Project description:Nuclear lamins contact the genome at the nuclear periphery through large domains and are involved in chromatin organization. Among broad peak calling algorithms available to date, none are suited for mapping lamin-genome interactions genome-wide. We disclose a novel algorithm, Enriched Domain Detector (EDD), for analysis of broad enrichment domains from ChIP-seq data. EDD enables discovery of genomic domains interacting with broadly distributed chromatin-associated proteins such as lamins. The main advantage of EDD over existing broad peak callers is sensitivity to domain width rather than enrichment strength at a particular site, and robustness against local variations. EDD is downloadable from http://github.com/eivindgl/edd. LMNA ChIP-seq experiments in human normal dermal fibroblasts (Lonza CC-2511; LDFs) and human normal primary dermal fibroblasts (Norwegian Stem Cell Center AD04DFs).
Project description:Nuclear lamins contact the genome at the nuclear periphery through large domains and are involved in chromatin organization. Among broad peak calling algorithms available to date, none are suited for mapping lamin-genome interactions genome-wide. We disclose a novel algorithm, Enriched Domain Detector (EDD), for analysis of broad enrichment domains from ChIP-seq data. EDD enables discovery of genomic domains interacting with broadly distributed chromatin-associated proteins such as lamins. The main advantage of EDD over existing broad peak callers is sensitivity to domain width rather than enrichment strength at a particular site, and robustness against local variations. EDD is downloadable from http://github.com/eivindgl/edd. RNA-seq experiments in human normal dermal fibroblasts (Lonza CC-2511; LDFs) and human normal primary dermal fibroblasts (Norwegian Stem Cell Center AD04DFs).
Project description:Hutchinson-Gilford progeria syndrome (HGPS) is a premature aging disease that is frequently caused by a de novo point mutation at position 1824 in LMNA. This mutation activates a cryptic splice donor site in exon 11, and leads to an in-frame deletion within the prelamin A mRNA and the production of a dominant negative lamin A protein, known as progerin. Here we show that HGPS cells experience genome-wide alterations in patterns of H3K27me3 deposition, changes in the associations of genomic loci with nuclear lamin A/C, and, at late passages, genome-wide loss of spatial compartmentalization of active and inactive chromatin domains that characterizes chromosome folding in normal cells. We further demonstrate that the H3K27me3 changes associate with gene expression alterations in HGPS cells. Our results support a model that the accumulation of progerin in the nuclear lamina leads to altered H3K27me3 marks in heterochromatin, possibly through the down-regulation of EZH2, and disrupts heterochromatin-lamina interactions. These changes may then lead to the genomic disorganization and changes in transcriptional regulation we observe in HGPS fibroblasts. ChIP-Seq was performed for H3K27me3 and lamin A/C in the human genome in three primary fibroblast cell lines: an HGPS patient (HGPS), normal cells from the father of the HGPS patient (Father), and age-matched normal cells (Age Control). Two biological replicates were performed.
Project description:Genomic information is encoded on a wide range of distance scales, ranging from tens of base pairs to megabases. We developed a multiscale framework to analyze and visualize the information content of genomic signals. Different types of signals, such as GC content or DNA methylation, are characterized by distinct patterns of signal enrichment or depletion across scales spanning several orders of magnitude. These patterns are associated with a variety of genomic annotations, including genes, nuclear lamina associated domains, and repeat elements. By integrating the information across all scales, as compared to using any single scale, we demonstrate improved prediction of gene expression from Polymerase II ChIP-seq measurements and we observed that gene expression differences in colorectal cancer are not most strongly related to gene body methylation, but rather to methylation patterns that extend beyond the single-gene scale. ChIP-seq data of six proteins in primary murine bone marrow macrophage cells (BMMs) under unstimulated and lipopolysaccharide (LPS) stimulated conditions. The BMMs were cultured from female C57BL/6 mice (age 8-12 weeks). Amongst these six proteins were three transcription factors (TFs), ATF340, NFκB/p50 and NFκB/p65, all of which are involved in regulating macrophage activation by microbial molecular components such as LPS. The other three ChIP-seq targets were RNA polymerase II (Pol II), and two chromatin modification marks: acetylation of histone H4 (H4ac) and tri-methylation of histone H3 lysine 27 (H3K27me3).
Project description:This SuperSeries is composed of the following subset Series: GSE41751: Correlated alterations in genome organization, histone methylation, and DNA-lamina interactions in Hutchinson-Gilford progeria syndrome (expression) GSE41757: Correlated alterations in genome organization, histone methylation, and DNA-lamina interactions in Hutchinson-Gilford progeria syndrome (ChIP-seq) GSE41763: Correlated alterations in genome organization, histone methylation, and DNA-lamina interactions in Hutchinson-Gilford progeria syndrome (Hi-C) Refer to individual Series
Project description:The glucocorticoid receptor (GR) binds the human genome at >10,000 sites, but only regulates the expression of hundreds of genes. To determine the functional effect of each site, we measured the glucocorticoid (GC) responsive activity of nearly all GR binding sites (GBSs) captured using chromatin immunoprecipitation (ChIP) in A549 cells. 13% of GBSs assayed had GC-induced activity. The responsive sites were defined by direct GR binding via a GC response element (GRE) and exclusively increased reporter- gene expression. Meanwhile, most GBSs lacked GC-induced reporter activity. The non-responsive sites had epigenetic features of steady state enhancers and clustered around direct GBSs. Together, our data support a model in which clusters of GBSs observed with ChIP-seq reflect interactions between direct and tethered GBSs over tens of kilobases. We further show that those interactions can synergistically modulate the activity of direct GBSs, and may therefore play a major role in driving gene activation in response to GCs. Glucoroticoid receptor binding site chip-seq libraries were cloned into STARR-seq for massively parallel functional analysis. The results were confirmed by ChIP-Exo performed on the GR in A549 cells treated with 100 nM dexamethasone for one hour. This dataset  contains ChIP-seq data for A549 cells treated with DEX or EtOH for 3 hours.
