ChIP-seq of MUTZ3 cells in experiments to investigate EVI1 regulation by CRISPR or compound treatment
Ontology highlight
ABSTRACT: ChIP-seq was conducted to evaluate the effect of deleting a MYB binding site in the GATA2 super-enhancer or treating MUTZ3 cells with a MYB inhibitor. H3K27ac, MYB and p300 ChIP-seq datasets were generated.
Project description:Following a CRISPR enhancer scan covering the GATA2 super-enhancer region, the top sgRNAs were selected for further inspection. MUTZ3 cells were thus treated with the selected sgRNAs and the region of interested was subjected to amplicons sequencing (amplicon-seq). To that end, we used the Illumina PCR-based custom amplicon sequencing method using the TruSeq Custom Amplicon index kit (Illumina). The same experiment was conducted in K562 cells, which do not harbor an inv(3)/t(3;3), to investigate the role of MYB in this enhancer in other leukemia settings
Project description:We investigated whether GATA2 enhancer-driven transcription of EVI1 in inv(3)/t(3;3) is reversible in leukemia cells. In primary inv(3)/t(3;3) AML, immature CD34+CD15- cells can be discriminated from more mature CD34-CD15- and CD34-CD15+ cells. Besides, we also investigated the effect of deleting a MYB binding site on these cells using CRISPR/Cas9.
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.
Project description:Open chromatin regions were analyzed by ATAC-seq in MUTZ3 and MOLM1 cell lines (both inv(3) AML) to characterize the GATA2 super-enhancer region. ATAC-seq was performed as described (Buenrostro et al., 2013) with a modification in the lysis buffer to reduce mitochondrial DNA contamination.
Project description:In skeletal myogenesis, the transcription factor MyoD activates distinct transcriptional programs in progenitors compared to terminally differentiated cells. Using ChIP-seq and gene expression analyses, we show that in primary myoblasts, Snail-HDAC1/2 repressive complex bind and exclude MyoD from its targets. Notably, Snail binds E-box motifs that are G/C-rich in their central dinucleotides, and such sites are almost exclusively associated with genes expressed during differentiation. By contrast, Snail does not bind the A/T-rich E-boxes associated with MyoD targets in myoblasts. Thus, Snai1-HDAC1/2 prevents MyoD occupancy on differentiation-specific regulatory elements and the change from Snail- to MyoD-binding often results in enhancer switching during differentiation. Furthermore, we show that a regulatory network involving Myogenic Regulatory Factors (MRFs), Snail/2, miR-30a and miR-206 acts as a molecular switch that controls entry into myogenic differentiation. Together, these results reveal a regulatory paradigm that directs distinct gene expression programs in progenitors versus terminally differentiated cells. Genome wide binding sites of various transcription factors and chromatin modifiers in muscle cells
Project description:In bacteria, Crp-Fnr superfamily transcription factors mediate 3',5'-cyclic adenosine monophosphate (cAMP) and 3',5'-cyclic guanosine monophosphate (cGMP) signaling. The CRP-like protein Clr of the soil-dwelling and plant-symbiotic α-proteobacterium Sinorhizobium meliloti was previously shown to activate target promoters in both its cAMP- and cGMP-bound states (Krol et al., Microbiology 62:1840–1856, 2016). In order to further characterize the overall regulon of Clr in S. meliloti Chromatin Immuno Precipitation DNA-Sequencing (ChIP-Seq) experiments were performed with C-terminally FLAG tagged Clr under four different growth conditions, namely growth in TY complex medium and MOPS minimal medium, each supplemented with either cAMP or cGMP. For each condition, the respective immunoprecipitated (IP) and non-immunoprecipitated (control) samples were analyzed and compared to locate genomic positions in which Clr-DNA binding occurs. In combining ChIP-Seq with Electrophoretic Mobility Shift Assays and promoter-probe assays we expanded the list of known Clr-regulated target promoters and showed that virtually all of these promoters containing a palindromic Clr binding site (CBS) motif are activated both by Clr•cAMP and Clr•cGMP.
Project description:This SuperSeries is composed of the following subset Series: GSE24811: Time Series of Mouse skeletal muscle cell differentiation GSE24852: ChIP-Seq of MyoD, Myf5, Snai1, HDAC1, HDAC2, E47 and empty vector controls in mouse skeletal myoblasts or myotubes GSE38236: RNA-Seq of si-Snai1, si-Snai2, si-Snai1/2 and si-Scrambled treated myoblasts Refer to individual Series
Project description:Earlier we have shown important roles of MYB in pancreatic tumor pathobiology. To better understand the role of MYB in the tumor microenvironment and identify MYB-associated secreted biomarker proteins, we conducted mass spectrometry analysis of the secretome from MYB-modulated and control pancreatic cancer cell lines. We also performed in silico analyses to determine MYB-associated biofunctions, gene networks and altered biological pathways. We further investigated the secreted proteins for their potential biomarker properties.
Project description:Examination of binding locations of Pax3 and Pax7 in primary myoblasts UCSC track hub available at: http://www.ogic.ca/projects/Soleimani_2012_Pax7_hub/hub.txt For details on viewing the track hub in the UCSC Genome Browser: http://altair.dartmouth.edu/ucsc/goldenPath/help/hgTrackHubHelp.html#View 3 Samples (Control, Pax7 ChIP, Pax3 ChIP)