Genome wide analysis of transcriptome modification after re-expression of PBRM1 WT and its BAH domain mutant in PBRM1 CRISPR-KO HEK293T cells.
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA630416 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR11687462_1.fastq.gz | Fastqsanger.gz | |||
| SRR11687462_2.fastq.gz | Fastqsanger.gz | |||
| SRR11687463_1.fastq.gz | Fastqsanger.gz | |||
| SRR11687463_2.fastq.gz | Fastqsanger.gz | |||
| SRR11687464_1.fastq.gz | Fastqsanger.gz |
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Project description:Purpose: To find out if cancer-associated mutation in BAH domain affects gene expression? Method: We used PBRM1 CRISPR KO HEK293T cells and ectopically expressed GFP-PBRM1-WT, GFP-PBRM1 P1048R, GFP-PBRM1 T1202K and GFP-PBRM1 C1233W plasmid for 48 hours. Result: We found that GFP-PBRM1 WT and GFP-PBRM1 T1202K mutant were identical in terms of gene expression. However, mutant P1048R and C1233W had very little difference in expression of only 15 (14 up- and 1 down) out of 10,160 genes.
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Project description:The Drosophila Trithorax group (TrxG) protein ASH1 remains associated with mitotic chromatin through mechanisms that are poorly understood. ASH1 dimethylates histone H3 at lysine 36 via its SET domain. Here we identify domains of the TrxG protein ASH1 that are required for mitotic chromatin attachment in living Drosophila. Quantitative live imaging demonstrates that ASH1 requires AT hooks and the BAH domain but not the SET domain for full chromatin binding in metaphase, and that none of these domains are essential for interphase binding. Genetic experiments show that disruptions of the AT hooks and the BAH domain together, but not deletion of the SET domain alone, are lethal. Transcriptional profiling demonstrates that intact ASH1 AT hooks and the BAH domain are required to maintain expression levels of a specific set of genes, including several involved in cell identity and survival. This study identifies in vivo roles for specific ASH1 domains in mitotic binding, gene regulation and survival that are distinct from its functions as a histone methyltransferase.
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Project description:The Polycomb Repressive Complex 2 (PRC2) and its trimethylation of histone H3 at lysine 27 (H3K27me3) control gene silencing, genome organization, cell-fate determination, as well as normal and pathological development. To date, functional transduction of H3K27me3 is believed to be achieved through the H3K27me3-‘recognizing’ chromodomain harbored within the chromobox (CBX) subunit of Polycomb Repressive Complex 1 (PRC1), which mediates gene silencing partly through H2A monoubiquitination. Here, we report a novel H3K27me3-readout mechanism in mammal, which utilizes an evolutionarily conserved Bromo-adjacent homology (BAH) domain of BAHCC1 (BAH domain and Coiled-Coil Containing 1) for silencing polycomb gene targets. Biochemical, structural and chromatin-immunoprecipitation followed by sequencing (ChIP-seq) analyses revealed that the BAH domain of BAHCC1 specifically engage H3K27me3 through a hydrophobic trimethyl-lysine-binding cage and multiple intermolecular interactions to its flanking residues, mediating co-localization of BAHCC1 with H3K27me3-marked genomic regions in cells. Additionally, we find that BAHCC1 is overexpressed in several human leukemia subtypes including T-cell acute lymphoblastic leukemia (T-ALL), and interacts with transcriptional repressors SAP30-binding protein (SAP30BP) and histone deacetylase 1 (HDAC1). BAHCC1 loss, or disrupting the BAH-mediated interaction of BAHCC1 with H3K27me3 via structure-based mutagenesis, causes chromatin remodeling at the H3K27me3-targeted loci and reactivates polycomb-related gene-silencing programs intimately associated with tumor suppression and cell differentiation, which leads to significantly suppressed T-ALL tumor growth in vitro and in vivo.
