De Novo Infection of B Cells during Murine Gammaherpesvirus 68 Latency

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.

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Evidence for a Multiprotein Gamma-2 Herpesvirus Entry Complex

Journal of Virology ◽

10.1128/jvi.01141-07 ◽

2007 ◽

Vol 81 (23) ◽

pp. 13082-13091 ◽

Cited By ~ 21

Author(s):

Laurent Gillet ◽

Philip G. Stevenson

Keyword(s):

B Cells ◽

Specific Antibody ◽

Cell Binding ◽

Murine Gammaherpesvirus ◽

Single Entry ◽

Single Complex ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68 ◽

Better Than

ABSTRACT Herpesviruses use multiple virion glycoproteins to enter cells. How these work together is not well understood: some may act separately or they may form a single complex. Murine gammaherpesvirus 68 (MHV-68) gB, gH, gL, and gp150 all participate in entry. gB and gL are involved in binding, gB and gH are conserved fusion proteins, and gp150 inhibits cell binding until glycosaminoglycans are engaged. Here we show that a gH-specific antibody coprecipitates gB and thus that gH and gB are associated in the virion membrane. A gH/gL-specific antibody also coprecipitated gB, implying a tripartite complex of gL/gH/gB, although the gH/gB association did not require gL. The association was also independent of gp150, and gp150 was not demonstrably bound to gB or gH. However, gp150 incorporation into virions was partly gL dependent, suggesting that it too contributes to a single entry complex. gp150− and gL− gp150− mutants bound better than the wild type to B cells and readily colonized B cells in vivo. Thus, gp150 and gL appear to be epithelial cell-adapted accessories of a core gB/gH entry complex. The cell binding revealed by gp150 disruption did not require gL and therefore seemed most likely to involve gB.

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The Murine Gammaherpesvirus 68 M2 Gene Is Required for Efficient Reactivation from Latently Infected B Cells

Journal of Virology ◽

10.1128/jvi.00066-11 ◽

2011 ◽

Vol 85 (6) ◽

pp. 3041-3041

Author(s):

J. H. Herskowitz ◽

M. A. Jacoby ◽

S. H. Speck

Keyword(s):

B Cells ◽

Murine Gammaherpesvirus ◽

Latently Infected ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

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The Absence of M1 Leads to Increased Establishment of Murine Gammaherpesvirus 68 Latency in IgD-Negative B Cells

Journal of Virology ◽

10.1128/jvi.01953-12 ◽

2013 ◽

Vol 87 (6) ◽

pp. 3597-3604 ◽

Cited By ~ 5

Author(s):

L. T. Krug ◽

A. G. Evans ◽

L. M. Gargano ◽

C. R. Paden ◽

S. H. Speck

Keyword(s):

B Cells ◽

Murine Gammaherpesvirus ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

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

Journal of Virology ◽

10.1128/jvi.00297-09 ◽

2009 ◽

Vol 83 (13) ◽

pp. 6484-6493 ◽

Cited By ~ 55

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.

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Function of Rta Is Essential for Lytic Replication of Murine Gammaherpesvirus 68

Journal of Virology ◽

10.1128/jvi.75.19.9262-9273.2001 ◽

2001 ◽

Vol 75 (19) ◽

pp. 9262-9273 ◽

Cited By ~ 71

Author(s):

Ting-Ting Wu ◽

Leming Tong ◽

Tammy Rickabaugh ◽

Samuel Speck ◽

Ren Sun

Keyword(s):

