Abstract
The three-dimensional structure of chromatin organized by genomic loops facilitates RNA polymerase II access to distal promoters. The Kaposi’s sarcoma-associated herpesvirus (KSHV) lytic transcriptional program is initiated by a single viral transactivator, K-Rta. Here we report the KSHV genomic structure and its relationship with K-Rta recruitment sites using Capture Hi–C analyses. High-resolution 3D viral genomic maps identify a number of direct physical, long-range, and dynamic genomic interactions. Mutant KSHV chromosomes harboring point mutations in the K-Rta responsive elements (RE) significantly attenuate not only the directly proximate downstream gene, but also distal gene expression in a domain-specific manner. Genomic loops increase in the presence of K-Rta, while abrogation of K-Rta binding impairs the formation of inducible genomic loops, decreases the expression of genes networked through the looping, and diminishes KSHV replication. Our study demonstrates that genomic architectural dynamics plays an essential role in herpesvirus gene expression.
Highlights
The three-dimensional structure of chromatin organized by genomic loops facilitates RNA polymerase II access to distal promoters
With the results reported here, we show that Kaposi’s sarcoma-associated herpesvirus (KSHV) has evolved spatiotemporal gene regulatory mechanisms to assemble viral genomic domains and recruit RNA polymerase II, which results in effective viral gene expression from uniquely structured viral episomes
We speculated that robust gene activation by K-Rta associates with a unique structure formed by the KSHV genome, which is prefabricated with an architecture designed for K-Rta recruitment
Summary
K-Rta binding sites on the KSHV genome. The KSHV immediate early gene, K-Rta, is both necessary and sufficient to switch latent KSHV into the lytic infection cycle[27,29,30]. We speculated that robust gene activation by K-Rta associates with a unique structure formed by the KSHV genome, which is prefabricated with an architecture designed for K-Rta recruitment To examine this hypothesis, the K-Rta binding sites in the KSHV genome were precisely mapped with ChIP-seq analysis in the context of the BCBL-1 cell line model. The results showed multiple recruitment sites of K-Rta, including the promoter regions for ORF6, 7, 8, K2, 70, PAN RNA, 31, 45, KRta, K-bZIP, ORF57, K12, 73, and 75, in addition to both ori-Lyt loci (Supplementary Table 1) These results were in good agreement with previous promoter analyses that identified K-Rta responsive promoters in reporter assays conducted with 293 cells[31]. We examined the relationship between KSHV genomic structure and K-Rta
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