Integrated Analysis of Chromatin Accessibility and Transcriptional Dynamics of the Viral Genome Following Pseudorabies Virus Infection

Pseudorabies virus (PRV) is an important α-herpesvirus. However, the chromatin state and transcriptional regulatory mechanisms of its genome upon entry into host cell nuclei remain poorly understood. In this study, we employed the Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) and RNA sequencing (RNA-seq) to analyze chromatin accessibility and the transcriptome of the PRV genome in PK15 cells at multiple time points post-infection. ATAC-seq analysis revealed that the proportion of viral reads increased progressively over time, from 0.01% at 4 h to approximately 1.09% and 13% at 8 h and 12 h, respectively. A total of 112 ATAC peaks were identified, distributed across the PRV genome without apparent low-accessibility regions. Fragment length analysis demonstrated that the PRV genome does not adopt the regularly phased nucleosome organization typical of the host genome. RNA-seq analysis detected 66 expressed PRV genes, which were classified into four kinetic expression clusters, corresponding to immediate-early/early (IE/E), early (E), early-to-late transitional (E-L), and late (L) gene groups. Virus–host correlation analysis revealed significant associations between PRV gene expression and multiple host inflammation-related genes. Integrated multi-omics analysis further confirmed the concordance between chromatin accessibility dynamics and transcriptional activity changes. Collectively, these findings indicate that the PRV genome maintains a predominantly open and accessible chromatin state throughout lytic infection, providing new insights into the epigenetic regulatory mechanisms of α-herpesviruses.