KSHV LANA acetylation-selective acidic domain reader sequence mediates virus persistence

Viruses modulate biochemical cellular pathways to permit infection. A recently described mechanism mediates selective protein interactions between acidic domain readers and unacetylated, lysine-rich regions, opposite of bromodomain function. Kaposi´s sarcoma (KS)-associated herpesvirus (KSHV) is tightly linked with KS, primary effusion lymphoma, and multicentric Castleman’s disease. KSHV latently infects cells, and its genome persists as a multicopy, extrachromosomal episome. During latency, KSHV expresses a small subset of genes, including the latency-associated nuclear antigen (LANA), which mediates viral episome persistence. Here we show that LANA contains two tandem, partially overlapping, acidic domain sequences homologous to the SET oncoprotein acidic domain reader. This domain selectively interacts with unacetylated p53, as evidenced by reduced LANA interaction after overexpression of CBP, which acetylates p53, or with an acetylation mimicking carboxyl-terminal domain p53 mutant. Conversely, the interaction of LANA with an acetylation-deficient p53 mutant is enhanced. Significantly, KSHV LANA mutants lacking the acidic domain reader sequence are deficient for establishment of latency and persistent infection. This deficiency was confirmed under physiological conditions, on infection of mice with a murine gammaherpesvirus 68 chimera expressing LANA, where the virus was highly deficient in establishing latent infection in germinal center B cells. Therefore, LANA’s acidic domain reader is critical for viral latency. These results implicate an acetylation-dependent mechanism mediating KSHV persistence and expand the role of acidic domain readers.

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Murine Gammaherpesvirus 68 Expressing Kaposi Sarcoma-Associated Herpesvirus Latency-Associated Nuclear Antigen (LANA) Reveals both Functional Conservation and Divergence in LANA Homologs

Journal of Virology ◽

10.1128/jvi.00992-17 ◽

2017 ◽

Vol 91 (19) ◽

Cited By ~ 7

Author(s):

Arundhati Gupta ◽

Darby G. Oldenburg ◽

Eduardo Salinas ◽

Douglas W. White ◽

J. Craig Forrest

Keyword(s):

Chronic Infection ◽

Ex Vivo ◽

Kaposi Sarcoma ◽

Lytic Replication ◽

Nuclear Antigen ◽

Content Type ◽

Murine Gammaherpesvirus ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

ABSTRACT Latency-associated nuclear antigen (LANA) is a multifunctional protein encoded by members of the Rhadinovirus genus of gammaherpesviruses. Studies using murine gammaherpesvirus 68 (MHV68) demonstrated that LANA is important for acute replication, latency establishment, and reactivation in vivo. Despite structural similarities in their DNA-binding domains (DBDs), LANA homologs from Kaposi sarcoma-associated herpesvirus (KSHV) and MHV68 exhibit considerable sequence divergence. We sought to determine if KSHV and MHV68 LANA homologs are functionally interchangeable. We generated an MHV68 virus that encodes KSHV LANA (kLANA) in place of MHV68 LANA (mLANA) and evaluated the virus's capacity to replicate, establish and maintain latency, and reactivate. kLANA knock-in (KLKI) MHV68 was replication competent in vitro and in vivo but exhibited slower growth kinetics and lower titers than wild-type (WT) MHV68. Following inoculation of mice, KLKI MHV68 established and maintained latency in splenocytes and peritoneal cells but did not reactivate efficiently ex vivo. kLANA repressed the MHV68 promoter for ORF50, the gene that encodes the major lytic transactivator protein RTA, while mLANA did not, suggesting a likely mechanism for the KLKI MHV68 phenotypes. Bypassing this repression by providing MHV68 RTA in trans rescued KLKI MHV68 replication in tissue culture and enabled detection of KLKI MHV68 reactivation ex vivo. These data demonstrate that kLANA and mLANA are functionally interchangeable for establishment and maintenance of latency and suggest that repression of lytic replication by kLANA, as previously shown with KSHV, is a kLANA-specific function that is transferable to MHV68. IMPORTANCE Kaposi sarcoma-associated herpesvirus (KSHV) and murine gammaherpesvirus 68 (MHV68) are members of the Rhadinovirus genus of gammaherpesviruses. These viruses establish lifelong infections that place their respective human and murine hosts at risk for cancer. Latency-associated nuclear antigen (LANA) is a conserved Rhadinovirus protein that is necessary for long-term chronic infection by these viruses. To better understand the conserved functions performed by LANA homologs, we generated a recombinant MHV68 virus that encodes the KSHV LANA protein in place of the MHV68 LANA homolog. We determined that the KSHV LANA protein is capable of supporting MHV68 latency in a mouse model of chronic infection but also functions to repress viral replication. This work describes an in vivo model system for defining evolutionarily conserved and divergent functions of LANA homologs in Rhadinovirus infection and disease.

