CTCF Binding Sites in the Herpes Simplex Virus 1 Genome Display Site-Specific CTCF Occupation, Protein Recruitment, and Insulator Function

Abstract

There are seven conserved CTCF binding domains in the herpes simplex virus 1 (HSV-1) genome. These binding sites individually flank the latency-associated transcript (LAT) and the immediate early (IE) gene regions, suggesting that CTCF insulators differentially control transcriptional domains in HSV-1 latency. In this work, we show that two CTCF binding motifs in HSV-1 display enhancer blocking in a cell-type-specific manner. We found that CTCF binding to the latent HSV-1 genome was LAT dependent and that the quantity of bound CTCF was site specific. Following reactivation, CTCF eviction was dynamic, suggesting that each CTCF site was independently regulated. We explored whether CTCF sites recruit the polycomb-repressive complex 2 (PRC2) to establish repressive domains through a CTCF-Suz12 interaction and found that Suz12 colocalized to the CTCF insulators flanking the ICP0 and ICP4 regions and, conversely, was removed at early times postreactivation. Collectively, these data support the idea that CTCF sites in HSV-1 are independently regulated and may contribute to lytic-latent HSV-1 control in a site-specific manner. The role of chromatin insulators in DNA viruses is an area of interest. It has been shown in several beta- and gammaherpesviruses that insulators likely control the lytic transcriptional profile through protein recruitment and through the formation of three-dimensional (3D) chromatin loops. The ability of insulators to regulate alphaherpesviruses has been understudied to date. The alphaherpesvirus HSV-1 has seven conserved insulator binding motifs that flank regions of the genome known to contribute to the establishment of latency. Our work presented here contributes to the understanding of how insulators control transcription of HSV-1.

Strength in diversity: Understanding the pathways to herpes simplex virus reactivation

“…Further investigation of how histone phosphorylation influences transcription of the viral genome will uncover mechanisms regulating HSV gene expression and provide insights on the control of cellular gene expression. Furthermore, there is clear evidence that the insulator protein CTCF and components of the polycomb repressive complex (Washington et al, 2018b) are evicted from sites on the viral genome following different reactivation stimuli, but the cause-and-effect relationships and mechanisms of these processes are less clear.…”

“…This apparent demarcation in the nature of the chromatin likely arises due to binding sites for the cellular insulator protein CCCTC-binding factor (CTCF) on the viral genome. Interestingly, CTCF eviction coincides with reactivation (Ertel et al, 2012; Washington et al, 2018b), and depletion of CTCF in vivo promotes reactivation (Washington et al, 2018a). Furthermore, one important binding site of CTCF, known as CTRL2, lies downstream of the LAT enhancer and separates it from the nearby lytic ICP0 gene (Amelio et al, 2006b).…”

“…It remains unclear if JNK plays a similar mechanistic role following explant of latently infected ganglia, and, as a whole, very little is known about the mechanisms linking cell signaling pathways and changes in viral chromatin following axotomy. However, a number of important observations about the viral chromatin have been made using the explant model, including changes in histone post-translational modification and changes in CTCF binding (Ertel et al, 2012; Washington et al, 2018b). Most notably, it has been shown that, following axotomy, there is a transient increase in the enrichment of euchromatic histone markers (acetylated H3K9/K14) at IE gene promoters (Amelio et al, 2006a).…”

“…This is in contrast to reactivation in intact ganglia, in which VP16 is required (Thompson et al, 2009), again indicating that the mechanisms of reactivation likely differ depending on the nature of the trigger. Lastly, it has been shown that CTCF domains positioned around IE (ICP0 and ICP4) and LAT gene regions lose CTCF occupancy following explant-induced reactivation (Ertel et al, 2012; Washington et al, 2018b), suggesting that disruption of CTCF-mediated chromatin domains may contribute to reactivation.…”

Small Alphaherpesvirus Latency-Associated Promoters Drive Efficient and Long-Term Transgene Expression in the Central Nervous System

“…The recruitment of specific TF from different host cells can modulate transgene transcription by the same mechanisms regulating resistance to inactivation during latency. Insulator elements like the CTCF-binding factor are independently regulated 67 and can protect promoter regions from repression by heterochromatin, maintaining long-lasting transcription 64 .…”