Cell Lines ◽

De Novo ◽

Lytic Replication ◽

Dominant Negative ◽

Early Gene ◽

Productive Infection ◽

Murine Gammaherpesvirus ◽

C Terminus ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

ABSTRACT Rta, encoded primarily by open reading frame 50, is well conserved among gammaherpesviruses. It has been shown that the Rta proteins of Epstein Barr virus (EBV), Kaposi's sarcoma-associated herpesvirus (KSHV, or HHV-8), and murine gammaherpesvirus 68 (MHV-68; also referred to as γHV68) play an important role in viral reactivation from latency. However, the role of Rta during productive de novo infection has not been characterized in gammaherpesviruses. Since there are cell lines that can support efficient productive de novo infection by MHV-68 but not EBV or KSHV, we examined whether MHV-68 Rta plays a role in initiating viral lytic replication in productively infected cells. Rta, functioning as a transcriptional activator, can activate the viral promoter of early lytic genes. The amino acid sequence alignments of the Rta homologues suggest that the organizations of their functional domains are similar, with the DNA binding and dimerization domains at the N terminus and the trans-activation domain at the C terminus. We constructed two mutants of MHV-68 Rta, Rd1 and Rd2, with deletions of 112 and 243 amino acids from the C terminus, respectively. Rd1 and Rd2 could no longer trans-activate the promoter of MHV-68 gene 57, consistent with the deletions of theirtrans-activation domains at the C terminus. Furthermore, Rd1 and Rd2 were able to function as dominant-negative mutants, inhibiting trans-activation of wild-type Rta. To study whether Rd1 and Rd2 blocked viral lytic replication, purified virion DNA was cotransfected with Rd1 or Rd2 into fibroblasts. Expression of viral lytic proteins was greatly suppressed, and the yield of infectious viruses was reduced up to 104-fold. Stable cell lines constitutively expressing Rd2 were established and infected with MHV-68. Transcription of the immediate-early gene, rta, and the early gene, tk, of the virus was reduced in these cell lines. The presence of Rd2 also led to attenuation of viral lytic protein expression and virion production. The ability of Rta dominant-negative mutants to inhibit productive infection suggests that the trans-activation function of Rta is essential for MHV-68 lytic replication. We propose that a single viral protein, Rta, governs the initiation of MHV-68 lytic replication during both reactivation and productive de novo infection.

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Selective Gene Expression of Latent Murine Gammaherpesvirus 68 in B Lymphocytes

Journal of Virology ◽

10.1128/jvi.77.13.7308-7318.2003 ◽

2003 ◽

Vol 77 (13) ◽

pp. 7308-7318 ◽

Cited By ~ 98

Author(s):

Sofia Marques ◽

Stacey Efstathiou ◽

K. G. Smith ◽

Matthias Haury ◽

J. Pedro Simas

Keyword(s):

Gene Expression ◽

B Cells ◽

B Cell ◽

Germinal Center ◽

Critical Role ◽

Murine Gammaherpesvirus ◽

Virus Transcription ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68 ◽

Germinal Center B Cells

ABSTRACT Intranasal infection of mice with murine gammaherpesvirus 68 (MHV-68), a virus genetically related to the human pathogen Kaposi's sarcoma-associated herpesvirus, results in a persistent, latent infection in the spleen and other lymphoid organs. Here, we have determined the frequency of virus infection in splenic dendritic cells, macrophages, and several B-cell subpopulations, and we quantified cell type-dependent virus transcription patterns. The frequencies of virus genome positive cells were maximal at 14 days postinfection in all splenic cell populations analyzed. Marginal zone and germinal center B cells harbored the highest frequency of infection and the former population accounted for approximately half the total number of infected B cells. Analysis of virus transcription during the establishment of latency revealed that virus gene expression in B cells was restricted and dependent on the differentiation stage of the B cell. Notably, transcription of ORF73 was detected in germinal center B cells, a finding in agreement with the predicted latent genome maintenance function of ORF73 in dividing cells. At late times after infection, virus DNA could only be detected in newly formed and germinal center B cells, which suggests that B cells play a critical role in facilitating life-long latency.

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Age-associated B cells are critical for the control of latent gammaherpesvirus 68 infection

10.1101/2021.12.29.474434 ◽

2021 ◽

Author(s):

Isobel C. Mouat ◽

Iryna Shanina ◽

Marc S. Horwitz

Keyword(s):

B Cells ◽

Viral Infections ◽

Acute Infection ◽

Effective Control ◽

Murine Gammaherpesvirus ◽

Barr Virus ◽

Gamma Herpesvirus ◽

Epstein Barr ◽

Gammaherpesvirus 68 ◽

Murine Gammaherpesvirus 68

Age-associated B cells (ABCs; CD19+CD11c+T-bet+) are increased during an array of viral infections, though their role during viral latency is unexplored. Here, we use murine gammaherpesvirus 68 (γHV68), a homolog of Epstein-Barr virus that latently infects B cells, to demonstrate that ABCs are necessary for the effective control of gamma-herpesvirus latency. We observe that ABCs expand in the spleen during acute infection and persist at least 150 days post-infection. During acute and latent infection ABCs secrete IFNγ and TNF. Using a strain of γHV68 that is cleared following acute infection, we show that ABCs persist in the absence of latent virus, though they secrete less IFNγ and TNF. With a fluorescent virus we demonstrate that ABCs are infected with γHV68 at similar rates to other mature B cells. We find that mice without ABCs display defects in anti-viral IgG2a/c antibodies and are less able to maintain γHV68 latency when challenged with heterologous infection. Together, these results indicate that ABCs are a persistent subset during latent viral infection that controls γHV68 reactivation from latency.

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