding Protein Encoded by Murine Gammaherpesvirus 68

2003 ◽  
Vol 77 (1) ◽  
pp. 624-630 ◽  
Author(s):  
Kristian K. Jensen ◽  
Shu-Cheng Chen ◽  
R. William Hipkin ◽  
Maria T. Wiekowski ◽  
Martin A. Schwarz ◽  
...  
Keyword(s):  
Lymphoid Tissues ◽  
Transgenic Expression ◽  
Murine Gammaherpesvirus ◽  
Chemokine Signaling ◽  
Specific Subset ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68 ◽  
Human Chemokines

ABSTRACT Chemokine-binding proteins represent a novel class of antichemokine agents encoded by poxviruses and herpesviruses. One such protein is encoded by the M3 gene present in the murine gammaherpesvirus 68 (MHV-68) genome. The M3 gene encodes a secreted 44-kDa protein that binds with high affinity to certain murine and human chemokines and has been shown to block chemokine signaling in vitro. However, there has been no direct evidence that M3 blocks chemokine activity in vivo, nor has the nature of M3-chemokine interaction been defined. To better understand the ability of M3 to block chemokine activity in vivo, we examined its interaction with a specific subset of chemokines expressed in lymphoid tissues, areas where gammaherpesviruses characteristically establish latency. Here we show that M3 blocks in vitro chemotaxis induced by CCL19 and CCL21, chemokines expressed constitutively in secondary lymphoid tissues. Moreover, we provide evidence that chemokine M3 binding exhibits positive cooperativity. In vivo, the expression of M3 in the pancreas of transgenic mice inhibits recruitment of lymphocytes induced by transgenic expression of CCL21 in this organ. The ability of M3 to block the biological activity of chemokines may represent an important strategy used by MHV-68 to evade immune detection and favor viral replication in the infected host.

scholarly journals Murine Gammaherpesvirus 68 ORF48 Is an RTA-Responsive Gene Product and Functions in both Viral Lytic Replication and Latency duringIn VivoInfection

2015 ◽  
Vol 89 (11) ◽  
pp. 5788-5800 ◽  
Author(s):  
Jing Qi ◽  
Chuanhui Han ◽  
Danyang Gong ◽  
Ping Liu ◽  
Sheng Zhou ◽  
...  
Keyword(s):  
Lytic Replication ◽  
Laboratory Mice ◽  
Transcription Activator ◽  
Murine Gammaherpesvirus ◽  
Responsive Gene ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68 ◽  
Lytic Genes

ABSTRACTReplication and transcription activator (RTA) of gammaherpesvirus is an immediate early gene product and regulates the expression of many downstream viral lytic genes. ORF48 is also conserved among gammaherpesviruses; however, its expression regulation and function remained largely unknown. In this study, we characterized the transcription unit ofORF48from murine gammaherpesvirus 68 (MHV-68) and analyzed its transcriptional regulation. We showed that RTA activates theORF48promoter via an RTA-responsive element (48pRRE). RTA binds to 48pRRE directlyin vitroand also associates with ORF48 promoterin vivo. Mutagenesis of 48pRRE in the context of the viral genome demonstrated that the expression of ORF48 is activated by RTA through 48pRRE duringde novoinfection. Through site-specific mutagenesis, we generated an ORF48-null virus and examined the function of ORF48in vitroandin vivo. The ORF48-null mutation remarkably reduced the viral replication efficiency in cell culture. Moreover, through intranasal or intraperitoneal infection of laboratory mice, we showed that ORF48 is important for viral lytic replication in the lung and establishment of latency in the spleen, as well as viral reactivation from latency. Collectively, our study identifiedORF48as an RTA-responsive gene and showed that ORF48 is important for MHV-68 replication bothin vitroandin vivo.IMPORTANCEThe replication and transcription activator (RTA), conserved among gammaherpesviruses, serves as a molecular switch for the virus life cycle. It works as a transcriptional regulator to activate the expression of many viral lytic genes. However, only a limited number of such downstream genes have been uncovered for MHV-68. In this study, we identifiedORF48as an RTA-responsive gene of MHV-68 and mapped theciselement involved. By constructing a mutant virus that is deficient in ORF48 expression and through infection of laboratory mice, we showed that ORF48 plays important roles in different stages of viral infectionin vivo. Our study provides insights into the transcriptional regulation and protein function of MHV-68, a desired model for studying gammaherpesviruses.

