CRISPR enhancer scan to detect critical elements for the regulation of EVI1 in a translocated enhancer
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ABSTRACT:
PROVIDER: PRJEB42274 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| ERR5029503.fastq.gz | Fastqsanger.gz |
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Project description:To uncover, in an unbiased fashion, which elements of the 18 kb translocated region control EVI1 transcription, we devised a CRISPR/Cas9-based enhancer scanning approach. We considered all possible sgRNA target sites containing a canonical Cas9 PAM site (NGG) on both strands of the minimal 18 kb translocated region. Deep-sequencing libraries were generated by PCR amplification of sgRNA guide strands using primers that tag the product with standard Illumina adapters and a 4 bp sample barcode in a 2 step-PCR protocol.
2021-07-26 | E-MTAB-9948 | biostudies-arrayexpress
Project description:In acute myeloid leukemia (AML) with inv(3)(q21;q26) or t(3;3)(q21;q26), a translocated GATA2 enhancer drives oncogenic expression of EVI1. We generated an EVI1-GFP AML model and applied an unbiased CRISPR/Cas9 enhancer scan to uncover sequence motifs essential for EVI1 transcription. Using this approach, we pinpoint a single regulatory element in the translocated GATA2 enhancer that is critically required for aberrant EVI1 expression while being dispensable for GATA2 expression from its endogenous locus. This element contains a DNA binding motif for the transcription factor MYB that heavily occupies this site specifically at the translocated allele. MYB knockout as well as peptidomimetic blockade of p300-dependent MYB function resulted in downregulation of EVI1 but not of GATA2. Targeting MYB or mutating its DNA-binding motif within the GATA2 enhancer resulted in myeloid differentiation and cell death, suggesting that interference with MYB-driven EVI1 transcription provides a potential entry point for therapy of inv(3)/t(3;3) AMLs.
| EGAS00001004839 | EGA
Project description:Chromosomal rearrangements are a frequent cause of oncogene deregulation in human malignancies. Overexpression of EVI1 is found in a subgroup of acute myeloid leukemia (AML) with 3q26 chromosomal rearrangements which are often therapy resistant. In a cohort of primary t(3;8)(q26;q24) AML samples we observed the translocation of a MYC super-enhancer to EVI1. We generated a patient-based t(3;8)(q26;q24) model in vitro using CRISPR-Cas9 technology and demonstrated hyper-activation of EVI1 by the hijacked MYC super-enhancer. One MYC super-enhancer element in particular, which recruits early hematopoietic regulators, is critical for EVI1 expression and enhancer-promoter interaction. This interaction is facilitated by a CTCF-bound motif upstream of the EVI1 promoter that acts as an enhancer-docking site in t(3;8) AML. Genomic analyses of 3q26-rearranged AML samples point to a common mechanism by which EVI1 uses this CTCF-bound enhancer-docking site to hijack early hematopoietic enhancers.
| EGAS00001004808 | EGA
Project description:We examined the effects of targeting the GATA2 super-enhancer on EVI1 expression in MUTZ3. To that end, we conducted genome editing with CRISPR and assessed H3K27 acetylation with Cut&Run. The protocol described by the Henikoff group was used to generate these data.
2021-07-26 | E-MTAB-9946 | biostudies-arrayexpress
Project description:Chromosomal aberrations in acute myeloid leukemia (AML), such as inv(3) and t(3;3), lead to deregulation of the EVI1 oncogene by the GATA2 distal hematopoietic enhancer (G2DHE). In this project, we aimed to study the transcription factor complexes involved in the regulation of the G2DHE sequence. We have identified PARPi as a member of the G2DHE complex. Here, we used RNA-Seq to analyze transcriptomic changes after PARP inhibition with olaparib and talazoparib and to compare those to EVI1 knockdown.
2021-04-27 | GSE167005 | GEO
Project description:Chromosomal aberrations in acute myeloid leukemia (AML), such as inv(3) and t(3;3), lead to deregulation of the EVI1 oncogene by the GATA2 distal hematopoietic enhancer (G2DHE). In this project, we aimed to study the transcription factor complexes involved in the regulation of the G2DHE sequence. We identified PARP1 as an interactor of G2DHE-associated transcription factors. In this dataset, we studied the interaction of genomic loci between the EVI1 promoter and G2DHE by 4C-Seq in the 3q-rearranged AML cell line MUTZ-3 treated with the PARP1 inhibitors olaparib, talazoparib or the DMSO vehicle control for 24 h.
