UV irradiation of HSV-1 virions induces the differential recruitment of host intrinsic and innate immune regulators to infecting viral genomes

Recognition of virus-derived nucleic acids is a key process in the activation of immune defences to viral infection. However, the spatiotemporal recruitment of host immune regulators to infecting viral DNA (vDNA) genomes remains poorly defined. Here we utilize 5-Ethynyl-2’-deoxycytidine (EdC) nucleotide labelling of WT or UV-irradiated HSV-1 genomes in combination with click chemistry and high-resolution confocal microscopy imaging to investigate the recruitment and proximity of key intracellular immune regulators to infecting vDNA during HSV-1 infection. We report that UV irradiation of HSV-1 virions restricts vDNA decompaction upon delivery to infected cells. UV treatment reduced the frequency in PML-NB recruitment to nuclear genomes relative to non-irradiated vDNA. These data demonstrate an important role of genome decompaction in the recruitment of intrinsic host factors to nuclear infecting vDNA. Additionally, UV-irradiated HSV-1 genomes favoured a premature cytosolic deposition phenotype, due to ineffective association with microtubules as shown by nocodazole treatment. The cytosolic UV irradiated vDNA showed enhanced association with both cGAS and STING. Interestingly, high-resolution Airyscan imaging identified a consistent unreported 3D interaction between both cGAS and STING, the strength of association of which was enhanced in the context of UV infection. This enhanced recruitment correlated with elevated levels of IFN-β transcription relative to non-irradiated vDNA, despite the presence of non-irradiated cytosolic genomes. Our data demonstrates that UV irradiation of herpesviruses, a well-established procedure for virus inactivation, has multiple effects on genome structure and sub-cellular localisation that differentially influence the spatiotemporal recruitment of intrinsic and innate host immune regulators to infecting genomes.