CGGBP1-dependent CTCF-binding sites restrict ectopic transcription
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ABSTRACT:
PROVIDER: PRJNA721628 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR14228534_1.fastq.gz | Fastqsanger.gz | |||
| SRR14228535_1.fastq.gz | Fastqsanger.gz |
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Project description:CGGBP1-dependent CTCF-binding sites identified in Patel et. al. 2019 [PMID 31547883] serve as barrier elements consistent with asymmetrical levels of H3K9me3 in the flanks and asymmetrical RNA levels at these sites in the genome. In this study we have characterised the function of such CGGBP1-depenedent CTCF binding sites which are usually repeat rich in nature. By cloning one such CTCF-binding site in an episomal system, we have studied the barrier activity of this CGGBP1-dependent CTCF-binding sites in the prsesnce and absence of CGGBP1 in HEK293T cells through various molecular assays and RNA-sequencing. Our results show that these sites act as barrier for the ectopic transcription as the depletion of CGGBP1 lead to starnd-specific bidirectional transcription as a result of loss of barrier activity concomitant with the loss of CTCF-binding. Further, analysing the RNA-seq revealed that weakly transcribed sites are flanked by the CTCF-binding at transcription start and end sites in the presence of CGGBP1. However, CGGBP1 depletion leads to loss of barrier activity maintained by CGGBP1 with dispersed CTCF binding and a loss of transcription restriction. Such CGGBP1-dependent CTCF-binding sites prevents ectopic transcription.
2022-01-06 | GSE171990 | GEO
Project description:Binding sites of the chromatin regulator protein CTCF function as important landmarks in the human genome. The recently characterized CTCF-binding sites at LINE-1 repeats depend on another repeat-regulatory protein CGGBP1. These CGGBP1-dependent CTCF-binding sites serve as potential barrier elements for epigenetic marks such as H3K9me3. Such CTCF-binding sites are associated with asymmetric H3K9me3 levels as well as RNA levels in their flanks. The functions of these CGGBP1-dependent CTCF-binding sites remain unknown. By performing targeted studies on candidate CGGBP1-dependent CTCF-binding sites cloned in an SV40 promoter-enhancer episomal system we show that these regions act as inhibitors of ectopic transcription from the SV40 promoter. CGGBP1-dependent CTCF-binding sites that recapitulate their genomic function of loss of CTCF binding upon CGGBP1 depletion and H3K9me3 asymmetry in immediate flanks are also the ones that show the strongest inhibition of ectopic transcription. By performing a series of strand-specific reverse transcription PCRs we demonstrate that this ectopic transcription results in the synthesis of RNA from the SV40 promoter in a direction opposite to the downstream reporter gene in a strand-specific manner. The unleashing of the bidirectionality of the SV40 promoter activity and a breach of the transcription barrier seems to depend on depletion of CGGBP1 and loss of CTCF binding proximal to the SV40 promoter. RNA-sequencing reveals that CGGBP1-regulated CTCF-binding sites act as barriers to transcription at multiple locations genome-wide. These findings suggest a role of CGGBP1-dependent binding sites in restricting ectopic transcription.
| S-EPMC8794514 | biostudies-literature
Project description:To study the role of CGGBP1 in regulation of CTCF occupancy, we performed CTCF ChIP-seq in HEK293T cells with three different levels of CGGBP1: (i) endogenous levels of CGGBP1 (ii) CGGBP1 knockdown, and (iii) CGGBP1 overexpression. Here, we show that CTCF binds to repeats as well as canonical CTCF-motifs and CGGBP1 determines the CTCF occupancy preference for repeats over canonical CTCF-motif. By combining CTCF ChIP-seq with histone modifications ChIP-seq (for H3K4me3, H3K9me3 and H3K27me3) under conditions of normal or CGGBP1 knockdown, we demonstrate that CTCF binding sites regulated by CGGBP1 correspond to chromatin barrier elements with profound effects on H3K9me3 distribution. In conclusion, these finding shows that CGGBP1 is a regulator of CTCF occupancy and serve as a regulator of barrier functions of CTCF-binding sites.
2019-09-23 | GSE129548 | GEO
Project description:To study if regulation of cytosine methylation and CTCF occupancy are interdependent and governed by the levels of CGGBP1, we specifically pulled-down methylated cytosines and found that some transcription factor binding sites including that of CTCF held out against the cytosine methylation changes. The cytosine methylation at CTCF-binding repeat-free motifs show a non-stochastic depdendence on CGGBP1 and occur at sites that mark cytosine methylation transition boundaries. We also show that allelic imbalance dictates stochastic methylation changes due to CGGBP1 depletion.
