Project description:Extensive changes in DNA methylation are common in cancer and may contribute to oncogenesis through transcriptional silencing of tumor suppressor genes. Genome-scale studies have yielded important insights into these changes, but have focused on CpG islands or gene promoters. We used whole-genome bisulfite sequencing (Bisulfite-Seq) to comprehensively profile a primary human colorectal tumor and adjacent-normal colon tissue at single-basepair resolution. Regions of focal hypermethylation in the tumor were located primarily at CpG islands and were concentrated within regions of long-range (>100 kb) hypomethylation. These hypomethylated domains covered nearly half the genome and coincided with late replication and attachment to the nuclear lamina in human cell lines. The confluence of hypermethylation and hypomethylation within these domains was confirmed in 25 diverse colorectal tumors with matched adjacent tissue. We propose that widespread DNA methylation changes in cancer are linked to silencing programs orchestrated by the 3D organization of chromatin within the nucleus. dbGAP study: phs000385 Primary tissue samples sequenced using 76-bp bisulfite sequencing (WGBS or Methyl-seq) using the Illumina GAII platform. Two independent libraries were constructed for each sample, and these libraries were combined into a single data file for each sample.
Project description:Extensive changes in DNA methylation are common in cancer and may contribute to oncogenesis through transcriptional silencing of tumor suppressor genes. Genome-scale studies have yielded important insights into these changes, but have focused on CpG islands or gene promoters. We used whole-genome bisulfite sequencing (Bisulfite-Seq) to comprehensively profile a primary human colorectal tumor and adjacent-normal colon tissue at single-basepair resolution. Regions of focal hypermethylation in the tumor were located primarily at CpG islands and were concentrated within regions of long-range (>100 kb) hypomethylation. These hypomethylated domains covered nearly half the genome and coincided with late replication and attachment to the nuclear lamina in human cell lines. The confluence of hypermethylation and hypomethylation within these domains was confirmed in 25 diverse colorectal tumors with matched adjacent tissue. We propose that widespread DNA methylation changes in cancer are linked to silencing programs orchestrated by the 3D organization of chromatin within the nucleus. dbGAP study: phs000385
Project description:A large fraction of the mammalian genome is organized into inactive chromosomal domains associated with the nuclear lamina. Using genomic repositioning assays we show that Lamina associated domains (LADs), spanning the developmentally regulated IgH and Cyp3a loci, contain transportable DNA regions that associate chromatin with the nuclear lamina and repress gene activity in fibroblasts. We characterized DNA regions within LADs that are functionally capable of positioning chromatin domains at the inner nuclear membrane (INM) lamina. We mapped and characterized the IgH and other LADs in murine fibroblasts. We show that these murine LADs have a unique chromatin structure with discrete boundaries. We demonstrate DNA regions within LADs that are capable of directing the association of chromatin domains with the INM-lamina as well as the silencing of a co-integrated reporter gene.
Project description:A large fraction of the mammalian genome is organized into inactive chromosomal domains associated with the nuclear lamina. Using genomic repositioning assays we show that Lamina associated domains (LADs), spanning the developmentally regulated IgH and Cyp3a loci, contain transportable DNA regions that associate chromatin with the nuclear lamina and repress gene activity in fibroblasts.
Project description:Structural Maintenance of Chromosomes flexible Hinge Domain containing 1 (SMCHD1) is an SMC-like protein that plays an important role in X chromosome inactivation and is mutated in two human diseases, a muscular dystrophy, and a rare developmental disorder. Here we characterized the function of SMCHD1 in human myoblasts and focused on its role on autosomes. We found that SMCHD1 colocalizes with Lamin B1 and the constitutive heterochromatin mark H3K9me3. Loss of SMCHD1 leads to a loss of heterochromatin at nuclear lamina regions and to extensive changes of the epigenome on most chromosomes. In SMCHD1-deficient cells, about half the regions that normally bind SMCHD1 loose H3K9me3, acquire long stretches of DNA hypermethylation and gain a variety of active chromatin marks including H3K4me3, H3K4me1, histone H3K27 acetylation, and H3K36 di- and trimethylation. Three-dimensional chromatin analysis using HiC shows that long range contacts between different heterochromatin blocks (B compartments) on the same chromosome are diminished while there is extensive formation of new topologically associated domains (TADs) and loops anchored at newly created CTCF/RAD21 binding sites. Inactivation of SMCHD1 promotes many inactive (B) to active (A) compartment transitions. These transitions are accompanied by activation of about 200 previously silenced genes with potential implications for the etiology and progression of SMCHD1-linked diseases. Our data suggest that SMCHD1 functions as a genome compartment organizer and as an important anchor for heterochromatin domains at the nuclear lamina ensuring that lamina- associated domains are protected from histone and DNA modification enzymes that typically operate in active chromatin.
