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Conservation of parental genomic imprinting by cortex generated from embryonic stem cells [RRBS-seq]

ABSTRACT: In vitro cortex generated from embryonic stem cells (ESCs) is a model system to investigate corticogenesis and a promising tool for cortical therapy. A fundamental question that has implications for understanding corticogenesis and for using stem cells therapeutically is to determine whether in vitro cortex reproduces some fine-tuned epigenetic modifications that are important for corticogenesis and function in vivo such as parent-of-origin dependent DNA methylation and expression of imprinted genes (IGs). Here, we have compared at single-base resolution the parent-of-origin dependent DNA methylation and expression of IGs in hybrid cortices generated either in vivo or in vitro from ESCs using Reduced Representation Bisulfite Sequencing (RRBS) and RNA-seq. We report that in vitro cortex strictly reproduced the in vivo parental expression of 41 IGs, including those involved in corticogenesis such as Mest (paternal) and Cdkn1c (maternal). The expressed allele was set in ESCs and maintained during in vitro corticogenesis for most IGs but some switched from a biallelic expression in ESCs to the monoallelic expression observed in vivo. RRBS experiments revealed that parent-of-origin dependent methylation at imprinted loci were also largely similar in in vitro and in vivo cortices except at a few loci. The most discordant locus was Gpr1-Zdbf2: Zdbf2 RNA was paternal in vivo and biallelic in vitro, and this was concomitant with an aberrant gain of methylation on the maternal allele in vitro. Thus, we conclude that the epigenetic mechanisms at imprinted loci are largely but not strictly preserved in vitro. We propose that in vitro corticogenesis, with its set of IGs displaying faithful parent-of-origin dependent expression and methylation, helps to define the poorly known mechanisms regulating imprinting in the brain and roles of IGs during corticogenesis.

ORGANISM(S): Mus musculus

PROVIDER: GSE75485 | GEO | 2016/04/21

SECONDARY ACCESSION(S): PRJNA304417

REPOSITORIES: GEO

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Project description:Genomic imprinting is a mechanism in which the expression of genes varies depending on their parent-of-origin. Imprinting occurs through differential DNA methylation and histone modifications on the two parental alleles, with most imprinted genes marked by CpG-rich differentially methylated regions (DMRs). DNA methylation profiling in cases of uniparental disomy (UPD) provides a unique system permitting the study of DNA derived from a single parent (PMID: 20631049). Approximately 70 human imprinted genes have been described, and imprinted loci have been associated with diseases such as diabetes and cancer. We profiled parent of origin DNA methylation marks to find novel imprinted loci. Methods: We have an unprecedented collection of whole blood DNA from XX patients with UPD covering 18 different chromosomes, allowing for the efficient detection of DMRs associated with imprinted genes for 84% of the human genome. Our study is complimented with Ovarian Teratoma DNA (maternal DNA) and Complete hydatidiform Mole (paternal DNA). DNA methylation was profiled using Illumina Infinium 450K Methylation BeadArrays. Imprinted DMRs were defined by sites at which the maternal and paternal methylation levels diverged significantly from the biparental average. We confirmed novel DMRs by bisulfite sequencing of informative trios and SequenomEpiTYPER assays. Allelic specific gene expression studies were also performed by RNA sequencing in independent biparental controls. Findings: Our results provide for the first comprehensive map of the human imprintome, doubling the number of known imprinted regions. We identified a total of 71 DMRs, 41 of which were novel. 27 novel DMRs were maternally methylated and 14 were paternally methylated. We identified DMRs on chromosomes 5, 21 and 22 previously considered devoid of imprinting, highlighting potential parent-of-origin effects in chromosomal aneuploidies such as Down syndrome. We also found DMRs in genes associated with Schizophrenia and epilepsy. Interpretation: Our data provide the first comprehensive genome-wide map of imprinted sites in the human genome, and provide novel insights into potential parent-of-origin effects in human disorders. 66 UPD samples analyzed in total, From each individual, whole bllod DNA was extracted and global DNA methylation levels were assessed using Illumina Infinium HumanMethylation450 BeadChip.

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Conservation of parental genomic imprinting by cortex generated from embryonic stem cells [RRBS-seq]

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