The H4K20-mono-methyltransferase SETD8 promotes global accessibility of infecting herpes simplex virus genomes.

Epigenetic modulation of herpes simplex virus (HSV) immediate early (IE) genes is a critical parameter governing lytic infection, latency, and viral reactivation. Multiple factors, including epigenetic complexes associated with the cellular transcriptional coactivator HCF-1, modulate the state of HSV-1 chromatin. Although some aspects of HSV-1 chromatin biology have been elucidated, many epigenetic factors controlling HSV-1 chromatin accessibility and transcriptional regulation remain unknown. To identify novel epigenetic regulators of the initial stage of HSV-1 infection, an epigenetic chemical probe library was screened for molecules that impacted viral IE gene expression. This screen identified several epigenetic inhibitors that modulated IE expression. Notably, UNC0379, an inhibitor of the histone H4K20 mono-methyltransferase SETD8, potently suppressed HSV-1 IE gene transcription during lytic infection. UNC0379 treatment also repressed HSV-1 reactivation in a mouse ganglia explant model, and topical application suppressed primary ocular infection in vivo. Importantly, SETD8 inhibition resulted in enhanced heterochromatin suppression and reduced viral genome accessibility, as evident by increased repressive H3K9me3 marks and decreased ATAC-seq signal across the HSV-1 genome. Reduced IE transcription was concomitant with disruption of H4K20me1 deposition and impaired recruitment of HCF-1 and RNA polymerase II (RNAPII). These findings are consistent with the established roles of SETD8 in promoting chromatin accessibility through H4K20me1 deposition and in regulating transcriptional elongation, suggesting that SETD8 may facilitate multiple steps in HSV-1 IE gene expression. Overall, these results identified SETD8 as an essential epigenetic modulator of HSV-1 chromatin accessibility and represent a promising novel antiviral therapeutic target. IMPORTANCE: Herpes simplex virus is a highly prevalent human pathogen, with persistence of the viral genome in sensory neurons serving as a reservoir for recurrent disease and viral shedding. Epigenetic pharmaceuticals have proven to be valuable tools in elucidating the chromatin landscape of the viral genome and its impact on viral gene expression. With the use of a chemical library screen, the histone methyltransferase SETD8 was identified as a key factor required for promoting accessibility of the herpesvirus genome to transcriptional regulators. Importantly, pharmacological inhibition of SETD8 suppressed herpes simplex virus lytic infection, reduced viral reactivation in sensory neurons, and when applied topically, inhibited primary ocular infection of mice. Collectively, these findings establish SETD8 as a critical regulator of viral gene expression during lytic infection and the initiation of reactivation from latency. These results highlight SETD8 as a potential novel target for antiviral therapy.