Centromeric DNA amplification triggered by viral proteins activates nuclear cGAS

The cGAS-cGAMP-STING pathway is crucial for antiviral immunity. While cytosolic cGAS detects viral DNA, most DNA viruses shield their genome and invade the nucleus, where chromatin restricts cGAS activation. How viruses may activate nuclear cGAS is not well understood. Here, we show that several herpesvirus proteins trigger nuclear cGAS activation by perturbing centromeres, where cGAS is enriched. The herpes simplex virus type 1 (HSV-1) ubiquitin ligase ICP0, which degrades centromeric proteins, promotes centromeric DNA amplification through the translesion DNA synthesis (TLS) pathway in quiescent monocyte-derived cells, thereby activating nuclear cGAS. During infection, HSV-1 evades this detection by also expressing UL36USP, a suppressor of TLS. Similarly to ICP0, the cytomegalovirus IE1 protein causes centromeric DNA amplification and cGAS activation. We define this mechanism as Viral-Induced Centromeric DNA Amplification and Recognition (VICAR), uncovering a non-mitotic, immune-activating role of centromeres. Overall design: CD14+ Human Peripheral Blood Mononuclear cells (PBMCs) were isolated from healthy human donors and selected for using anti-CD14 magnetic beads. They are then differentiated into Monocyte-derived Dendritic Cells (MDDCs) by treatment with GM-CSF and IL-4 and transduced with either control lentivector or lentivector expressing wild-type ICP0 or mutant ICP0-FxE protein. After 4 days of differentiation and transduction, cells are collected for chromatin accessibility profiling by ATAC-seq. Details are described in the Methods section of the paper.