Dynamics of Viral and Host 3D Genome Structure upon Infection

Abstract

Eukaryotic chromatin is highly organized in the 3D nuclear space and dynamically regulated in response to environmental stimuli. This genomic organization is arranged in a hierarchical fashion to support various cellular functions, including transcriptional regulation of gene expression. Like other host cellular mechanisms, viral pathogens utilize and modulate host chromatin architecture and its regulatory machinery to control features of their life cycle, such as lytic versus latent status. Combined with previous research focusing on individual loci, recent global genomic studies employing conformational assays coupled with high-throughput sequencing technology have informed models for host and, in some cases, viral 3D chromosomal structure re-organization during infection and the contribution of these alterations to virus-mediated diseases. Here, we review recent discoveries and progress in host and viral chromatin structural dynamics during infection, focusing on a subset of DNA (human herpesviruses and HPV) as well as RNA (HIV, influenza virus and SARS-CoV-2) viruses. An understanding of how host and viral genomic structure affect gene expression in both contexts and ultimately viral pathogenesis can facilitate the development of novel therapeutic strategies.

Keywords: Chromatin structure; EBV; HIV; HPV; KSHV; SARS-CoV-2; influenza virus.

Fig. 1. Schematic representation of assays for studying chromosome architecture.

For 3C-based methods, nuclei are first treated with appropriate fixatives (i.e., formaldehyde, DSG, etc.). In GAM, cryosections are cut from paraformaldehydefixed and sucrose-embedded samples. In 3C, 4C and 5C, fixed nuclei are treated with restriction enzymes, ligated, and the ligation frequency is measured by PCR or NGS. For Hi-C, Plac-seq, and cHi-C, chromosomal DNA is digested by restriction enzymes while micro-C uses MNase for finer resolution. The digested DNA ends are repaired with biotin-labeled nucleotides followed by blunt-end ligation. The ligated biotin-labeled contacts are sheared and purified with streptavidin beads prior to NGS sequencing. In Plac-seq and cHi-C, antibody pull-down or RNA oligo-mediated DNA pull-down is performed for target enrichment, respectively. In the SPRITE method, crosslinked chromatin is fragmented by sonication, each interacting complex is uniquely tagged by multiple rounds of split-pool barcoding, and the final material is sequenced. In the GAM method, the DNA contents from cryosections are extracted, fragmented, and sequenced. Appropriate computational analysis of the sequencing data from each approach is necessary to detect physical interactions between genomic loci.

Fig. 2. Models of viral and host regulation related to chromatin conformation.

(A) In herpesviruses, host architectural proteins (i.e., CTCF and cohesin) bind the viral episome and influence viral gene expression depending on the state of the virus (i.e., latent vs. lytic) via conformational changes to the episome. (B) Viral gene products can modulate host gene expression by targeting host enhancers or promoters (i.e., EBNAs and LANA). (C) The herpesvirus episome is associated with a repressed compartment in the host genome during latency. Upon reactivation, the interaction between the episome and euchromatin may increase. This viral-host chromosomal interplay could modulate both viral and host gene expression. (D) CTCF binding at the E2 promoter acts as a suppressor of viral oncogene E6/E7 by modulating HPV chromatin architecture. Disruption of CTCF binding causes transcriptional activation of E6/E7 in the HPV episome. (E) Viral integration into the host genome weakens looping between the E2 and E6/E7 loci, which also results in activation of the latter. The integrated viral sequence can hijack host enhancers for transcription of viral gene products or can act as an enhancer itself to stimulate neighboring cancer-associated genes. (F) Upon IAV infection, the viral protein NS1 inhibits the termination of transcription in a subset of highly transcribed genes. This inhibition leads to RNA Pol II-dependent dissociation of CTCF from its binding sites and disruption of chromatin loops (left). IAV protein NP competitively binds the host protein Suv4-20h2 to release Suv4-20h2- bound cohesin. Liberated cohesin is then recruited to the HoxC8-HoxC6 locus where it induces gene expression by forming an active chromatin loop (right).