2020-07-20 | GSE145300 | GEO
Project description:Truncated forms of CGGBP1 with (N-term) or without (C-term) the DNA-binding domain (DBD) have been used to to assay global cytosine methylation. HEK293T cells with endogenous CGGBP1 knocked down were used to over-express truncated forms of CGGBP1 followed by MeDIP-Seq analysis. Genome-wide analyses of cytosine methylation and binding of CGGBP1 DBD show that CGGBP1 restricts cytosine methylation in a manner that depends on its DBD and its DNA-binding. Our findings suggest that CGGBP1 protects transcription factor binding sites (TFBS) from cytosine methylation-associated loss. A superimposition of our results and evolution of CGGBP1 suggests that mitigation of cytosine methylation is majorly achieved by its N-terminal DBD. Our results position CGGBP1 DNA-binding as a major evolutionarily acquired mechanism through which it keeps cytosine methylation under check and regulates TFBS retention.
2024-11-17 | GSE269791 | GEO
Project description:CGGBP1 regulates chromatin barrier activity and CTCF occupancy at repeats
| PRJNA531700 | ENA
Project description:CGGBP1-regulated cytosine methylation at CTCF-binding motifs resists stochasticity
| PRJNA606696 | ENA
Project description:Truncated forms of CGGBP1 with (N-term) or without (C-term) the DNA-binding domain (DBD) have been used to to assay global gene expression. HEK293T cells with endogenous CGGBP1 knocked down were used to over-express truncated forms of CGGBP1 followed by RNA extraction and one-coloured global gene expression analysis. The data suggests that while the C-term of CGGBP1 is the major repressor of transcription, just the N-term containing the DBD fails to achieve so. Proximal promoters of CGGBP1-repressed genes, although significantly GC-poor, contain GC-rich transcription factor binding motifs and exhibit base compositions indicative of low C-T transition rates due to targeted prevention of cytosine methylation. Our findings suggest that CGGBP1 protects transcription factor binding sites (TFBS) from cytosine methylation-associated loss and thereby regulates gene expression. By analysing orthologous promoter sequences, we show that protection from cytosine methylation is a function of CGGBP1 progressively acquired during vertebrate evolution.
2024-11-17 | GSE269654 | GEO
Project description:Pervasive usage of alternative promoters leads to deregulation of gene expression in carcinogenesis and may drive the emergence of new genes in spermatogenesis. However, little is known regarding the mechanisms underpinning the activation of alternative promoters. In our present study, we uncovered a novel mechanism by which alternative cancer-testis-specific transcription is activated from the intergenic and intronic clustered CTCF binding sites, which are transcriptionally inert in normal somatic cells. BORIS/CTCFL, a paralog of CTCF with cancer-testis-specific expression forms a heterodimer with CTCF at the clustered binding sites thus triggering epigenetic reprogramming of these sites into units of active transcription. BORIS binding to CTCF sites leads to the recruitment of chromatin-remodeling factor SRCAP, with subsequent replacement of H2A histone with H2A.Z, therefore creating a more relaxed chromatin state in the nucleosomes flanking the clustered binding sites. This facilitates opening of chromatin beyond CTCF/BORIS binding sites and paves the way to the recruitment of multiple additional transcription factors, thereby activating transcription from a given binding site. We demonstrate that the CTCF binding sites, epigenetically reprogrammed by ectopic BORIS expression can drive the expression of cancer-testis genes, long-noncoding RNAs, retro-pseudogenes, and dormant transposable elements harbored by activated long transcripts. Taking together, our results reveal that BORIS functions as a transcription factor that epigenetically reprograms clustered CTCF binding sites into transcriptional start sites, promoting transcription from alternative promoters in both germ cells and cancer cells.
2024-01-03 | GSE207058 | GEO
Project description:Alu SINEs are the most numerous frequently occurring transcription units in our genome and possess sequence competence for transcription by RNA Pol III. However, through poorly understood mechanisms, the Alu RNA levels are maintained at very low levels in normal somatic cells with obvious benefits of low rates of Alu retrotransposition and energy-economical deployment of RNA Pol III to the tRNA genes which share promoter structure and polymerase requirements with Alu SINEs. Using comparative ChIP sequencing, we unveil that a repeat binding protein, CGGBP1, binds to the transcriptional regulatory regions of Alu SINEs thereby impeding Alu transcription by inhibiting RNA Pol III recruitment. We show that this Alu-silencing depends on growth factor stimulation of cells and subsequent tyrosine phosphorylation of CGGBP1. Importantly, CGGBP1 ensures a sequence-specific discriminative inhibition of RNA Pol III activity at Alu promoters, while sparing the structurally similar tRNA promoters. Our data suggest that CGGBP1 contributes to growth-related transcription by preventing the hijacking of RNA Pol III by Alu SINEs. This study was used to find out the effect of CGGBP1 on serum-induced changes in gene expression and effect of serum on gene expression regulation by CGGBP1. Gene expression profiling of normal human fibroblasts under 4 different experimental perturbations: serum starvation or serum stimulation and CGGBP1 depletion or normal CGGBP1 levels.
2015-12-21 | E-GEOD-54058 | biostudies-arrayexpress