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Evi1 governs Kdm6b-mediated histone demethylation to finely regulate the Laptm4b-driven mTORC pathway in vivo [CUT&RUN]

ABSTRACT: Ecotropic viral integration site 1 (EVI1/MECOM) overexpression is common in myeloid malignancies. We present a new Evi1 transgenic mouse model with inducible expression in hematopoietic stem cells (HSCs) and progenitor cells (HPCs) at lower levels. Upon exogenous Evi1 induction, mice displayed anemia, thrombocytopenia, lymphopenia, and dysplasia in erythroid and megakaryocyte cells with a significant expansion of committed myeloid progenitor cells, resembling human Myelodysplastic syndrome/Myeloproliferative neoplasm (MDS/MPN)-like disease. Methods: Lin-C-Kit+ cells were isolated from 2 Evi1 overexpressing mice for CUT&RUN assay using flag antibody and IgG. Lin-C-Kit+ cells were isolated from 2 pair of WT mice and Evi1 overexpressing mice for CUT&RUN assay using H3K27me3 antibody.Around 5X105 Lin-C-Kit+ cells for each group were used in CUT&RUN assay. About 10 ng of the purified CUT&RUN DNA was used for preparation of multiplexed libraries with the NEB Ultra II DNA Library Prep Kit per manufacturer's instruction (NEB #E7103). Sequencing was conducted using an Illumina NextSeq 500 Sequencing System. Results: Molecular pathways altered in Lin-C-Kit+ cells from Evi1-OE mice. Conclusions: Evi1 has a special binding profiling in Lin-C-Kit+ cells. The modification profile of H3K27me3 was altered in Evi1-OE hematopoietic stem and progenitor cells.

ORGANISM(S): Mus musculus

PROVIDER: GSE235439 | GEO | 2023/06/27

REPOSITORIES: GEO

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Evi1 governs Kdm6b-mediated histone demethylation to finely regulate the Laptm4b-driven mTORC pathway in vivo [LK]

Project description:Ecotropic viral integration site 1 (EVI1/MECOM) overexpression is common in myeloid malignancies. We present a new Evi1 transgenic mouse model with inducible expression in hematopoietic stem cells (HSCs) and progenitor cells (HPCs) at lower levels. Upon exogenous Evi1 induction, mice displayed anemia, thrombocytopenia, lymphopenia, and dysplasia in erythroid and megakaryocyte cells with a significant expansion of committed myeloid progenitor cells, resembling human Myelodysplastic syndrome/Myeloproliferative neoplasm (MDS/MPN)-like disease. Methods: Lin-C-Kit+ cells were isolated from 3 pairs of WT and Evi1 overexpressing mice. Around 1X106 Lin-C-Kit+ cells were harvested from each mouse. Then the cells were lysated by Trizol and total RNA was extracted by phenol-chloroform. Results: Molecular pathways altered in Lin-C-Kit+ cells from Evi1-OE mice. Conclusions: Multiple molecular pathways changed in Evi1 overexpressing hematopoietic stem and progenitor cells.

2023-06-27 | GSE235440 | GEO

The SWI/SNF subunit Dpf2 licenses Nrf2-dependent gene expression to enforce multilineage control of inflammation [CUT&RUN]

Project description:Purpose: Multi-omics characterization of the consequences of deleting the SWI/SNF subunit Dpf2 in the transcriptome (RNAseq), accessibility of the genome (ATACseq) and binding of SWI/SNF subunits (CUT&RUN) in hematopoietic stem/progenitor cells, resting and polarized bone-marrow derived macrophages and resting and activated CD4+ T cells obtained from Dpf2f/f and Dpf2D/D mice. Methods: Lin- cKit+ cells were sorted from bone marrow of 28-days old Dpf2f/f and Dpf2D/D mice. Bone-marrow derived macrophages were obtained from 28-days old Dpf2f/f and Dpf2D/D mice and polarized with IFNg and LPS, or IL-4 for 24hours. CD4+ splenic T helper cells were obtained from 28-days old Dpf2f/f and Dpf2D/D mice and activated by treatment with PMA and Ionomycin. Methods: For CUT&RUN experiments, Lin- cKit+ cells were obtained from bone marrow of 28-days old Dpf2f/f and Dpf2D/D mice. Cells were crosslinked (1 min, 1% formaldehyde), quenched (125mM Glycine) and washed in buffers following the CUTANA ChIC/CUT&RUN kit according to the manufacturer’s CUT&RUN cross-linking protocol (EpiCypher, 14-1048). Results: For CUT&RUN experiments, 5 million (paired-end, 75bp) reads were obtained per sample. Pair-end fastq files were processed with the ENCODE Transcription Factor and Histone ChIP-Seq processing pipeline (https://github.com/ENCODE-DCC/chip-seq-pipeline2). Reads were trimmed using cutadapt v2.5. and aligned to the mm10 genome using Bowtie2 v2.3.4.3. SAMtools v1.9 were used to convert the output file to BAM format. Duplicates were removed using Picard Tools v2.20.7. Peak calling was performed with MACS2 v2.2.4. Conclusions: The absence of Dpf2 in LK cells results in downregulation of anti-oxidative and anti-inflammatory gene expression programs in bone marrow LK cells, bone-marrow derived macrophages and CD4+ T cells.