Project description:Structural Maintenance of Chromosomes flexible Hinge Domain containing 1 (SMCHD1) is an SMC-like protein that plays an important role in X chromosome inactivation and is mutated in two human diseases, a muscular dystrophy, and a rare developmental disorder. Here we characterized the function of SMCHD1 in human myoblasts and focused on its role on autosomes. We found that SMCHD1 colocalizes with Lamin B1 and the constitutive heterochromatin mark H3K9me3. Loss of SMCHD1 leads to a loss of heterochromatin at nuclear lamina regions and to extensive changes of the epigenome on most chromosomes. In SMCHD1-deficient cells, about half the regions that normally bind SMCHD1 loose H3K9me3, acquire long stretches of DNA hypermethylation and gain a variety of active chromatin marks including H3K4me3, H3K4me1, histone H3K27 acetylation, and H3K36 di- and trimethylation. Three-dimensional chromatin analysis using HiC shows that long range contacts between different heterochromatin blocks (B compartments) on the same chromosome are diminished while there is extensive formation of new topologically associated domains (TADs) and loops anchored at newly created CTCF/RAD21 binding sites. Inactivation of SMCHD1 promotes many inactive (B) to active (A) compartment transitions. These transitions are accompanied by activation of about 200 previously silenced genes with potential implications for the etiology and progression of SMCHD1-linked diseases. Our data suggest that SMCHD1 functions as a genome compartment organizer and as an important anchor for heterochromatin domains at the nuclear lamina ensuring that lamina- associated domains are protected from histone and DNA modification enzymes that typically operate in active chromatin.
Project description:Structural Maintenance of Chromosomes flexible Hinge Domain containing 1 (SMCHD1) is an SMC-like protein that plays an important role in X chromosome inactivation and is mutated in two human diseases, a muscular dystrophy, and a rare developmental disorder. Here we characterized the function of SMCHD1 in human myoblasts and focused on its role on autosomes. We found that SMCHD1 colocalizes with Lamin B1 and the constitutive heterochromatin mark H3K9me3. Loss of SMCHD1 leads to a loss of heterochromatin at nuclear lamina regions and to extensive changes of the epigenome on most chromosomes. In SMCHD1-deficient cells, about half the regions that normally bind SMCHD1 loose H3K9me3, acquire long stretches of DNA hypermethylation and gain a variety of active chromatin marks including H3K4me3, H3K4me1, histone H3K27 acetylation, and H3K36 di- and trimethylation. Three-dimensional chromatin analysis using HiC shows that long range contacts between different heterochromatin blocks (B compartments) on the same chromosome are diminished while there is extensive formation of new topologically associated domains (TADs) and loops anchored at newly created CTCF/RAD21 binding sites. Inactivation of SMCHD1 promotes many inactive (B) to active (A) compartment transitions. These transitions are accompanied by activation of about 200 previously silenced genes with potential implications for the etiology and progression of SMCHD1-linked diseases. Our data suggest that SMCHD1 functions as a genome compartment organizer and as an important anchor for heterochromatin domains at the nuclear lamina ensuring that lamina- associated domains are protected from histone and DNA modification enzymes that typically operate in active chromatin.