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Project description:PBRM1 is a component of the PBAF chromatin remodelling complex and has been observed to be deregulated in a significant proportion of patients with clear-cell Renal Cell Carcinoma (ccRCC). This study employs RNA-Seq to identify differentially expressed genes in cellular models of ccRCC by expressing PBRM1 in PBRM1-deficient Caki2 cells.
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Project description:Trimethylation of histone H3 lysine 27 (H3K27me3) is important for gene silencing, (epi)genome organization and organismal development. In a prevalent model, the functional “readout” of H3K27me3 in mammalian cells is achieved through the H3K27me3-recognizing chromodomain harbored within the chromobox (CBX) component of canonical Polycomb repressive complex 1 (cPRC1), which induces chromatin compaction and gene repression. Here, we report that binding of H3K27me3 by a Bromo Adjacent Homology (BAH) domain harbored within BAH domain-containing protein 1 (BAHD1) is required for overall BAHD1 targeting to chromatin and for optimal repression of the H3K27me3-demarcated genes in mammalian cells. Disruption of direct interaction between BAHD1-BAH and H3K27me3 by point mutagenesis leads to chromatin remodeling, notably, increased histone acetylation, at its Polycomb gene targets. Mice carrying an H3K27me3-interaction-defective mutation of Bahd1BAH causes marked embryonic lethality, showing a requirement of this pathway for normal development. Altogether, this work demonstrates an H3K27me3-initiated signaling cascade that operates through a conserved BAH “reader” module within BAHD1 in mammals.
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Project description:Here we utilize a forward genetics approach in Neurospora crassa to identify novel effectors of Polycomb repression. We recovered several mutant alleles of a gene (NCU07505),which we deemed effector of Polycomb repression 1 (epr-1), encoding a protein with a bromo-adjacent homology (BAH) domain and plant homeodomain (PHD) finger. Although epr-1mutants display phenotypic and gene expression changes similar to strains lacking PRC2 components, H3K27 methylation is notably unaffected. We demonstrate that EPR-1 forms nuclear foci, reminiscent of Polycomb bodies {Pirrotta:2012br}, and its genomic distribution is limited to and dependent upon H3K27-methylated chromatin, which it recognizes through its BAH domain. Finally, we discover that EPR-1 homologs are widely distributed across eukaryotes, contrary to previous reports {LopezGonzalez:2014id, Li:2018kg}, suggesting an ancient role of EPR-1 homologs in Polycomb repression that was secondarily lost in select lineages.
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Project description:Subunits of SWI/SNF chromatin remodeling complexes are frequently mutated in human malignancies. The PBAF complex is composed of multiple subunits, including the putative tumor suppressor proteins PBRM1 (BAF180) and ARID2 (BAF200) that are unique to this SWI/SNF complex. PBRM1 is mutated in various cancers, with a high mutation frequency in clear cell renal cell carcinoma (ccRCC). Here, we integrate RNA-seq, ARID2 and histone mark ChIP-seq, and ATAC-seq data to show that PBAF acts to enhance or repress gene expression depending on the genomic context. At baseline, ARID2 binds to areas of open chromatin at both active enhancers and promoters. Depletion of PBRM1 leads to attenuated and redistributed ARID2 chromatin binding that correlates significantly with gene expression changes. At enhancers, ARID2 binding loss leads to diminishment of the histone mark H3K4me1 and gene downregulation. Alternatively, at a subset of promoters, ARID2 binding loss derepresses gene expression. Interestingly, ARID2, which remains bound to other PBAF subunits after loss of PBRM1, is essential for many of the pro-tumorigenic transcriptional changes observed after loss of PBRM1, whereas other core SWI/SNF components are dispensable. Upon loss of PBRM1, ARID2 positively regulates cancer-related genes and pathways, including the cancer stem cell marker ALDH1A1 and EGF signaling, to stimulate tumor cell growth. Therefore, ARID2 is crucial for maintaining the transformed state of PBRM1-deficient ccRCC cells. In total, this study suggests a novel mechanism of transcriptional control by PBRM1, whereby its loss alters the chromatin distribution of the residual PBAF complex leading to altered transcription that promotes tumorigenesis.
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