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Role of CXCR3 in the Immune Response to Murine Gammaherpesvirus 68

Journal of Virology ◽

10.1128/jvi.79.14.9351-9355.2005 ◽

2005 ◽

Vol 79 (14) ◽

pp. 9351-9355 ◽

Cited By ~ 13

Author(s):

Bong Joo Lee ◽

Francesca Giannoni ◽

Ashley Lyon ◽

Shinichiro Yada ◽

Bao Lu ◽

...

Keyword(s):

Immune Response ◽

Cytolytic Activity ◽

Murine Gammaherpesvirus ◽

Latently Infected ◽

Virus Specific Antibody ◽

Antibody Titers ◽

Infected Cells ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

ABSTRACT The chemokine IP-10 (CXCL10) and its cellular receptor CXCR3 are upregulated in the lung during murine gammaherpesvirus 68 (MHV-68) infection. In order to determine the role of the CXCR3 chemokine receptor in the immune response to MHV-68, CXCR3−/− mice were infected with the virus. CXCR3−/− mice showed delayed clearance of replicating MHV-68 from the lungs. This correlated with delayed T-cell recruitment to the lungs and reduced cytolytic activity prior to viral clearance. Splenomegaly and the numbers of latently infected cells per spleen were transiently increased. Ηowever, CXCR3−/− mice showed normal virus-specific antibody titers and effective long-term control of MHV-68 infection.

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A role of hypoxia-inducible factor 1 alpha in Murine Gammaherpesvirus 68 (MHV68) lytic replication and reactivation from latency

PLoS Pathogens ◽

10.1371/journal.ppat.1008192 ◽

2019 ◽

Vol 15 (12) ◽

pp. e1008192 ◽

Cited By ~ 3

Author(s):

Darlah M. López-Rodríguez ◽

Varvara Kirillov ◽

Laurie T. Krug ◽

Enrique A. Mesri ◽

Samita Andreansky

Keyword(s):

Hypoxia Inducible Factor ◽

Lytic Replication ◽

Murine Gammaherpesvirus ◽

Hypoxia Inducible Factor 1 ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

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Absence of the Uracil DNA Glycosylase of Murine Gammaherpesvirus 68 Impairs Replication and Delays the Establishment of LatencyIn Vivo

Journal of Virology ◽

10.1128/jvi.03111-14 ◽

2015 ◽

Vol 89 (6) ◽

pp. 3366-3379 ◽

Cited By ~ 9

Author(s):

Nana Minkah ◽

Marc Macaluso ◽

Darby G. Oldenburg ◽

Clinton R. Paden ◽

Douglas W. White ◽

...

Keyword(s):

