Project description:We analyzed RNA-seq, ATAC-seq, ChIP-seq and 4C-seq data to find that CTCF binding site located between HOXA7 and HOXA9 genes (CBS7/9) is critical for establishing and maintaining aberrant HOXA9-HOXA13 gene expression in AML. Disruption of the CBS7/9 boundary resulted in spreading of repressive H3K27me3 into the posterior active HOXA chromatin domain that subsequently impaired enhancer/promoter chromatin accessibility and disrupted ectopic long-range interactions among the posterior HOXA genes. Consistent with the role of the CBS7/9 boundary in HOXA locus chromatin organization, attenuation of the CBS7/9 boundary function reduced posterior HOTTIP lincRNA and HOXA gene expression and altered myeloid specific transcriptome profiles important for pathogenesis of myeloid malignancies.
Project description:Chromatin insulators partition the genome into the functional units to control gene expression especially in complex chromosomal regions. The CCCTC-binding factor (CTCF) is an insulator-binding protein which functions for transcriptional regulation and higher-order chromatin formation. Here we report that a removable CTCF insulator is responsible for the retinoic acid (RA)-mediated higher-order chromatin remodeling and selective gene expression in the human HOXA gene locus. Our detailed chromatin analyses characterized multiple CTCF-enriched sites and RA-responsive enhancers in this locus. These regulatory elements and transcriptionally silent HOXA genes were closely positioned under the basal condition. Notably, upon the RA signaling, the transcription factor RAR/RXR induced the loss of an adjacent CTCF binding and changed the higher-order chromatin conformation of the overall locus, establishing transcriptionally active, poised and repressive domains that were separated by stably localizing CTCF insulators. This study uncovers that removable insulator spatiotemporally switches higher-order chromatin and multiple gene activities at the chromosomal domain levels, via the cooperation of CTCF and key transcription factors.
Project description:Chromatin insulators partition the genome into the functional units to control gene expression especially in complex chromosomal regions. The CCCTC-binding factor (CTCF) is an insulator-binding protein which functions for transcriptional regulation and higher-order chromatin formation. Here we report that a removable CTCF insulator is responsible for the retinoic acid (RA)-mediated higher-order chromatin remodeling and selective gene expression in the human HOXA gene locus. Our detailed chromatin analyses characterized multiple CTCF-enriched sites and RA-responsive enhancers in this locus. These regulatory elements and transcriptionally silent HOXA genes were closely positioned under the basal condition. Notably, upon the RA signaling, the transcription factor RAR/RXR induced the loss of an adjacent CTCF binding and changed the higher-order chromatin conformation of the overall locus, establishing transcriptionally active, poised and repressive domains that were separated by stably localizing CTCF insulators. This study uncovers that removable insulator spatiotemporally switches higher-order chromatin and multiple gene activities at the chromosomal domain levels, via the cooperation of CTCF and key transcription factors.
Project description:We found that posterior HOXA-associated HOTTIP lncRNA is aberrantly activated in MLL-rearranged AMLs and required for the posterior HOXA chromatin structure and gene expression. Knock-out of HOTTIP attenuates leukemic progressions of the transplanted humanized AML mice by blocking posterior HOXA-associated AML gene expression programs while the dCas-VP160 mediated reactivation of HOTTIP restores HOXA locus chromatin structure and HOXA9-A13 gene expression in the CBS7/9 boundary depleted AML cells. Finally, transgenic expression of HOTTIP lncRNA in mouse bone marrow hematopoietic cells resulted in perturbation of the balance of HSC self-renewal and differentiation and development of AML like disease by aberrant altering HOXA associated chromatin structure and transcription program. Thus, HOTTIP lncRNA acts as oncogene to reprogram leukemic associated chromatin and gene transcription.
