Author(s): Washington SD, Singh P, Johns RN, Edwards TG, Mariani M, Frietze S, Bloom DC, Neumann DM. The CCCTC binding factor, CTRL2, modulates heterochromatin deposition and the establishment of HSV-1 latency in vivo. J Virol. 2019 Apr 17. pii: JVI.00415-19. doi: 10.1128/JVI.00415-19. [Epub ahead of print]
Journal: Journal Of Virology, Apr 2019
The cellular insulator protein CTCF plays a role in HSV-1 latency through the establishment and regulation of chromatin boundaries. We previously found that the CTRL2 regulatory element downstream from the LAT enhancer was bound by CTCF during latency and underwent CTCF eviction at early times post-reactivation in mice latent with 17syn+. We also showed that CTRL2 was a functional enhancer-blocking insulator in both epithelial and neuronal cell lines. We hypothesized that CTRL2 played a direct role in silencing lytic gene expression during the establishment of HSV-1 latency. To test this hypothesis, we used a recombinant virus with a 135-bp deletion spanning only the core CTRL2 insulator domain (ΔCTRL2) in the 17syn+ background. Deletion of CTRL2 resulted in restricted viral replication in epithelial cells, but not neuronal cells. Following ocular infection, mouse survival decreased in the ΔCTRL2 cohort and we found a significant decrease in the number of viral genomes in mouse trigeminal ganglia (TG) infected with ΔCTRL2, indicating that the CTRL2 insulator was required for the efficient establishment of latency. Immediate early (IE) gene expression significantly increased in the ganglia infected with ΔCTRL2 by 31 days post-infection, relative to 17syn+, indicating that deletion of the CTRL2 insulator disrupted the organization of chromatin domains during HSV-1 latency. Finally, ChIP-seq analyses of TG from ΔCTRL2 infected mice confirmed that the distribution of the repressive H3K27me3 mark on the ΔCTRL2 recombinant genomes was altered compared to the wild-type, indicating that CTRL2 site modulates the repression of IE genes during latency.IMPORTANCE It is becoming increasingly clear that chromatin insulators play a key role in the transcriptional control of DNA viruses. The gamma herpesviruses EBV and KSHV utilize chromatin insulators to order protein recruitment and dictate the formation of 3-dimensional DNA loops that spatially control transcription and latency. The contribution of chromatin insulators in alpha herpesvirus transcriptional control is less understood. The work presented here begins to bridge that gap in knowledge by showing how one insulator site in HSV-1 modulates lytic gene transcription and heterochromatin deposition as the HSV-1 genome establishes latency.