Project description:We previously mapped ETV1 using ChIP-Seq in GIST48 cells (GSE22441). Here, we map the enhancer landscape marked by histone H3K4me1 and the promoter landscape marked by histone H3K4me3 in GIST48 cells. Crosslink ChIP-Seq of H3K4me1 and H3K4me3 in GIST48 cells
Project description:FOXA1 is a pioneer factor that is important in hormone dependent cancer cells to stabilise nuclear receptors, such as estrogen receptor (ER) to chromatin. FOXA1 binds to enhancers regions that are enriched in H3K4mono- and dimethylation (H3K4me1, H3K4me2) histone marks and evidence suggests that these marks are requisite events for FOXA1 to associate with enhancers to initate subsequent gene expression events. However, exogenous expression of FOXA1 has been shown to induce H3K4me1 and H3K4me2 signal at enhancer elements and the order of events and the functional importance of these events is not clear. We performed a FOXA1 Rapid Immunoprecipitation Mass Spectrometry of Endogenous Proteins (RIME) screen in ERα-positive MCF-7 breast cancer cells in order to identify FOXA1 interacting partners and we found histone-lysine N-methyltransferase (MLL3) as the top FOXA1 interacting protein. MLL3 is typically thought to induce H3K4me3 at promoter regions, but recent findings suggest it may contribute to H3K4me1 deposition, in line with our observation that MLL3 associates with an enhancer specific protein. We performed MLL3 ChIP-seq in breast cancer cells and unexpectedly found that MLL3 binds mostly at non-promoter regions enhancers, in contrast to the prevailing hypothesis. MLL3 was shown to occupy regions marked by FOXA1 occupancy and as expected, H3K4me1 and H3K4me2. MLL3 binding was dependent on FOXA1, indicating that FOXA1 recruits MLL3 to chromatin. Motif analysis and subsequent genomic mapping revealed a role for Grainy head like protein-2 (GRHL2) which was shown to co-occupy regions of the chromatin with MLL3. Regions occupied by all three factors, namely FOXA1, MLL3 and GRHL2, were most enriched in H3K4me1. MLL3 silencing decreased H3K4me1 at enhancer elements, but had no appreciable impact on H3K4me3 at enhancer elements. We identify a complex relationship between FOXA1, MLL3 and H3K4me1 at enhancers in breast cancer and propose a mechanism whereby the pioneer factor FOXA1 can interact with a chromatin modifier MLL3, recruiting it to chromatin to facilitate the deposition of H3K4me1 histone marks, subsequently demarcating active enhancer elements.
Project description:Here we present the whole genome ChIP-Seq analyses of a wide variety of histone marks, H3K27ac, H3K4me1, H3K4me3, and H3K27me3 in the brain, heart, and liver, along with the RNA-seq data of these organs of early human embryos 12 weeks after gestation. In total, brain, heart, and liver of early human post-implantation embryos were used, and four histone modifications were detected, including H3K27ac, H3K4me1, H3K4me3 and H3K27me3. Also, the transcriptomes of these three organs were analyzed.
Project description:ChIP-seq was performed using Drosophila Kc167 cells using antibodies against H3K4me3 to identify active promoters and H3K4me1 to identify active enhancers. H3K27ac ChIPseq was performed to identify active promoters and enhancers. Once enhancers and promoters were identified, JIL-1 and histone phosphorylation, H3K9acS10ph and H3K27acS28ph, ChIP-seq was performed to look at binding trends. JIL-1 and phosphoacetlation is found at low levels at inactive enhancers and shows increase at active enhancers and promoters. Here we examine histone phosphorylation by JIL-1 and acetylation of H3K27ac by CBP at transcriptionally active vs. inactive promoters and enhancers. ChIP-seq is performed in Kc167 Drosophila cells using antibodies against JIL-1, H3K27acS28ph, H3K9acS10ph, H3K4me3, H3K4me1, and H3K27ac.
