Project description:Substantial evidence supports the hypothesis that enhancers are critical regulators of cell type determination, orchestrating both positive and negative transcriptional programs; however, the basic mechanisms by which enhancers orchestrate interactions with cognate promoters during activation and repression events remain incompletely understood. Here we report the required actions of the LIM domain binding protein, LDB1/CLIM2/NLI, interacting with the enhancer binding protein, ASCL1, to mediate looping to target gene promoters and target gene regulation in corticotrope cells. LDB1-mediated enhancer:promoter looping appears to be required for both activation and repression of these target target gene promoter genes. While LDB1-dependend activated genes are regulated at the level of transcriptional initiation, the LDB1-dependent repressed transcription units appear to be regulated primarily at the level of promoter pausing, with LDB1 regulating recruitment of MTA2, a component of the NuRD complex, on these negative enhancers, required for the repressive enhancer function. These results indicate that LDB1-dependent looping events can deliver repressive cargo to cognate promoters to mediate promoter pausing events in a pituitary cell type. ChIP assay followed by high throughput sequencing (ChIP-seq)

2015-02-03 | E-GEOD-64483 | biostudies-arrayexpress

Enhancer-bound LDB1 Regulates a Corticotrope Promoter Pausing Repression Program [GRO-seq]

Project description:Substantial evidence supports the hypothesis that enhancers are critical regulators of cell type determination, orchestrating both positive and negative transcriptional programs; however, the basic mechanisms by which enhancers orchestrate interactions with cognate promoters during activation and repression events remain incompletely understood. Here we report the required actions of the LIM domain binding protein, LDB1/CLIM2/NLI, interacting with the enhancer binding protein, ASCL1, to mediate looping to target gene promoters and target gene regulation in corticotrope cells. LDB1-mediated enhancer:promoter looping appears to be required for both activation and repression of these target target gene promoter genes. While LDB1-dependend activated genes are regulated at the level of transcriptional initiation, the LDB1-dependent repressed transcription units appear to be regulated primarily at the level of promoter pausing, with LDB1 regulating recruitment of MTA2, a component of the NuRD complex, on these negative enhancers, required for the repressive enhancer function. These results indicate that LDB1-dependent looping events can deliver repressive cargo to cognate promoters to mediate promoter pausing events in a pituitary cell type. Global Run On (GRO) assay followed by high throughput sequencing (GRO-seq)

2015-02-03 | E-GEOD-64515 | biostudies-arrayexpress

Enhancer-bound LDB1 Regulates a Corticotrope Promoter Pausing Repression Program [GRO-seq]

2015-02-03 | GSE64515 | GEO

Enhancer-bound LDB1 Regulates a Corticotrope Promoter Pausing Repression Program [ChIP-seq]

2015-02-03 | GSE64483 | GEO

Role of LDB1 in the transition from chromatin looping to transcription activation

Project description:Many questions remain about how close association of genes and distant enhancers occurs and how this is linked to transcription activation. In erythroid cells, LDB1 is recruited to the β-globin locus via LMO2 and is required for looping of the β-globin locus control region (LCR) to the active β-globin promoter. We show that the LDB1 dimerization domain (DD) is necessary and, when fused to LMO2 is sufficient, to completely restore LCR-promoter looping and transcription in LDB1 depleted cells. The looping function of the DD is unique and irreplaceable by heterologous dimerization domains. Dissection of the DD revealed distinct functional properties of conserved subdomains. Notably, a conserved helical region (DD4/5) is dispensable for LDB1 dimerization and chromatin looping but essential for transcriptional activation. DD4/5 is required for the recruitment of the co-regulators FOG1 and NuRD complex. Lack of DD4/5 alters histone acetylation and RNA polymerase II recruitment and results in failure of the locus to migrate to the nuclear interior as normally occurs during erythroid maturation. These results uncouple enhancer-promoter looping from nuclear migration and transcription activation and reveal new roles for LDB1in these processes. RNA-seq in LDB1 knockdown, LDB1 delta4/5 construct, LDB1 full-length construct, and control in induced MEL cells; three replicates each.

2014-07-02 | E-GEOD-54549 | biostudies-arrayexpress

Pax3 cooperates with Ldb1 to direct local chromosome architecture during lineage specification

Project description:Chromatin looping allows enhancer-bound regulatory factors to influence transcription. Large domains, referred to as Topologically Associated Domains (TADs), participate in genome organization but the mechanisms underlining interactions within TADs, that actually control gene expression, are largely unknown. Here we report that activation of embryonic myogenesis is associated with establishment of long-range chromatin interactions centered on Pax3-bound loci. Using mass spectrometry and genomic studies, we identified the ubiquitously expressed LIM-domain binding protein 1 (Ldb1) as the mediator of looping interactions at a subset of Pax3 binding sites. Ldb1 is recruited to Pax3-bound elements independently of CTCF-Cohesin, and is necessary for efficient deposition of H3K4me1 at these sites and chromatin looping. When Ldb1 is deleted in Pax3-expressing cells in vivo, specification of migratory myogenic progenitors is severely impaired. These results highlight Ldb1 requirement for Pax3 myogenic activity and demonstrate how a transcription factor promotes formation of sub-TAD interactions associated with lineage specification.

2019-04-16 | GSE126362 | GEO

RNA polymerase II and PARP1 shape enhancer-promoter contacts [CUT&Tag]

Project description:What controls enhancer-promoter communication to allow for cis-transcriptional regulation remains a mystery. Here we studied how transcriptional dynamics shapes enhancer-promoter contacts. We demonstrate that enhancer function is reflected in enhancer-promoter contacts frequency which highly depend on active transcription. Pol II pausing, which is widespread across metazoan enhancers and promoters, has a direct effect on focal enhancer-promoter contacts. We confirmed this effect by depleting NELFB, a subunit of the negative elongation factor complex and a pivotal factor in Pol II pausing. Moreover, the catalytic activity of PARP1, a regulator of transcription and chromatin condensation, stabilizes enhancer-promoter contacts globally, but can destabilize contacts by promoting Pol II escape from pausing. Based on these findings, we propose an updated model that couples transcription and enhancer-promoter contacts.

2022-06-18 | GSE206022 | GEO