Abstract
Interferon regulatory factor 8 (IRF8), also known as interferon consensus sequence-binding protein (ICSBP), is a transcription factor of the IRF family. IRF8 plays a key role in normal B cell differentiation, a cellular process that is intrinsically associated with Epstein-Barr virus (EBV) reactivation. However, whether IRF8 regulates EBV lytic replication remains unknown. In this study, we utilized a CRISPR/Cas9 genomic editing approach to deplete IRF8 and found that IRF8 depletion dramatically inhibits the reactivation of EBV upon lytic induction. We demonstrated that IRF8 depletion suppresses the expression of a group of genes involved in apoptosis and thus inhibits apoptosis induction upon lytic induction by B cell receptor (BCR) stimulation or chemical induction. The protein levels of caspase-1, caspase-3 and caspase-8 all dramatically decreased in IRF8-depleted cells, which led to reduced caspase activation and the stabilization of KAP1, PAX5 and DNMT3A upon BCR stimulation. Interestingly, caspase inhibition blocked the degradation of KAP1, PAX5 and DNMT3A, suppressed EBV lytic gene expression and viral DNA replication upon lytic induction, suggesting that the reduced caspase expression in IRF8-depleted cells contributes to the suppression of EBV lytic replication. We further demonstrated that IRF8 directly regulates CASP1 (caspase-1) gene expression through targeting its gene promoter and knockdown of caspase-1 abrogates EBV reactivation upon lytic induction, partially through the stabilization of KAP1. Together our study suggested that, by modulating the activation of caspases and the subsequent cleavage of KAP1 upon lytic induction, IRF8 plays a critical role in EBV lytic reactivation.
Highlights
Epstein-Barr virus (EBV), a ubiquitous human gammaherpesvirus, is associated with malignant diseases, including Burkitt’s lymphoma, Hodgkin’s lymphoma, nasopharyngeal carcinoma, and NK/T cell lymphoma [1]
We demonstrate that interferon regulatory factor 8 (IRF8) is required for EBV lytic replication
Interferon regulatory factor 8 (IRF8) directly regulates caspase-1 expression and caspase activation upon B cell receptor (BCR) stimulation and chemical induction, which leads to the cleavage and de-stabilization of several host factors suppressing lytic replication, including KAP1
Summary
Epstein-Barr virus (EBV), a ubiquitous human gammaherpesvirus, is associated with malignant diseases, including Burkitt’s lymphoma, Hodgkin’s lymphoma, nasopharyngeal carcinoma, and NK/T cell lymphoma [1]. The genome of EBV is approximately 170 kb in length and encodes more than 80 genes. EBV infects both B lymphocytes and some epithelial cells and the life cycle of EBV is divided into latent or lytic phases. EBV expresses all lytic genes and progeny virus particles are packaged and released from the cell [2]. The reactivation of EBV from latent to lytic phase can be triggered by expression of two viral immediate-early gene products, ZTA ( called BZLF1 or Z) and RTA ( known as BRLF1 or R). B cell receptor (BCR) activation is a philologically relevant stimulus for triggering EBV reactivation from latency since this occurs in tumor cell lines and in freshly isolated B cells from patients [17,18]
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