Regulation of Epstein-Barr virus gene expression via CRISPR-mediated targeting of DNA methylation enzymes

Abstract

Epstein-Barr virus (EBV) can establish an oncogenic presence in cells to cause transformation. Globally, EBV drives a number of human cancers, including Natural Killer/T cell Lymphoma (NKTL). NKTL is an aggressive malignancy thought to express a limited number of EBV proteins, namely EBV nuclear antigen 1 (EBNA1) and latent membrane proteins (LMPs) 1 and 2. However, apart from these, NKTLs also express a wide range of EBV transcripts that do not appear to be translated. LMP1 transcript levels have been positively correlated whereas late lytic gene levels are negatively correlated with patient survival. The regulation of LMP1 and lytic gene expression is not fully understood. Therefore, in this study, the role of DNA methylation enzymes, DNMT1, DNMT3A and UHRF1, in regulating LMP1, BZLF1 and BRRF2 were clarified. Additionally, LMP1 was investigated for its regulation on these genes. Through gene editing and expression analyses, DNMT1, UHRF1, DNMT3A downregulate LMP1 expression. Intriguingly, while DNMT3A downregulated lytic genes, DNMT1 and UHRF1 upregulated BZLF1 and BRRF2, contrary to current understanding. Furthermore, LMP1 influences both latency and lytic gene expression. Thus, DNMT1, DNMT3A, UHRF1 and LMP1 could be potential targets in increasing immunogenicity in NKTL in order to enhance immune clearance of tumour mass.