Project description:DNA Methyltransferase 3A (DNMT3A) is frequently mutated in various hematopoietic malignancies; however, the underlying oncogenic mechanisms remain elusive. Here, we report that DNMT3A mutational âhotspotâ at Arg882 (DNMT3A-R882H) cooperates with constitutively activated RAS in transforming murine hematopoietic stem/progenitor cells (HSPCs) ex vivo and inducing acute leukemias in vivo. DNMT3A-R882H potentiates aberrant transactivation of âstemnessâ gene expression programs, notably transcription factors Meis1, Hox-A, Mn1 and Mycn. Mechanistically, R882-mutated DNMT3A directly binds to cis-regulatory elements of these genes and induces focal CpG hypomethylation reminiscent of what was seen in human leukemias bearing DNMT3A R882 mutation. Furthermore, DNMT3A-R882H induced DNA hypomethylation facilitates gene enhancer/promoter activation and recruitment of Dot1l-associated transcription elongation machineries. Inactivation of Dot1l represses DNMT3AR882H-mediated stem cell gene dysregulation and acute leukemogenicity. In this dataset, we provided R882H-mutated DNMT3A, H3K4me1, H3K4me3 and H3K27me3 ChIP-seq profiling data of RH-RAS LSCs, and H3K4me1 ChIP-seq data in HOXA9-MEIS1 LSCs. Genome-wide binding of R882H-mutated DNMT3A (Myc tagged; ChIP-seq with 9e10 anti-Myc antibodies) and histone modification profiles for H3K4me1, H3K4me3 and H3K27me3 were generated by ChIP-seq using specific antibodies in RH-RAS LSCs. Genome-wide H3K4me1 histone modification profiles were generated by ChIP-seq using H3K4me1 specific antibody in HOXA9-MEIS1 LSCs.
Project description:Here we used ChIP-MS to quantitatively profile chromatin-associated proteins that are specifically associated with H3K4me1- and H3K4me3-modified nucleosomes in IMR-90 chromatin.
Project description:Histone H3 lysine 4 (H3K4) can be mono-, di-, and trimethylated by members of the COMPASS (COMplex of Proteins ASsociated with Set1) family from yeast to human and these modifications can be found at distinct regions of the genome. Monomethylation of histone H3K4 (H3K4me1) is relatively more enriched at metazoan enhancer regions compared to trimethylated histone H3K4 (H3K4me3), which are found at transcription start sites in all eukaryotes. Our recent studies in Drosophila demonstrated that the Trithorax-related (Trr) branch of the COMPASS family regulates enhancer activity and is responsible for the implementation of H3K4me1 at these regions. There are six COMPASS family members in mammals, two of which, MLL3 and MLL4, are most closely related to Drosophila Trr. Here, we use ChIP-seq of this class of COMPASS family members in both human HCT116 cells and mouse embryonic stem cells and find that MLL4 is preferentially found at enhancer regions. MLL3 and MLL4 are frequently mutated in cancer, and indeed, the widely used HCT116 cancer cell line contains inactivating mutations in the MLL3 gene. Using HCT116 cells in which MLL4 has also been knocked out, we demonstrate that MLL4 is a major regulator of H3K4me1 in these cells, with the greatest loss of monomethylation at enhancer regions. Moreover, we found a redundant role between Mll3 and Mll4 in enhancer H3K4 monomethylation in mouse embryonic fibroblast (MEF) cells. These findings suggest that mammalian MLL3/MLL4 function in the regulation of enhancer activity and enhancer-promoter communication during gene expression and that mutations of MLL3 and MLL4 found in cancer could exert their properties through enhancer malfunction. ChIP-Seq in mouse embryonic stem (mES) cells for MLL4. ChIP-seq of MLL4 and p300 in human parental HCT116 cells. ChIP-seq of H3K4me1, H3K4me2 and H3K4me3 in parental HCT116 cells and HCT116 cells with Mll4∆set.
Project description:We performed ChIP-Seq for Ets1 and histone modifications in P5424 thymic cells, without and with Ets1 knockdown via shRNA. Overall, we find that loss of Ets1 results in specific, higher occupancy of H3K4me1-marked nucleosomes at the Ets1 binding site, but not H3K4me3 nucleosomes. We verified the specificity of this mechanism as Ets1-dependent by also computing H3K4me1 and 3 nucleosome occupancy in hypersensitive, Ets1-depleted sites. This effect was also found in primary WT DN+DP Ets1 sites. Overall, this suggests that Ets1 induces chromatin remodeling and that its loss increases H3K4me1 nucleosome occupancy in bound sites, with levels of H3K4me3 comparatively unchanged, thus reducing the activation status of these enhancers Genome-wide analysis via ChIP-Seq for Ets1, H3K4me1 and H3K4me3 in WT, sh-scramble and sh-Ets1 the P5424 thymic cell line
Project description:Phf8 is a histone demethylase with links to human behavior. To gain insight into Phf8's function in the brain, we have performed ChIP-seq for the histone methylation marks H3K4me3, H3K4me1, H3K27me2 and H3K9me2.