Abstract
Numerous pieces of evidence support the complex, 3D spatial organization of the genome dictates gene expression. CTCF is essential to define topologically associated domain boundaries and to facilitate the formation of insulated chromatin loop structures. To understand CTCF’s direct role in global transcriptional regulation, we integrated the miniAID-mClover3 cassette to the endogenous CTCF locus in a human pediatric B-ALL cell line, SEM, and an immortal erythroid precursor cell line, HUDEP-2, to allow for acute depletion of CTCF protein by the auxin-inducible degron system. In SEM cells, CTCF loss notably disrupted intra-TAD loops and TAD integrity in concurrence with a reduction in CTCF-binding affinity, while showing no perturbation to nuclear compartment integrity. Strikingly, the overall effect of CTCF’s loss on transcription was minimal. Whole transcriptome analysis showed hundreds of genes differentially expressed in CTCF-depleted cells, among which MYC and a number of MYC target genes were specifically downregulated. Mechanically, acute depletion of CTCF disrupted the direct interaction between the MYC promoter and its distal enhancer cluster residing ∼1.8 Mb downstream. Notably, MYC expression was not profoundly affected upon CTCF loss in HUDEP-2 cells suggesting that CTCF could play a B-ALL cell line specific role in maintaining MYC expression.
Highlights
CCCTC-binding factor (CTCF) is a transcription factor with 11 zinc finger domains ubiquitously expressed in cells of eukaryotes
Current characterization of chromatin loops discovered by high-resolution Hi-C, Hi-ChIP and ChIA-PET has to date only identified ∼7000 CTCF-mediated loops [77,78,79,80] suggesting that most -binding sites are not involved in facilitating DNA interactions, but in other roles such as direct transcriptional regulation
We observed that acute depletion of CTCF in SEM and HUDEP-2 cells did not lead to severe genome-wide transcriptional dysregulation
Summary
CCCTC-binding factor (CTCF) is a transcription factor with 11 zinc finger domains ubiquitously expressed in cells of eukaryotes. Conditional knock-out mouse studies using Ctcfflox/flox mice crossed with various tissue-specific Cre-expressing strains confirmed the requirement of Ctcf during organ development [6,7,8,9]. CTCF is well-characterized as a transcription factor that binds to insulator elements to define TAD boundaries, and along with cohesin proteins, promote the formation of CTCF–CTCF insulated chromatin loop structures. These structures spatially compartmentalize interacting promoters and enhancers into regulatory domains that allow for dynamic gene transcriptional regulation through proximal and distal intra-TAD chromatin looping [11,18,19,20]. IntraTAD DNA loops between enhancer and promoter regions have been shown to be facilitated by CTCF [18,21]
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