2023-06-06 | GSE192777 | GEO

CUT&RUN-seq of H3K4me3, H3K27me3 and H3K27ac in mll2wt and mll2[Y2602A] mESC

Project description:CUT&RUN-seq of H3K4me3, H3K27me3 and H3K27ac in mouse ESCs carrying a homozygous point mutation in the catalytic domain of mll2[Y2602A]. CUT&RUN was performed according to Skene & Henikoff, 2017 and purified DNA was using for library preparation with NEBNext DNA Ultra II kit (NEB E7645S; input: 5 ng of DNA). Libraries were multiplexed and sequenced on a NextSeq500 (Paired-End; read length 40). Each sample is present in 3 biological replicates.

2023-11-15 | E-MTAB-13466 | biostudies-arrayexpress

Transcriptome analysis of bone marrow cells of Evi1-overexpressing leukemia mouse

Project description:Puropose: To detemine transcriptome profile of Evi1-overexpressing leukemia cells by RNA sequencing Method: RNA samples were isolated from GFP+Lin-c-kit+ mouse bone marrow cells

2020-03-31 | GSE147736 | GEO

Evi1 governs Kdm6b-mediated histone demethylation to finely regulate the Laptm4b-driven mTORC pathway in vivo [LSK]

Project description:Ecotropic viral integration site 1 (EVI1/MECOM) overexpression is common in myeloid malignancies. We present a new Evi1 transgenic mouse model with inducible expression in hematopoietic stem cells (HSCs) and progenitor cells (HPCs) at lower levels. Upon exogenous Evi1 induction, mice displayed anemia, thrombocytopenia, lymphopenia, and dysplasia in erythroid and megakaryocyte cells with a significant expansion of committed myeloid progenitor cells, resembling human Myelodysplastic syndrome/Myeloproliferative neoplasm (MDS/MPN)-like disease. Methods: LSK cells were sorted by Sony SH800 from 3 pairs of mice transplanted with WT BM cells and Evi1-OE BM cells after pIpC injection. Around 5X103 LSK cells were harvested from each mouse. Then the cells were lysated by Trizol and total RNA was extracted by phenol-chloroform. Results: Molecular pathways altered in LSK cells from recipient mice transplanted with Evi1-OE BM cells. Conclusions: Transcription profile changed in Evi1 overexpressing hematopoietic stem and progenitor cells.

2023-06-27 | GSE235441 | GEO

A mucus production programme promotes classical pancreatic ductal adenocarcinoma

Project description:CUT&RUN for IgG, H3K4me3 and HA-SPDEF in human PDA HPAF-II cells and CUT&RUN HA-Spdef in murine KPC 2D FC1245 cells We performed CUT&RUN assay in human and murine PDA cells to profile the direct binding of SPDEF to target genes.

2024-01-24 | GSE250519 | GEO

Bromodomain member BRD4 is required for hematopoietic stem cell functions (CUT&RUN)

Project description:To investigate the role of BRD4 in normal hematopoiesis, Brd4 conditional KO mouse model was generated. We purified the Lin-cKit+ cells from bone marrow of WT and Brd4 KO mice and performed the CUT&RUN to study the histone modification and Brd4 occupancy around genome in HSC/HPCs population.

2023-08-20 | GSE189833 | GEO

Exploring the binding motif in genome wide for p53, Pygo2 and H3K4-Me3 in mouse (Pten/Smad4 KO) PS cell line

Project description:Since our data support that Pygo2 function is p53 Wild type status dependent and the interaction of Pygo2 with H3K4-Me3 is needed for the c-Kit regulation by Pygo2, however, neither Pygo2 nor p53 bind at c-Kit promoter region. To find out the potential co-binding locia for Pygo2, p53 and H3K4-Me3, we performed a Cut&Run assay followed by Miseq Sequencing.

2022-12-23 | GSE196486 | GEO

Cut&Run-seq of MUTZ3 cells to investigate changes in a translocated enhancer following CRISPR treatment

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

H3K4me3 associated chromatin by CUT&RUN on c-kit+ cells from chimeras

Project description:Sequencing of chromatin associated with H3K4me3 histone marks in live c-kit+ cells isolated by CUT&RUN from chimeras after transplantation with Ctrl-BM, MSC-BM and CpG-BM.

2023-03-24 | GSE199338 | GEO

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