Project description:Catalytic activity of the ISWI family of remodelers is critical for nucleosomal organization and transcription factor binding, including the insulator protein CTCF. To define which subcomplex mediates these diverse functions we phenotyped a panel of isogenic mouse stem cell lines each lacking one of six ISWI accessory subunits. Individual deletions of either CERF, RSF1, ACF, WICH or NoRC subcomplexes only moderately affect the chromatin landscape, while removal of the NURF-specific subunit BPTF leads to drastic reduction in chromatin accessibility and Snf2h ATPase localization around CTCF sites. While this reduces distances to the adjacent nucleosomes it only modestly impacts CTCF binding itself. In absence of accessibility, the insulator function of CTCF is nevertheless impaired resulting in lower occupancy of cohesin and cohesin-loading factors, and reduced insulation at these sites, highlighting the need of NURF-mediated remodeling for open chromatin and proper CTCF function. Our comprehensive analysis reveals a specific role for NURF in mediating Snf2h localization and chromatin opening at bound CTCF sites showing that local accessibility is critical for cohesin binding and insulator function.
Project description:Embryonic stem cells (ESCs) cells run a self-renewal gene expression program, requiring the expression of certain transcription factors accompanied by a particular chromosome organization to maintain a balance between pluripotency and the capacity for rapid differentiation. However, how transcriptional regulation is linked to chromosome organization in ESCs remains enigmatic. Here we show that Cohesin exhibits a functional role in maintaining ESC identity through association with the pluripotency transcriptional network. ChIP-seq analyses of the cohesin subunit Rad21 reveal an ESC specific cohesin binding pattern that is characterized by a CTCF independent colocalization of cohesin with pluripotency related transcription factors. Upon ESC differentiation, these binding sites disappear and instead new CTCF independent Rad21 binding sites emerge, which are enriched for binding sites of transcription factors implicated in early differentiation. Furthermore, knock-down of cohesin subunits causes expression changes that are reminiscent of the depletion of key pluripotency transcription factors, demonstrating the functional relevance of the cohesin - pluripotency transcriptional network association. Finally, we show that Nanog physically interacts with the cohesin interacting proteins Stag1 and Wapl, further substantiating this association. Based on these findings we propose that a dynamic placement of cohesin by pluripotency transcription factors contributes to a chromosome organization supporting the ESC expression program. This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
Project description:The homeotic genes (Hox genes) encode transcription factors (HOX-TFs) that are key regulators of animal development. Single and compound deletion of Hox genes in mice revealed that they act in a partially redundant manner to pattern the vertebrate limb. Biochemical screens probing the sequence specificity of the DNA-binding domains showed that HOX-TFs recognize largely similar DNA sequences, but also emphasized the important role of co-factors in HOX DNA-binding. However, due to their high sequence homology and overlapping expression patterns, little is known about the genome-wide binding of these transcription factors Here, we set out to systematically compare the effects of the nine limb-bud expressed HOX-TFs on cell differentiation and gene regulation, and compare their genome-wide binding characteristics. We find that HOX-TFs induce distinct regulatory programs in transduced cells. Through genome-wide DNA binding profiling we find that the posterior HOX-TFs can be separated into two groups with distinct binding motifs and association with co-factors. Through this unexpected grouping, we characterize the CCCTC-binding factor (CTCF) as a novel co-factor of HOX-TFs and show that one, but not the other group of HOX-TFs binds to thousands of CTCF-occupied sites in the chicken genome.
Project description:The homeotic genes (Hox genes) encode transcription factors (HOX-TFs) that are key regulators of animal development. Single and compound deletion of Hox genes in mice revealed that they act in a partially redundant manner to pattern the vertebrate limb. Biochemical screens probing the sequence specificity of the DNA-binding domains showed that HOX-TFs recognize largely similar DNA sequences, but also emphasized the important role of co-factors in HOX DNA-binding. However, due to their high sequence homology and overlapping expression patterns, little is known about the genome-wide binding of these transcription factors Here, we set out to systematically compare the effects of the nine limb-bud expressed HOX-TFs on cell differentiation and gene regulation, and compare their genome-wide binding characteristics. We find that HOX-TFs induce distinct regulatory programs in transduced cells. Through genome-wide DNA binding profiling we find that the posterior HOX-TFs can be separated into two groups with distinct binding motifs and association with co-factors. Through this unexpected grouping, we characterize the CCCTC-binding factor (CTCF) as a novel co-factor of HOX-TFs and show that one, but not the other group of HOX-TFs binds to thousands of CTCF-occupied sites in the chicken genome.