scholarly journals Murine gammaherpesvirus-68 ORF28 encodes a non-essential virion glycoprotein

2005 ◽  
Vol 86 (4) ◽  
pp. 919-928 ◽  
Author(s):  
Janet S. May ◽  
Heather M. Coleman ◽  
Jessica M. Boname ◽  
Philip G. Stevenson
Keyword(s):  
Function Analysis ◽  
Lytic Replication ◽  
Specific Gene ◽  
Virion Protein ◽  
Major Function ◽  
Murine Gammaherpesvirus ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68

Murine gammaherpesvirus-68 (MHV-68) ORF28 is a gammaherpesvirus-specific gene of unknown function. Analysis of epitope-tagged ORF28 protein indicated that it was membrane-associated and incorporated into virions in N-glycosylated, O-glycosylated and unglycosylated forms. The extensive glycosylation of the small ORF28 extracellular domain – most forms of the protein appeared to be mainly carbohydrate by weight – suggested that a major function of ORF28 is to attach a variety of glycans to the virion surface. MHV-68 lacking ORF28 showed normal lytic replication in vitro and in vivo and normal latency establishment. MHV-68 ORF28 therefore encodes a small, membrane-bound and extensively glycosylated virion protein, whose function is entirely dispensable for normal, single-cycle host colonization.

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.

scholarly journals ORF23 of murine gammaherpesvirus 68 is non-essential for in vitro and in vivo infection

2012 ◽  
Vol 93 (5) ◽  
pp. 1076-1080 ◽  
Author(s):  
S. Ohno ◽  
B. Steer ◽  
C. Sattler ◽  
H. Adler
Keyword(s):  
Deletion Mutant ◽  
Protein Product ◽  
Lytic Replication ◽  
Murine Gammaherpesvirus ◽  
The Face ◽  
Infected Cells ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68

Although ORF23 is conserved among gammaherpesviruses, its role during infection is unknown. Here, we studied the expression of ORF23 of murine gammaherpesvirus 68 (MHV-68) and its role during infection. ORF23 mRNA was detected in infected cells as a late transcript. The ORF23 protein product could be expressed and detected as an N-terminally FLAG-tagged protein by Western blot and indirect immunofluorescence. To investigate the role of ORF23 in the infection cycle of a gammaherpesvirus, we constructed an ORF23 deletion mutant of MHV-68. The analysis of the ORF23 deletion mutant suggested that ORF23 of MHV-68 is neither essential for replication in cell culture nor for lytic or latent infection in vivo. A phenotype of the ORF23 deletion mutant, reflected by a moderate reduction in lytic replication and latency amplification, was only detectable in the face of direct competition to the parental virus.

scholarly journals The M10 Locus of Murine Gammaherpesvirus 68 Contributes to both the Lytic and the Latent Phases of Infection

2009 ◽  
Vol 83 (16) ◽  
pp. 8163-8172 ◽  
Author(s):  
B. Flach ◽  
B. Steer ◽  
N. N. Thakur ◽  
J. Haas ◽  
H. Adler
Keyword(s):  
Lytic Replication ◽  
Mutant Virus ◽  
Wild Type ◽  
Coding Region ◽  
Murine Gammaherpesvirus ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68 ◽  
Revertant Virus

