KSHV Topologically Associating Domains in Latent and Reactivated Viral Chromatin

Eukaryotic genomes are structurally organized via the formation of multiple loops that create gene expression regulatory units called topologically associating domains (TADs). Here we revealed the KSHV TAD structure at 500 base pair resolution and constructed a 3D KSHV genomic structural model with 2kb binning. The latent KSHV genome formed very similar genomic architectures in three different naturally infected PEL cell lines and in an experimentally infected epithelial cell line. The majority of the TAD boundaries were occupied by CTCF and SMC1, and the KSHV transactivator was recruited to these sites during reactivation. Triggering KSHV gene expression decreased pre-wired genomic loops within the regulatory unit, while contacts extending outside of regulatory borders increased, leading to formation of a larger regulatory unit with a shift from repressive to active compartments (B to A). The 3D genomic structural model proposes that the immediate-early promoter region is localized on the periphery of the 3D viral genome during latency, while highly inducible non-coding RNA regions moved toward the inner space of the structure, resembling the configuration of a “bird cage” during reactivation. The compartment-like properties of viral episomal chromatin structure and its reorganization during the transition from latency may help coordinate viral gene transcription. Importance The 3D architecture of chromatin allows for efficient arrangement, expression, and replication of genetic material. The genomes of all organisms studied to date have been found to be organized through some form of tiered domain structures. However, the architectural framework of the genomes of large double-stranded DNA viruses such as the herpesvirus family has not been reported. Prior studies with Kaposi’s sarcoma-associated herpesvirus (KSHV) have indicated that the viral chromatin shares many biological properties exhibited by the host cell genome, essentially behaving as a mini human chromosome. Thus, we hypothesized that the KSHV genome may be organized in a similar manner. In this report, we describe the domain structure of the latent and lytic KSHV genome at 500 base pair resolution and present a 3D genomic structural model for KSHV under each condition. These results add new insights into the complex regulation of the viral lifecycle.