2021-04-27 | GSE153306 | GEO
Project description:We studied the variations of mRNA amounts after Flag-EVI1, Flag-EVI1?324, or Flag expression in HeLa cells. Despites EVI1 discovery in 1988, its recognized role as a dominant oncogene in myeloid leukemia and more recently in epithelial cancers, only a few target genes were known and it was not clear why EVI1 was involved in cancer progression. Here we obtained the genomic binding occupancy and expression data for EVI1 in human cells. We identified numerous EVI1 target cancer genes and genes controlling cell migration and adhesion. Moreover, we characterized a transcriptional cooperation between AP1 and EVI1 that regulated proliferation and adhesion through a feed-forward loop. This study provides human genome-wide mapping and expression analyses for EVI1 that will be useful for the research community. 12 samples were collected. Each condition was done in 4 replicates, collected 24 hours after transfection (for mild expression of EVI1 or EVI1?324). Transfections with Flag-expressing vector were used as controls.
2012-11-14 | E-GEOD-42251 | biostudies-arrayexpress
Project description:Acute myeloid leukemia (AML) driven by the activation of EVI1 due to chromosome 3q26/MECOM rearrangements is incurable with current chemotherapy regimens. Insight into the mechanism by which EVI1 drives myeloid transformation is needed to target EVI1 in those leukemias. Here we demonstrate recurrent interaction of CTBP1/2 with a unique PLDLS motif in EVI1, which is indispensable for leukemic transformation of 3q26/MECOM rearranged AML. A PLDLS competitor construct outcompetes EVI1 to CTBP1/2 binding and inhibits AML proliferation in xenotransplant models. This proof-of-concept study opens the possibility to target one of the most incurable forms of AML using specific inhibitors directed to EVI1/CTBP1/2 interaction. Our findings have important implications for other tumour types with aberrant expression of EVI1 or other oncogenic transcription factors that also depend on CTBP1/2 interaction.
2024-05-21 | PXD048760 | Pride
Project description:Acute myeloid leukemia (AML) driven by the activation of EVI1 due to chromosome 3q26/MECOM rearrangements is incurable with current chemotherapy regimens. Insight into the mechanism by which EVI1 drives myeloid transformation is needed to target EVI1 in those leukemias. Here we demonstrate recurrent interaction of CTBP1/2 with a unique PLDLS motif in EVI1, which is indispensable for leukemic transformation of 3q26/MECOM rearranged AML. A PLDLS competitor construct outcompetes EVI1 to CTBP1/2 binding and inhibits AML proliferation in xenotransplant models. This proof-of-concept study opens the possibility to target one of the most incurable forms of AML using specific inhibitors directed to EVI1/CTBP1/2 interaction. Our findings have important implications for other tumour types with aberrant expression of EVI1 or other oncogenic transcription factors that also depend on CTBP1/2 interaction.
2023-07-13 | PXD043333 | Pride
Project description:Cancer cells utilize epigenetic alterations to acquire autonomous capabilities for tumor maintenance. Here, we show that pancreatic ductal adenocarcinoma (PDA) cells hijack super-enhancers (SEs) to activate the EVI1 gene with its tumor-intrinsic program. We demonstrate that SE is the vital cis-acting element to maintain aberrant EVI1 transcription in PDA cells. Related to disease progression and inferior survival outcomes in human PDA, functional experiments further show that EVI1 upregulation is the cause of aggressive tumor phenotypes. In particular, EVI1 mediates the resistance to anchorage-independent growth and enhanced motility in vitro, with efficient tumor propagation in vivo. EVI1-dependent activation of gene expression program attributes to these phenotypes, by which EVI1 drives the stepwise configuration of the active enhancer chromatin. In sum, our findings support the promise that EVI1 is a crucial driver of oncogenic transcription program in PDA cells. Further, we emphasize the instructive role of epigenetic aberrancy in establishing PDA tumorigenesis.
2021-05-20 | GSE166510 | GEO