Stop Codon ◽

Critical Role ◽

Dna Glycosylase ◽

Uracil Dna Glycosylase ◽

Murine Gammaherpesvirus ◽

Enzymatic Function ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

ABSTRACTUracil DNA glycosylases (UNG) are highly conserved proteins that preserve DNA fidelity by catalyzing the removal of mutagenic uracils. All herpesviruses encode a viral UNG (vUNG), and yet the role of the vUNG in a pathogenic course of gammaherpesvirus infection is not known. First, we demonstrated that the vUNG of murine gammaherpesvirus 68 (MHV68) retains the enzymatic function of host UNG in anin vitroclass switch recombination assay. Next, we generated a recombinant MHV68 with a stop codon in ORF46/UNG (ΔUNG) that led to loss of UNG activity in infected cells and a replication defect in primary fibroblasts. Acute replication of MHV68ΔUNG in the lungs of infected mice was reduced 100-fold and was accompanied by a substantial delay in the establishment of splenic latency. Latency was largely, yet not fully, restored by an increase in virus inoculum or by altering the route of infection. MHV68 reactivation from latent splenocytes was not altered in the absence of the vUNG. A survey of host UNG activity in cells and tissues targeted by MHV68 indicated that the lung tissue has a lower level of enzymatic UNG activity than the spleen. Taken together, these results indicate that the vUNG plays a critical role in the replication of MHV68 in tissues with limited host UNG activity and this vUNG-dependent expansion, in turn, influences the kinetics of latency establishment in distal reservoirs.IMPORTANCEHerpesviruses establish chronic lifelong infections using a strategy of replicative expansion, dissemination to latent reservoirs, and subsequent reactivation for transmission and spread. We examined the role of the viral uracil DNA glycosylase, a protein conserved among all herpesviruses, in replication and latency of murine gammaherpesvirus 68. We report that the viral UNG of this murine pathogen retains catalytic activity and influences replication in culture. The viral UNG was impaired for productive replication in the lung. This defect in expansion at the initial site of acute replication was associated with a substantial delay of latency establishment in the spleen. The levels of host UNG were substantially lower in the lung compared to the spleen, suggesting that herpesviruses encode a viral UNG to compensate for reduced host enzyme levels in some cell types and tissues. These data suggest that intervention at the site of initial replicative expansion can delay the establishment of latency, a hallmark of chronic herpesvirus infection.

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Murine Gammaherpesvirus 68 LANA Is Essential for Virus Reactivation from Splenocytes but Not Long-Term Carriage of Viral Genome

Journal of Virology ◽

10.1128/jvi.00133-10 ◽

2010 ◽

Vol 84 (14) ◽

pp. 7214-7224 ◽

Cited By ~ 22

Author(s):

Clinton R. Paden ◽

J. Craig Forrest ◽

Nathaniel J. Moorman ◽

Samuel H. Speck

Keyword(s):

Viral Genome ◽

Nuclear Antigen ◽

Type I ◽

Virus Reactivation ◽

Intranasal Inoculation ◽

Murine Gammaherpesvirus ◽

Marker Rescue ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68 ◽

Rescue Virus

ABSTRACT ORF73, which encodes the latency-associated nuclear antigen (LANA), is a conserved gamma-2-herpesvirus gene. The murine gammaherpesvirus 68 (MHV68) LANA (mLANA) is critical for efficient virus replication and the establishment of latent infection following intranasal inoculation. To test whether the initial host immune response limits the capacity of mLANA-null virus to traffic to and establish latency in the spleen, we infected type I interferon receptor knockout (IFN-α/βR−/−) mice via intranasal inoculation and observed the presence of viral genome-positive splenocytes at day 18 postinfection at approximately 10-fold-lower levels than in the genetically repaired marker rescue-infected mice. However, no mLANA-null virus reactivation from infected IFN-α/βR−/− splenocytes was observed. To more thoroughly define a role of mLANA in MHV68 infection, we evaluated the capacity of an mLANA-null virus to establish and maintain infection apart from restriction in the lungs of immunocompetent mice. At day 18 following intraperitoneal infection of C57BL/6 mice, the mLANA-null virus was able to establish a chronic infection in the spleen albeit at a 5-fold-reduced level. However, as in IFN-α/βR−/− mice, little or no virus reactivation could be detected from mLANA-null virus-infected splenocytes upon explant. An examination of peritoneal exudate cells (PECs) following intraperitoneal inoculation revealed nearly equivalent frequencies of PECs harboring the mLANA-null virus relative to the marker rescue virus. Furthermore, although significantly compromised, mLANA-null virus reactivation from PECs was detected upon explant. Notably, at later times postinfection, the frequency of mLANA-null genome-positive splenocytes was indistinguishable from that of marker rescue virus-infected animals. Analyses of viral genome-positive splenocytes revealed the absence of viral episomes in mLANA-null infected mice, suggesting that the viral genome is integrated or maintained in a linear state. Thus, these data provide the first evidence that a LANA homolog is directly involved in the formation and/or maintenance of an extrachromosomal viral episome in vivo, which is likely required for the reactivation of MHV68.

