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Coordination of KSHV Latent and Lytic Gene Control by CTCF-Cohesin Mediated Chromosome Conformation

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      Herpesvirus persistence requires a dynamic balance between latent and lytic cycle gene expression, but how this balance is maintained remains enigmatic. We have previously shown that the Kaposi's Sarcoma-Associated Herpesvirus (KSHV) major latency transcripts encoding LANA, vCyclin, vFLIP, v-miRNAs, and Kaposin are regulated, in part, by a chromatin organizing element that binds CTCF and cohesins. Using viral genome-wide chromatin conformation capture (3C) methods, we now show that KSHV latency control region is physically linked to the promoter regulatory region for ORF50, which encodes the KSHV immediate early protein RTA. Other linkages were also observed, including an interaction between the 5′ and 3′ end of the latency transcription cluster. Mutation of the CTCF-cohesin binding site reduced or eliminated the chromatin conformation linkages, and deregulated viral transcription and genome copy number control. siRNA depletion of CTCF or cohesin subunits also disrupted chromosomal linkages and deregulated viral latent and lytic gene transcription. Furthermore, the linkage between the latent and lytic control region was subject to cell cycle fluctuation and disrupted during lytic cycle reactivation, suggesting that these interactions are dynamic and regulatory. Our findings indicate that KSHV genomes are organized into chromatin loops mediated by CTCF and cohesin interactions, and that these inter-chromosomal linkages coordinate latent and lytic gene control.

      Author Summary

      Multiple mechanisms have been implicated in the control of herpesvirus latent and lytic gene regulation, but few mechanisms account for coordinate regulation of these two life cycles. Here, we show that the transcription control elements for KSHV latent and lytic genes are in close physical proximity. Mutations in the CTCF binding sites of the KSHV latency control region caused a loss of cohesin binding, and derepression of latent transcripts. Loss of CTCF binding also caused a loss of KSHV DNA copy number, and a failure to express lytic genes, including the immediate early gene Rta. Chromatin conformation capture (3C) methods indicated that the CTCF binding sites in the latency control region are linked to the promoter region of Rta. Additional chromatin linkages were detected between the 5′ and 3′ ends of the major latency transcripts, suggesting that chromatin loops organize both latent and lytic gene clusters. The interaction between latent and lytic control regions was subject to cell cycle regulation, consistent with earlier studies implicating cell cycle control of cohesin binding and viral transcription patterns. KSHV chromosome conformation was also disrupted by lytic cycle reactivation. We propose that CTCF-cohesin form dynamic linkages between viral regulatory domains to both insulate and coordinate latent and lytic gene expression.

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            Author and article information

            [1 ]The Wistar Institute, Philadelphia, Pennsylvania, United States of America
            [2 ]The Kyungpook National University, College of Pharmacy, Daegu, Korea
            [3 ]The University of Pennsylvania, School of Dentistry, Philadelphia, Pennsylvania, United States of America
            [4 ]The University of Pennsylvania, School of Medicine, Philadelphia, Pennsylvania, United States of America
            Emory University, United States of America
            Author notes

            Conceived and designed the experiments: HK PML . Performed the experiments: HK AW. Analyzed the data: HK YY ER PML. Contributed reagents/materials/analysis tools: YY ER. Wrote the paper: HK PML.

            Role: Editor
            PLoS Pathog
            PLoS Pathogens
            Public Library of Science (San Francisco, USA )
            August 2011
            August 2011
            18 August 2011
            : 7
            : 8
            Kang et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
            Pages: 15
            Research Article

            Infectious disease & Microbiology


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