Dataset Information

An insulator element 3' to the CFTR gene binds CTCF and reveals an active chromatin hub in primary cells.

ABSTRACT: Regulation of expression of the CFTR gene is poorly understood. Elements within the basal promoter of the gene do not fully explain CFTR expression patterns, suggesting that cis-regulatory elements are located elsewhere, either within the locus or in adjacent chromatin. We previously mapped DNase I hypersensitive sites (DHS) in 400 kb spanning the CFTR locus including a cluster of sites close to the 3'-end of the gene. Here we focus on a DHS at +6.8 kb from the CFTR translation end-point to evaluate its potential role in regulating expression of the gene. This DHS, which encompasses a consensus CTCF-binding site, was evident in primary human epididymis cells that express abundant CFTR mRNA. We show by DNase I footprinting and electophoretic mobility shift assays that the cis-regulatory element within this DHS binds CTCF in vitro. We further demonstrate that the element functions as an enhancer blocker in a well-established in vivo assay, and by using chromatin immunoprecipitation that it recruits CTCF in vivo. Moreover, we reveal that in primary epididymis cells, the +6.8 kb DHS interacts closely with the CFTR promoter, suggesting that the CFTR locus exists in a looped conformation, characteristic of an active chromatin hub.

SUBMITTER: Blackledge NP

PROVIDER: S-EPMC2651798 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature

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An insulator element 3' to the CFTR gene binds CTCF and reveals an active chromatin hub in primary cells.

Blackledge Neil P NP Ott Christopher J CJ Gillen Austin E AE Harris Ann A

Nucleic acids research 20090107 4

Regulation of expression of the CFTR gene is poorly understood. Elements within the basal promoter of the gene do not fully explain CFTR expression patterns, suggesting that cis-regulatory elements are located elsewhere, either within the locus or in adjacent chromatin. We previously mapped DNase I hypersensitive sites (DHS) in 400 kb spanning the CFTR locus including a cluster of sites close to the 3'-end of the gene. Here we focus on a DHS at +6.8 kb from the CFTR translation end-point to eval ...[more]

PMID: 19129223

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Project description:Herpes Simplex Virus 1 (HSV-1) is a human pathogen that has the ability to establish a lifelong infection in the host. During latency, HSV-1 genomes are chromatinized and are abundantly associated with histones in sensory neurons, yet the mechanisms that govern the latent-lytic transition remain unclear. We hypothesize that the latent-lytic switch is controlled by CTCF insulators, positioned within the HSV-1 latent genome. CTCF insulators, together with the cohesin complex, have the ability to establish and maintain chromtin loops that allow distance separated gene regions to be spatially oriented for transcriptional control. In this current study, we demonstrated that the cohesin subunit Rad21 was recruited to latent HSV-1 genomes near four of the CTCF insulators during latency. We showed that the CTCF insulator known as CTRS1/2, positioned downstream from the essential transactivating IE region of ICP4 was only enriched in Rad21 prior to but not during latency, suggesting that the CTRS1/2 insulator is not required for the maintenance of latency. Further, deletion of the CTRL2 insulator, positioned downstream from the LAT enhancer, resulted in a loss of Rad21 enrichment at insulators flanking the ICP4 region at early times post-infection in mice ganglia, suggesting that these insulators are interdependent. Finally, deletion of the CTRL2 insulator resulted in a loss of Rad21 enrichment at the CTRL2 insulator in a cell-type specific manner, and this loss of Rad21 enrichment was correlated to decreased LAT expression, suggesting that Rad21 recruitment to viral genomes is important for efficient gene expression.ImportanceCTCF insulators are important for transcriptional control and increasing evidence suggests that that CTCF insulators, together with the cohesin complex, regulate viral transcription in DNA viruses. The CTCF-cohesin interaction is important for the formation of chromatin loops, structures that orient distance separated elements in close spatial proximity for transcriptional control. Herpes Simplex Virus 1 (HSV-1) has seven putative CTCF insulators that flank the LAT and the IE, indicating that CTCF insulators play a role in the transition from latency to reactivation. Contributions from the work presented here include the finding that CTCF insulators in HSV-1 genomes are differentially enriched in the cohesin subunit Rad21, suggesting that CTCF-cohesin interactions could be establishing and anchoring chromatin loop structures to control viral transcription.

Dataset Information

An insulator element 3' to the CFTR gene binds CTCF and reveals an active chromatin hub in primary cells.

Publications

An insulator element 3' to the CFTR gene binds CTCF and reveals an active chromatin hub in primary cells.

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