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Endosomal Toll-Like Receptors 7 and 9 Cooperate in Detection of Murine Gammaherpesvirus 68 Infection

Journal of Virology ◽

10.1128/jvi.01173-18 ◽

2018 ◽

Vol 93 (3) ◽

Cited By ~ 8

Author(s):

Kendra A. Bussey ◽

Sripriya Murthy ◽

Elisa Reimer ◽

Baca Chan ◽

Bastian Hatesuer ◽

...

Keyword(s):

Epstein Barr Virus ◽

Toll Like Receptor ◽

Murine Gammaherpesvirus ◽

Barr Virus ◽

Epstein Barr ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68 ◽

Deficient Mice

ABSTRACTMurine gammaherpesvirus 68 (MHV68) is a small-animal model suitable for study of the human pathogens Epstein-Barr virus and Kaposi’s sarcoma-associated herpesvirus. Here, we have characterized the roles of the endosomal Toll-like receptor (TLR) escort protein UNC93B, endosomal TLR7, -9, and -13, and cell surface TLR2 in MHV68 detection. We found that the alpha interferon (IFN-α) response of plasmacytoid dendritic cells (pDC) to MHV68 was reduced inTlr9−/−cells compared to levels in wild type (WT) cells but not completely lost.Tlr7−/−pDC responded similarly to WT. However, we found that inUnc93b−/−pDC, as well as inTlr7−/−Tlr9−/−double-knockout pDC, the IFN-α response to MHV68 was completely abolished. Thus, the only pattern recognition receptors contributing to the IFN-α response to MHV68 in pDC are TLR7 and TLR9, but the contribution of TLR7 is masked by the presence of TLR9. To address the role of UNC93B and TLR for MHV68 infectionin vivo, we infected mice with MHV68. Lytic replication of MHV68 after intravenous infection was enhanced in the lungs, spleen, and liver of UNC93B-deficient mice, in the spleen of TLR9-deficient mice, and in the liver and spleen ofTlr7−/−Tlr9−/−mice. The absence of TLR2 or TLR13 did not affect lytic viral titers. We then compared reactivation of MHV68 from latently infected WT,Unc93b−/−,Tlr7−/−Tlr9−/−,Tlr7−/−, andTlr9−/−splenocytes. We observed enhanced reactivation and latent viral loads, particularly fromTlr7−/−Tlr9−/−splenocytes compared to levels in the WT. Our data show that UNC93B-dependent TLR7 and TLR9 cooperate in and contribute to detection and control of MHV68 infection.IMPORTANCEThe two human gammaherpesviruses, Epstein-Barr virus (EBV) and Kaposi’s sarcoma-associated herpesvirus (KSHV), can cause aggressive forms of cancer. These herpesviruses are strictly host specific, and therefore the homolog murine gammaherpesvirus 68 (MHV68) is a widely used model to obtainin vivoinsights into the interaction between these two gammaherpesviruses and their host. Like EBV and KSHV, MHV68 establishes lifelong latency in B cells. The innate immune system serves as one of the first lines of host defense, with pattern recognition receptors such as the Toll-like receptors playing a crucial role in mounting a potent antiviral immune response to various pathogens. Here, we shed light on a yet unanticipated role of Toll-like receptor 7 in the recognition of MHV68 in a subset of immune cells called plasmacytoid dendritic cells, as well as on the control of this virus in its host.

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Establishment and Maintenance of Long-Term Murine Gammaherpesvirus 68 Latency in B Cells in the Absence of CD40

Journal of Virology ◽

10.1128/jvi.79.5.2891-2899.2005 ◽

2005 ◽

Vol 79 (5) ◽

pp. 2891-2899 ◽

Cited By ~ 19

Author(s):

David O. Willer ◽

Samuel H. Speck

Keyword(s):

