RESUMO
Humans are infected with two distinct strains (Type 1 (T1) and Type 2 (T2)) of Epstein-Barr virus (EBV) that differ substantially in their EBNA2 and EBNA 3A/B/C latency genes and the ability to transform B cells in vitro. While most T1 EBV strains contain the "prototype" form of the BZLF1 immediate-early promoter ("Zp-P"), all T2 strains contain the "Zp-V3" variant, which contains an NFAT binding motif and is activated much more strongly by B-cell receptor signalling. Whether B cells infected with T2 EBV are more lytic than cells infected with T1 EBV is unknown. Here we show that B cells infected with T2 EBV strains (AG876 and BL5) have much more lytic protein expression compared to B cells infected with T1 EBV strains (M81, Akata, and Mutu) in both a cord blood-humanized (CBH) mouse model and EBV-transformed lymphoblastoid cell lines (LCLs). Although T2 LCLs grow more slowly than T1 LCLs, both EBV types induce B-cell lymphomas in CBH mice. T1 EBV strains (M81 and Akata) containing Zp-V3 are less lytic than T2 EBV strains, suggesting that Zp-V3 is not sufficient to confer a lytic phenotype. Instead, we find that T2 LCLs express much higher levels of activated NFATc1 and NFATc2, and that cyclosporine (an NFAT inhibitor) and knockdown of NFATc2 attenuate constitutive lytic infection in T2 LCLs. Both NFATc1 and NFATc2 induce lytic EBV gene expression when combined with activated CAMKIV (which is activated by calcium signaling and activates MEF2D) in Burkitt Akata cells. Together, these results suggest that B cells infected with T2 EBV are more lytic due to increased activity of the cellular NFATc1/c2 transcription factors in addition to the universal presence of the Zp-V3 form of BZLF1 promoter.
Assuntos
Linfócitos B/metabolismo , Fatores de Transcrição NFATC/genética , Animais , Linfócitos B/virologia , Linhagem Celular , Proteínas de Ligação a DNA/metabolismo , Infecções por Vírus Epstein-Barr/genética , Antígenos Nucleares do Vírus Epstein-Barr , Expressão Gênica/genética , Regulação Viral da Expressão Gênica/genética , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/metabolismo , Herpesvirus Humano 4/patogenicidade , Humanos , Camundongos , Regiões Promotoras Genéticas/genética , Transativadores/genética , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Proteínas Virais/metabolismo , Ativação Viral , Latência ViralRESUMO
Using GFP to mark recent thymic emigrants (RTEs) in mice carrying a GFP transgene driven by the recombination-activating gene 2 promoter, we demonstrate that RTEs are readily detectable even in 2-year-old mice, despite the fact that the proportion of the peripheral T cell pool comprised of RTEs declines with age. Although the number of RTEs decreases after reaching a peak at 6 weeks of age, thymic output as a function of thymic size is surprisingly age-independent. The CD4:CD8 ratio of RTEs declines with age, partly because of a striking decrease in steady-state proliferation of CD4+ RTEs in older mice. RTEs in aged mice undergo phenotypic maturation in the lymphoid periphery with delayed kinetics compared with young mice. RTEs from aged mice secrete less IL-2, proliferate less well, and achieve only weak expression of early-activation markers compared with more mature naïve peripheral T cells from the same mice. The proportion of GFP- cells in the CD4+ and CD8+ thymic compartments increases with age, partly as a result of leakiness in the aged thymus, allowing reentry of naïve peripheral T cells.
Assuntos
Envelhecimento/imunologia , Timo/imunologia , Animais , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD8-Positivos/citologia , Movimento Celular , Proliferação de Células , Células Cultivadas , Imunidade Inata , Cinética , Contagem de Linfócitos , Camundongos , Tamanho do Órgão , Fenótipo , Timo/citologiaRESUMO
Mouse CD4(+)Vbeta5(+) T cells recognize a peripherally expressed superantigen encoded by an endogenous retrovirus. Ag encounter tolerizes the mature CD4 T cell compartment, either by deletion of autoreactive cells or by TCR revision. This latter process is driven by TCRbeta rearrangement through RAG activity and results in the rescue of cells expressing novel TCRs that no longer recognize the tolerogen. Consistent with the notion that revising T cells represent a distinct peripheral T cell population, we now show that these lymphocyte blasts express a hybrid effector/memory phenotype and are not undergoing cell division. A population of revising T cells is CD40(+), expresses the germinal center (GC) marker CXCR5, and is Vbeta5(low)Thy-1(low). Histology reveals that, consistent with their surface Ag phenotype, T cells undergoing TCR revision are enriched in splenic GCs. These data demonstrate that TCR revision is a multistep tolerance pathway supported by the unique microenvironment provided by GCs.