RESUMO
Epstein-Barr virus (EBV) has been implicated in the pathogenesis of human malignancies, but its contribution to tumorigenesis is not well understood. EBV carriage is associated with increased genomic instability in Burkitt's lymphoma, suggesting that viral products may induce this tumor phenotype. Using a panel of transfected sublines of the B-lymphoma line BJAB expressing the viral genes associated with latent infection, we show that the EBV nuclear antigens, EBNA-1 and EBNA-3C, and the latent membrane protein 1, LMP-1, independently promote genomic instability, as detected by nonclonal chromosomal aberrations, DNA breaks and phosphorylation of histone H2AX. EBNA-1 promotes the generation of DNA damage by inducing reactive oxygen species (ROS), whereas DNA repair is inhibited in LMP-1-expressing cells through downregulation of the DNA damage-sensing kinase, ataxia telangiectasia mutated (ATM), reduction of phosphorylation of its downstream targets Chk2 and inactivation of the G(2) checkpoint. EBNA-3C enhances the propagation of damaged DNA through inactivation of the mitotic spindle checkpoint and transcriptional downregulation of BubR1. Thus, multiple cellular functions involved in the maintenance of genome integrity seem to be independently targeted by EBV, pointing to the induction of genomic instability as a critical event in viral oncogenesis.
Assuntos
Antígenos Virais/genética , Linfoma de Burkitt/genética , Linfoma de Burkitt/virologia , Dano ao DNA , Reparo do DNA , Instabilidade Genômica , Herpesvirus Humano 4/fisiologia , Proteínas Mutadas de Ataxia Telangiectasia , Linfoma de Burkitt/metabolismo , Ciclo Celular , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/metabolismo , Regulação para Baixo , Antígenos Nucleares do Vírus Epstein-Barr/genética , Herpesvirus Humano 4/genética , Humanos , Proteínas Serina-Treonina Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Fuso Acromático , Transfecção , Proteínas Supressoras de Tumor/metabolismo , Proteínas da Matriz Viral/genética , Latência ViralRESUMO
Epstein-Barr virus (EBV) has been implicated in the pathogenesis of human malignancies but the mechanisms of oncogenesis remain largely unknown. Genomic instability and chromosomal aberrations are hallmarks of malignant transformation. We report that EBV carriage promotes genomic instability in Burkitt's lymphoma (BL). Cytogenetic analysis of EBV- and EBV+ BL lines and their sublines derived by EBV conversion or spontaneous loss of the viral genome revealed a significant increase in dicentric chromosomes, chromosome fragments and chromatid gaps in EBV-carrying cells. Expression of EBV latency I was sufficient for this effect, whereas a stronger effect was observed in cells expressing latency III. Telomere analysis by fluorescent in situ hybridization revealed an overall increase of telomere size and prevalence of telomere fusion and double strand-break fusion in dicentric chromosomes from EBV+ cells. Phosphorylated H2AX, a reporter of DNA damage and ongoing repair, was increased in virus-carrying cells in the absence of exogenous stimuli, whereas efficient activation of DNA repair was observed in both EBV+ and EBV- cells following treatment with etoposide. These findings point to induction of telomere dysfunction and DNA damage as important mechanisms for EBV oncogenesis.