B Cells ◽

B Cell ◽

Virus Replication ◽

Receptor Expression ◽

Cell Phenotype ◽

Memory B Cell ◽

Murine Gammaherpesvirus ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

ABSTRACT Murine gammaherpesvirus 68 (γHV68), like Epstein-Barr virus (EBV), establishes a chronic infection in its host by gaining access to the memory B-cell reservoir, where it persists undetected by the host's immune system. EBV encodes a membrane protein, LMP1, that appears to function as a constitutively active CD40 receptor, and is hypothesized to play a central role in EBV-driven differentiation of infected naive B cells to a memory B-cell phenotype. However, it has recently been shown that there is a critical role for CD40-CD40L interaction in B-cell immortalization by EBV (K.-I. Imadome, M. Shirakata, N. Shimizu, S. Nonoyama, and Y. Yamanashi, Proc. Natl. Acad. Sci. USA 100:7836-7840, 2003), indicating that LMP1 does not adequately recapitulate all of the necessary functions of CD40. The role of CD40 receptor expression on B cells for the establishment and maintenance of γHV68 latency is unclear. Data previously obtained with a competition model, demonstrated that in the face of CD40-sufficient B cells, γHV68 latency in CD40-deficient B cells waned over time in chimeric mice (I.-J. Kim, E. Flano, D. L. Woodland, F. E. Lund, T. D. Randall, and M. A. Blackman, J. Immunol. 171:886-892, 2003). To further investigate the role of CD40 in γHV68 latency in vivo, we have characterized the infection of CD40 knockout (CD40−/−) mice. Here we report that, consistent with previous observations, γHV68 efficiently established a latent infection in B cells of CD40−/− mice. Notably, unlike the infection of normal C57BL/6 mice, significant ex vivo reactivation from splenocytes harvested from infected CD40−/− mice 42 days postinfection was observed. In addition, in contrast to γHV68 infection of C57BL/6 mice, the frequency of infected naive B cells remained fairly stable over a 3-month period postinfection. Furthermore, a slightly higher frequency of γHV68 infection was observed in immunoglobulin D (IgD)-negative B cells, which was stably maintained over a period of 3 months postinfection. The presence of virus in IgD-negative B cells indicates that γHV68 may either directly infect memory B cells present in CD40−/− mice or be capable of driving differentiation of naive CD40−/− B cells. A possible explanation for the apparent discrepancy between the failure of γHV68 latency to be maintained in CD40-deficient B cells in the presence of CD40-sufficient B cells and the stable maintenance of γHV68 B-cell latency in CD40−/− mice came from examining virus replication in the lungs of infected CD40−/− mice, where we observed significantly higher levels of virus replication at late times postinfection compared to those in infected C57BL/6 mice. Taken together, these findings are consistent with a model in which chronic virus infection of CD40−/− mice is maintained through virus reactivation in the lungs and reseeding of latency reservoirs.

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Role for MyD88 Signaling in Murine Gammaherpesvirus 68 Latency

Journal of Virology ◽

10.1128/jvi.02577-07 ◽

2008 ◽

Vol 82 (8) ◽

pp. 3853-3863 ◽

Cited By ~ 37

Author(s):

Lisa M. Gargano ◽

Janice M. Moser ◽

Samuel H. Speck

Keyword(s):

B Cells ◽

B Cell ◽

Cell Response ◽

Wild Type ◽

Tlr Signaling ◽

Murine Gammaherpesvirus ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68 ◽

B Cell Response

ABSTRACT Toll-like receptors (TLRs) are known predominantly for their role in activating the innate immune response. Recently, TLR signaling via MyD88 has been reported to play an important function in development of a B-cell response. Since B cells are a major latency reservoir for murine gammaherpesvirus 68 (MHV68), we investigated the role of TLR signaling in the establishment and maintenance of MHV68 latency in vivo. Mice deficient in MyD88 (MyD88−/−) or TLR3 (TLR3−/−) were infected with MHV68. Analysis of splenocytes recovered at day 16 postinfection from MyD88−/− mice compared to those from wild-type control mice revealed a lower frequency of (i) activated B cells, (ii) germinal-center B cells, and (iii) class-switched B cells. Accompanying this substantial defect in the B-cell response was an approximately 10-fold decrease in the establishment of splenic latency. In contrast, no defect in viral latency was observed in TLR3−/− mice. Analysis of MHV68-specific antibody responses also demonstrated a substantial decrease in the kinetics of the response in MyD88−/− mice. Analysis of wild-type × MyD88−/− mixed-bone-marrow chimeric mice demonstrated that there is a selective failure of MyD88−/− B cells to participate in germinal-center reactions as well as to become activated and undergo class switching. In addition, while MHV68 established latency efficiently in the MyD88-sufficient B cells, there was again a ca. 10-fold reduction in the frequency of MyD88−/− B cells harboring latent MHV68. This phenotype indicates that MyD88 is important for the establishment of MHV68 latency and is directly related to the role of MyD88 in the generation of a B-cell response. Furthermore, the generation of a B-cell response to MHV68 was intrinsic to B cells and was independent of the interleukin-1 receptor, a cytokine receptor that also signals through MyD88. These data provide evidence for a unique role for MyD88 in the establishment of MHV68 latency.