Project description:Structural Maintenance of Chromosomes flexible Hinge Domain containing 1 (SMCHD1) is an SMC-like protein that plays an important role in X chromosome inactivation and is mutated in two human diseases, a muscular dystrophy, and a rare developmental disorder. Here we characterized the function of SMCHD1 in human myoblasts and focused on its role on autosomes. We found that SMCHD1 colocalizes with Lamin B1 and the constitutive heterochromatin mark H3K9me3. Loss of SMCHD1 leads to a loss of heterochromatin at nuclear lamina regions and to extensive changes of the epigenome on most chromosomes. In SMCHD1-deficient cells, about half the regions that normally bind SMCHD1 loose H3K9me3, acquire long stretches of DNA hypermethylation and gain a variety of active chromatin marks including H3K4me3, H3K4me1, histone H3K27 acetylation, and H3K36 di- and trimethylation. Three-dimensional chromatin analysis using HiC shows that long range contacts between different heterochromatin blocks (B compartments) on the same chromosome are diminished while there is extensive formation of new topologically associated domains (TADs) and loops anchored at newly created CTCF/RAD21 binding sites. Inactivation of SMCHD1 promotes many inactive (B) to active (A) compartment transitions. These transitions are accompanied by activation of about 200 previously silenced genes with potential implications for the etiology and progression of SMCHD1-linked diseases. Our data suggest that SMCHD1 functions as a genome compartment organizer and as an important anchor for heterochromatin domains at the nuclear lamina ensuring that lamina- associated domains are protected from histone and DNA modification enzymes that typically operate in active chromatin.
Project description:Structural Maintenance of Chromosomes flexible Hinge Domain containing 1 (SMCHD1) is an SMC-like protein that plays an important role in X chromosome inactivation and is mutated in two human diseases, a muscular dystrophy, and a rare developmental disorder. Here we characterized the function of SMCHD1 in human myoblasts and focused on its role on autosomes. We found that SMCHD1 colocalizes with Lamin B1 and the constitutive heterochromatin mark H3K9me3. Loss of SMCHD1 leads to a loss of heterochromatin at nuclear lamina regions and to extensive changes of the epigenome on most chromosomes. In SMCHD1-deficient cells, about half the regions that normally bind SMCHD1 loose H3K9me3, acquire long stretches of DNA hypermethylation and gain a variety of active chromatin marks including H3K4me3, H3K4me1, histone H3K27 acetylation, and H3K36 di- and trimethylation. Three-dimensional chromatin analysis using HiC shows that long range contacts between different heterochromatin blocks (B compartments) on the same chromosome are diminished while there is extensive formation of new topologically associated domains (TADs) and loops anchored at newly created CTCF/RAD21 binding sites. Inactivation of SMCHD1 promotes many inactive (B) to active (A) compartment transitions. These transitions are accompanied by activation of about 200 previously silenced genes with potential implications for the etiology and progression of SMCHD1-linked diseases. Our data suggest that SMCHD1 functions as a genome compartment organizer and as an important anchor for heterochromatin domains at the nuclear lamina ensuring that lamina- associated domains are protected from histone and DNA modification enzymes that typically operate in active chromatin.
Project description:How gene positioning to the nuclear periphery regulates transcription remains largely unclear. We have previously observed the differential compartmentalization of transcription factors and histone modifications at the nuclear periphery in mouse C2C12 myoblasts. Here, we have integrated high throughput DNA sequencing into the DNA adenine methyltransferase identification (DamID) assay, and have identified ~15, 000 sequencing-based Lamina-Associated Domains (sLADs) in mouse 3T3 fibroblasts and C2C12 myoblasts. These genomic regions range from a few kb to over 1 Mb and cover ~30% of the genome, and are spatially proximal to the nuclear lamina (NL). Active histone modifications such as H3K4me2, H3K9Ac, H3K36me3 and H3K79me2 are all localized away from the nuclear periphery microscopically, and distributed predominantly out of sLADs genome-wide. Therefore, the spatial compartmentalization of active histone modifications likely characterizes a major portion of chromatin at the nuclear periphery in mammalian cells. Genomic regions around transcription start sites of expressed sLAD genes display reduced associations with the NL and possess active histone modifications; in contrast, gene bodies of expressed sLAD genes possess very low levels of active histone modifications. Our genome-wide analyses of NL-associated chromatin have enabled functional and mechanistic dissections of gene positioning on transcription regulation. generate DamID maps of genome-NL interaction for mouse 3T3 fibroblasts and C2C12 myoblasts