Project description:Inborn defects in DNA repairare associated with complex developmental disorders whose causal mechanisms are poorly understood. Using an in vivo biotinylation tagging approach in mice, we show that the nucleotide excision repair (NER) structure-specific endonuclease ERCC1-XPF complex interacts with the insulator binding protein CTCF, the cohesin subunits SMC1A and SMC3 and with MBD2; the factors co-localize with ATRX at the promoters and control regions (ICRs) of imprinted genes during postnatal hepatic development. Loss of Ercc1or exposure to mitomycin C triggers the localization of CTCF to heterochromatin, the dissociation of the CTCF-cohesin complex and ATRXfrom promoters and ICRs,altered histone marks and the aberrant developmental expression of imprinted genes without altering DNA methylation. We propose that ERCC1-XPF cooperates with CTCF and the cohesinto facilitatet he developmental silencing of imprinted genes and that persistent DNA damage triggers chromatin changes that affect gene expression programs associated with NER disorders.
Project description:Parental imprinting results in a monoallelic parent-of-origin dependent gene expression. However, many imprinted genes identified by differential methylation do not exhibit complete monoallelic expression. Previous studies demonstrated a complex tissue-dependent expression patterns for some imprinted genes. Still, the complete magnitude of this phenomenon remains largely unknown. Differentiating human parthenogenetic induced pluripotent stem cells into different cell types and combining DNA methylation with novel 5' RNA sequencing methodology, enabled us to identify tissue- and isoform-dependent imprinted genes in a genome wide manner. We show that nearly half of all imprinted genes expresses both biallelic and monoallelic isoforms, that are controlled by tissue specific alternative promoters. This study provides the first global analysis of tissue-specific imprinting in humans, and implies that alternative promoters are central in the regulation of imprinted genes.
Project description:Parental imprinting results in a monoallelic parent-of-origin dependent gene expression. However, many imprinted genes identified by differential methylation do not exhibit complete monoallelic expression. Previous studies demonstrated a complex tissue-dependent expression patterns for some imprinted genes. Still, the complete magnitude of this phenomenon remains largely unknown. Differentiating human parthenogenetic induced pluripotent stem cells into different cell types and combining DNA methylation with novel 5' RNA sequencing methodology, enabled us to identify tissue- and isoform-dependent imprinted genes in a genome wide manner. We show that nearly half of all imprinted genes expresses both biallelic and monoallelic isoforms, that are controlled by tissue specific alternative promoters. This study provides the first global analysis of tissue-specific imprinting in humans, and implies that alternative promoters are central in the regulation of imprinted genes. Gene expression analysis was performed on a total of 6 human cell lines, including 4 iPSCs differentiated to neural progenitors and sorted using NCAM1 (2 control and 2 Parthenogenetic), and 2 iPSCs differentiated to endodermal progenitors (1 Control and 1 Parthenogenetic)
Project description:Large-scale transcriptome and methylome data analyses obtained by high-throughput technologies have been enabling the identification of novel imprinted genes. We investigated genome-wide DNA methylation patterns in multiple human tissues, using a high-resolution microarray to uncover hemimethylated CpGs located in promoters overlapping CpG islands, aiming to identify novel candidate imprinted genes. Using our approach, we recovered ~30% of the known human imprinted genes, further 168 candidates were identified, 61 of which with at least three hemimethylated CpGs shared by more than two tissue types. Thirty-four of these candidate genes are members of the protocadherins cluster on 5q31.3; in mice, protocadherin genes have non-imprinted monoallelic randomic expression, which might be the case in humans. Among the remaining 27 genes, ZNF331 was recently validated as an imprinted gene, and six of them have been reported as candidates, supporting our prediction. Five candidates (CCDC166, ARC, PLEC, TONSL and VPS28) map to 8q24.3, and might constitute a novel imprinted cluster. Additionally, we performed a comprehensive compilation of known human and mice imprinted genes from literature and databases, and a comparison among high-throughput imprinting studies in humans. The screening for hemimethylated CpGs shared by multiple human tissues, together with the extensive review, appears as a useful approach to reveal candidate imprinted genes.
Project description:As nucleosomes are widely replaced by protamine in mature human sperm, epigenetic contributions to embryo development appear limited. However, our genome-wide approaches find nucleosomes at low levels genome-wide, but also significantly enriched at imprinted gene clusters, miRNA clusters, HOX gene clusters, and the promoters of other developmental transcription and signaling factors. Developmental promoters were often DNA hypomethylated, and bore histone modifications localized to discrete locations: H3K4me2 is enriched at certain developmental promoters, whereas large blocks of H3K4me3 localize to a subset of developmental promoters, regions in HOX loci, certain non-coding RNAs, and generally to paternally-expressed imprinted loci. In contrast, H3K4me3 is generally absent at paternally-repressed imprinted loci. Interestingly, repressive H3K27me3 is enriched at many developmental promoters that lack early expression in embryos, with significant overlap with bivalent (H3K4me3/H3K27me3) promoters in ES cells. Taken together, epigenetic marking in sperm is extensive, and correlated with developmental regulators.
Project description:As nucleosomes are widely replaced by protamine in mature human sperm, epigenetic contributions to embryo development appear limited. However, our genome-wide approaches find nucleosomes at low levels genome-wide, but also significantly enriched at imprinted gene clusters, miRNA clusters, HOX gene clusters, and the promoters of other developmental transcription and signaling factors. Developmental promoters were often DNA hypomethylated, and bore histone modifications localized to discrete locations: H3K4me2 is enriched at certain developmental promoters, whereas large blocks of H3K4me3 localize to a subset of developmental promoters, regions in HOX loci, certain non-coding RNAs, and generally to paternally-expressed imprinted loci. In contrast, H3K4me3 is generally absent at paternally-repressed imprinted loci. Interestingly, repressive H3K27me3 is enriched at many developmental promoters that lack early expression in embryos, with significant overlap with bivalent (H3K4me3/H3K27me3) promoters in ES cells. Taken together, epigenetic marking in sperm is extensive, and correlated with developmental regulators.