Project description:To evaluate the effect of CG methylation on DNA binding of sequence-specific B-ZIP transcription factors (TFs) in a high-throughput manner, we enzymatically methylated the cytosine in the CG dinucleotide on protein binding microarrays. Using this novel technology, we show that CG methylation enhanced binding for CEBPA and CEBPB and inhibited binding for CREB, ATF4, JUN, JUND, CEBPD and CEBPG. The CEBPB|ATF4 heterodimer bound a novel motif CGAT|GCAA 10-fold better when methylated. EMSA confirmed these results. CEBPB ChIP-seq data using primary female mouse dermal fibroblasts with 50X methylome coverage for each strand indicate that the methylated sequences well-bound on the arrays are also bound in vivo. CEBPB bound 39% of the methylated canonical 10-mers ATTGC|GCAAT in the mouse genome. After ATF4 protein induction by thapsigargin which results in ER stress, CEBPB binds methylated CGAT|GCAA in vivo, recapitulating what was observed on the arrays. mRNA-seq of primary female mouse dermal fibroblasts with and without thapsigargin identified differentially expressed genes. Genes that are commonly bound by CEBPB and ATF4 to TGAT|GCAA (the best-bound 8-mer in the array) at the promoters were highly expressed and up-regulated or remained unchanged in the thapsigargin treated primary female mouse dermal fibroblasts. RNA-Seq: Examination of whole genome transcriptome profiles (RNA-seq) of primary mouse dermal fibroblasts with and without Thapsigargin treatment ChIP-Seq: Examination of transcription factor binding in dermal fibroblasts with and without Thapsigargin teratment BS-Seq: Determination of whole genome DNA methylation profiles (BS-seq) of primary mouse dermal fibroblasts
Project description:Little is known about the epigenomics of liposarcoma (LPS). Here, we profiled the global expression of 9 epigenetic marks in well differentiated (WD) and dedifferentiated (DD) LPS from 151 patients and found increased H3K9me3 among DDLPS tumors. We performed ChIP-seq and gene expression profiling of patient derived cell lines to discover functionally significant regions of differential H3K9me3 enrichment between WDLPS and DDLPS associated with concomitant gene expression changes. We performed genome-wide transcriptional profiling of H3K9me3 in dedifferentiated liposarcoma DDLPS and well differentiated liposarcoma WDLPS cell lines.
Project description:Kir2.1, an inward rectifier potassium channel, plays an important role in controlling membrane potential specially in cardiomyocytes. Here we explore its interactome via proximity biotin labeling and affinity purification approach (BioID). First, BioID membrane controls were generated with randomly selected transmembrane domains from yeast proteins, demonstrating the identification of membrane-associated proteins more than GFP or NLS controls. Using this control and a mutation from Andersen-Tawil syndrome which causes Kir2.1 Golgi accumulation, we have identified the most comprehensive Kir2.1 interactome and also found clues to its spatial information in subcellular levels. The interactome is showing that Kir2.1 in plasma membrane is surrounded by desmosome, integrin, cadherin and dystrophin-glycoprotein complex complexes, and supported by MAGUK family scaffold proteins. On the other hand, Kir2.1 mutant BioID revealed the COPII-mediated delivery of Kir2.1 from ER to Golgi and lysosome-mediated degradation of Golgi-accumulated and/or retrograde Kir2.1. Validating the interactome with co-immunoprecipitation, confocal microscope, and patch clamp analysis, we concluded that Kir2.1 is located in the defined membrane environment and is actively regulated by endosomal sorting for lysosome degradation.
Project description:Mono-methylation of histone H3 on lysine 4 (H3K4me1) and acetylation of histone H3 on lysine 27 (H3K27ac) are histone modifications that are highly enriched over the body of actively transcribed genes and enhancers. Although in yeast all H3K4 methylation patterns including H3K4me1 are implemented by Set1/COMPASS, there are three classes of COMPASS-like complexes in Drosophila that could carry out H3K4me1 on enhancers: dSet1, Trithorax and Trithorax-related (Trr). Here, we report that Trr, the Drosophila homolog of mammalian Mll3/4, can function as a major H3K4 mono-methyltransferase on enhancers in vivo. Loss of Trr results in a global decrease of H3K4me1 and H3K27ac in various tissues. Assays with the cut wing margin enhancer imply a functional role for Trr in enhancer-mediated processes. A genome-wide analysis demonstrates that Trr is required for H3K4me1 and H3K27ac on chromatin signatures that resemble the histone modification patterns described for enhancers. Since Trr and mammalian Mll3/4 complexes are distinguished by bearing a unique subunit, the H3K27 demethylase UTX, we propose a model in which the H3K4 mono-methyltransferase Trr, and the H3K27 demethylase, UTX, cooperate to regulate the transition from inactive/poised to active enhancers. ChIP-seq of Trr, LPT, UTX in Drosophila S2 Cells. ChIP-seq of H3K4me1, H3K4me3, H3K27ac, H3K27me3 in WT and Trr knock-down Drosophila S2 cells. ChIP-seq of H3K4me1, H3K27me3 in LPT knock-down Drosophila S2 cells. ChIP-seq of LPT and UTX in Trr knock-down Drosophila S2 cells. ChIP-seq of H3K4me1 and H3K27me3 in MLL1(+/+), MLL1(-/-), MLL3(+/+), and MLL3(-/-) Mouse Embryonic Fibroblasts (MEFs).