ABSTRACT Murine gammaherpesvirus 68 (MHV-68) is closely related to Epstein-Barr virus and Kaposi's sarcoma-associated herpesvirus (KSHV) and provides a small-animal model to study the pathogenesis of gammaherpesvirus (γHV) infections. According to the colinear organization of the γHV genomes, the M10 locus is situated at a position equivalent to the K12 locus of KSHV, which codes for proteins of the kaposin family. The M10 locus of MHV-68 has been predicted to code for three overlapping open reading frames (M10a, M10b, and M10c [M10a-c]) with unknown function. In addition, the M10 locus contains a lytic origin of replication (oriLyt). To elucidate the function of the M10 locus during lytic and latent infections, we investigated, both in vitro and in vivo, the following four recombinant viruses which were generated using MHV-68 cloned as a bacterial artificial chromosome: (i) a mutant virus with a deletion which affects both the coding region for M10a-c and the oriLyt; (ii) a revertant virus in which both the M10a-c coding region and the oriLyt were reverted to those of the wild type; (iii) a virus with an ectopic insertion of the oriLyt, which restores the function of the oriLyt but not the M10a-c coding region; and (iv) a mutant virus with a deletion in the oriLyt only. While the mutants were slightly attenuated with regard to lytic replication in cell culture, they showed severe growth defects in vivo. Both lytic replication and latency amplification were strongly reduced. In contrast, both the revertant virus and the virus with the ectopic oriLyt insertion grew very similarly to the parental wild-type virus both in vitro and in vivo. Thus, we provide genetic evidence that mutation of the oriLyt, and not of putative protein coding sequences within the M10a-c region, is responsible for the observed phenotype. We conclude that the oriLyt in the M10 locus plays an important role during infection of mice with MHV-68.

scholarly journals Murine Gammaherpesvirus 68 Open Reading Frame 11 Encodes a Nonessential Virion Component

2005 ◽  
Vol 79 (5) ◽  
pp. 3163-3168 ◽  
Author(s):  
Jessica M. Boname ◽  
Janet S. May ◽  
Philip G. Stevenson
Keyword(s):  
Lytic Replication ◽  
Open Reading Frame ◽  
Reading Frame ◽  
Murine Gammaherpesvirus ◽  
Infected Cells ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68

ABSTRACT Open reading frame 11 (ORF11) is a conserved gammaherpesvirus gene that remains undefined. We identified the product of murine gammaherpesvirus 68 (MHV-68) ORF11, p43, as a virion component with a predominantly perinuclear distribution in infected cells. MHV-68 lacking p43 grew normally in vitro but showed reduced lytic replication in vivo and a delay in seeding to the spleen. Subsequent latency amplification was normal. Thus, MHV-68 ORF11 encoded a virion component that was important for in vivo lytic replication but dispensable for the establishment of latency.

scholarly journals Murine gammaherpesvirus 68 bcl-2 homologue contributes to latency establishment in vivo

2005 ◽  
Vol 86 (1) ◽  
pp. 31-40 ◽  
Author(s):  
Brigitte D. de Lima ◽  
Janet S. May ◽  
Sofia Marques ◽  
J. Pedro Simas ◽  
Philip G. Stevenson
Keyword(s):  
Cell Activation ◽  
Ex Vivo ◽  
Lytic Replication ◽  
B Cell Activation ◽  
Murine Gammaherpesvirus ◽  
Gammaherpesvirus 68 ◽  
Murine Gammaherpesvirus 68

The gammaherpesviruses are characteristically latent in lymphocytes and exploit lymphocyte proliferation to establish a large, persistent pool of latent genomes. Murine gammaherpesvirus 68 (MHV-68) allows the in vivo analysis of viral genes that contribute to this and other aspects of host colonization. In this study, the MHV-68 bcl-2 homologue, M11, was disrupted either in its BH1 homology domain or upstream of its membrane-localizing C-terminal domain. Each M11 mutant showed normal lytic replication in vitro and in vivo, but had a reduction in peak splenic latency. Lower infectious-centre titres correlated with lower in vivo B-cell activation, lower viral genome loads and reduced viral tRNA expression. This was therefore a true latency deficit, rather than a deficit in ex vivo reactivation. Stable, long-term levels of splenic latency were normal. M11 function therefore contributed specifically to viral latency amplification in infected lymphoid tissue.

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