KSHV encoded ORF59 modulates histone arginine methylation of the viral genome to promote viral reactivation

Abstract

Kaposi’s sarcoma associated herpesvirus (KSHV) persists in a highly-ordered chromatin structure inside latently infected cells with the majority of the viral genome having repressive marks. However, upon reactivation the viral chromatin landscape changes into ‘open’ chromatin through the involvement of lysine demethylases and methyltransferases. Besides methylation of lysine residues of histone H3, arginine methylation of histone H4 plays an important role in controlling the compactness of the chromatin. Symmetric methylation of histone H4 at arginine 3 (H4R3me2s) negatively affects the methylation of histone H3 at lysine 4 (H3K4me3), an active epigenetic mark deposited on the viral chromatin during reactivation. We identified a novel binding partner to KSHV viral DNA processivity factor, ORF59-a protein arginine methyl transferase 5 (PRMT5). PRMT5 is an arginine methyltransferase that dimethylates arginine 3 (R3) of histone H4 in a symmetric manner, one hallmark of condensed chromatin. Our ChIP-seq data of symmetrically methylated H4 arginine 3 showed a significant decrease in H4R3me2s on the viral genome of reactivated cells as compared to the latent cells. Reduction in arginine methylation correlated with the binding of ORF59 on the viral chromatin and disruption of PRMT5 from its adapter protein, COPR5 (cooperator of PRMT5). Binding of PRMT5 through COPR5 is important for symmetric methylation of H4R3 and the expression of ORF59 competitively reduces the association of PRMT5 with COPR5, leading to a reduction in PRMT5 mediated arginine methylation. This ultimately resulted in a reduced level of symmetrically methylated H4R3 and increased levels of H3K4me3 marks, contributing to the formation of an open chromatin for transcription and DNA replication. Depletion of PRMT5 levels led to a decrease in symmetric methylation and increase in viral gene transcription confirming the role of PRMT5 in viral reactivation. In conclusion, ORF59 modulates histone-modifying enzymes to alter the chromatin structure during lytic reactivation.

Author summary

Kaposi’s sarcoma-associated herpesvirus (KSHV) must carefully regulate both phases of its lifecycle in order to persist and proliferate effectively in the infected cells. In this study, we show the importance of dynamic epigenetic modifications on the viral chromatin in dictating whether KSHV displays the latent or lytic phase of its life cycle. Various chromatin-modifying enzymes are responsible for adding activating or repressive ‘marks’ on chromatin, one of these is a PRMT5 (protein arginine methyltransferase 5), which symmetrically dimethylates arginine 3 of histone H4 (H4R3me2s) and associates with condensed chromatin leading to restricted gene expression. An early lytic protein of KSHV, ORF59 associates with PRMT5 to disrupt its binding with the chromatin leading to a loss of repressive, H4R3me2s mark and corresponding gain of activating H3K4me3 during lytic reactivation.