Epstein-Barr virus (EBV) infects >95% of adults and contributes to several human cancers. EBV can remain latent where viral lytic genes are silenced, precluding the use of antiviral agents such as ganciclovir. Little is known about the host factors involved in EBV latency. Here we performed a human genome-wide CRISPR-Cas9 screen in Burkitt lymphoma B cells, which identified lysine-specific histone demethylase 1 (LSD1) and its corepressors REST corepressor 1 (CoREST) and zinc finger protein 217 (ZNF217) as critical for EBV latency. Gene knockout or LSD1 inhibition triggered EBV reactivation, and the latter sensitized cells to ganciclovir cytotoxicity, including in murine tumour xenografts. Mechanistically, ZNF217 recruits LSD1 and CoREST to form a complex that binds a specific DNA motif associated with regions implicated in EBV reactivation. It removes histone 3 lysine 4 (H3K4) methylation marks and restricts host DNA looping. Alternatively, the H3K4 lysine methyltransferase 2D supports EBV lytic reactivation. Our results highlight H3K4 methylation as a major EBV lytic switch regulator and therapeutic target.