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A Gammaherpesvirus Ubiquitin-Specific Protease Is Involved in the Establishment of Murine Gammaherpesvirus 68 Infection

Journal of Virology ◽

10.1128/jvi.01017-09 ◽

2009 ◽

Vol 83 (20) ◽

pp. 10644-10652 ◽

Cited By ~ 20

Author(s):

Sara Gredmark-Russ ◽

Marisa K. Isaacson ◽

Lisa Kattenhorn ◽

Evelyn J. Cheung ◽

Nicki Watson ◽

...

Keyword(s):

Acute Infection ◽

Mutant Virus ◽

Tegument Protein ◽

Murine Gammaherpesvirus ◽

Enzymatic Function ◽

Significant Difference ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

ABSTRACT Murine gammaherpesvirus 68 (MHV-68) contains a ubiquitin (Ub)-specific cysteine protease (USP) domain embedded within the large tegument protein ORF64, as do all other herpesviruses. The biological role of this protease is still unclear, but for the alphaherpesvirus Marek's disease virus, its USP is involved in T-cell lymphoma formation. We here study the role of the MHV-68 USP, encoded by ORF64. By constructing a mutant virus with a single cysteine-to-alanine replacement in the active site of ORF64, we demonstrate that the USP activity of ORF64 is abolished. The mutant virus replicates less efficiently in vitro, and plaque size is reduced compared to that of a revertant virus. Electron microscopy of infected cells did not reveal any obvious differences in virion morphogenesis or differences in egress for the mutant and revertant viruses. Intraperitoneal infection of C57/BL6 mice demonstrates that the mutant virus is generally cleared by day 7, indicating a role for the USP in the persistence of MHV-68 infection or efficient replication. However, the USP activity in MHV-68 is unlikely to be involved in the establishment of latency or reactivation, since we observed no significant difference in viral DNA genome copy number in the spleen or in the number of cells that reactivate MHV-68 from latency. Our results for MHV-68 ORF64 are consistent with an enzymatic function of the tegument protein that is beneficial to the virus during acute infection, particularly in vivo.

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Protein Kinase C θ Is Not Essential for T-Cell-Mediated Clearance of Murine Gammaherpesvirus 68

Journal of Virology ◽

10.1128/jvi.79.11.6808-6813.2005 ◽

2005 ◽

Vol 79 (11) ◽

pp. 6808-6813 ◽

Cited By ~ 31

Author(s):

Francesca Giannoni ◽

Ashley B. Lyon ◽

Mark D. Wareing ◽

Peter B. Dias ◽

Sally R. Sarawar

Keyword(s):

T Cell ◽

T Cell Activation ◽

Cell Activation ◽

Cytokine Production ◽

Viral Clearance ◽

Murine Gammaherpesvirus ◽

Kinase C ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

ABSTRACT Murine gammaherpesvirus 68 (MHV-68) is a naturally occurring rodent pathogen with significant homology to human pathogens Epstein-Barr virus and Kaposi's sarcoma-associated herpesvirus. T cells are essential for primary clearance of MHV-68 and survival of mice following intranasal infection. Previous reports have suggested that protein kinase C θ (PKCθ) is essential for T-cell activation and cytokine production in vitro. To determine the role of this molecule in vivo during the immune response to a viral infection, PKCθ−/− mice were infected with MHV-68. Despite the essential role of T cells in viral clearance, PKCθ−/− mice survived infection, cleared lytic virus, and maintained effective long-term control of latency. CD8 T-cell expansion, trafficking to the lung, and cytotoxic activity were similar in PKCθ+/+ and PKCθ−/− mice, whereas antiviral antibody and T-helper cell cytokine production were significantly lower in PKCθ−/− mice than in PKCθ+/+ mice. These studies demonstrate a differential requirement for PKCθ in the immune response to MHV-68 and show that PKCθ is not essential for the T-cell activation events leading to viral clearance.

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