Female human iPS cells retain an inactive X-chromosome
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE22246 | GEO | 2010/08/19
SECONDARY ACCESSION(S): PRJNA128759
REPOSITORIES: GEO
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Project description:Comparison of human iPSC lines, ESC and fibroblasts to determine their expression patterns. All early passage female lines profiled expressed XIST RNA which is an indicator of an inactive X chromosome. Genes on the X-chromosome were also analyzed for overall levels of gene expression compared to human fibroblasts. hiPSC lines were generated and extracted for RNA with TriZOL and hybridization on Affymetrix arrays.
2010-08-18 | E-GEOD-22246 | biostudies-arrayexpress
Project description:Female human ESC-lines can carry active X-chromosomes (Xa) or an XIST-RNA- coated inactive X-chromosome (Xi XIST+ ). Additionally, many ESC-lines have abnormal X-chromosome-inactivation (XCI)-states where the Xi no longer expresses XIST-RNA and has transcriptionally active regions (eroded Xi=Xe). The fate of each XCI-state upon differentiation is unclear because individual lines often contain a mixture of XCI-states. Here, we established homogeneous XiXa, XeXa, and XaXa ESC-lines. We found that these lines were unable to initiate XIST-expression and X-chromosome- wide silencing upon differentiation indicating that the ESC XCI-state is maintained in differentiated cells. Consequently, differentiated XeXa and XaXa cells displayed higher levels of X-linked gene-expression than XiXa cells. Although global transcriptional compensation between X-chromosomes and autosomes is not required for female ESC-differentiation, the degree of X-chromosome- silencing influences differentiation efficiencies. Our data suggest that the Xi XIST+ Xa state is inherent to human ESCs and that all other XCI-states, including XaXa, are abnormal and arise during ESC-derivation or maintenance.
2016-12-15 | GSE91072 | GEO
Project description:Female human ESC-lines can carry active X-chromosomes (Xa) or an XIST-RNA-coated inactive X-chromosome (XiXIST+). Additionally, many ESC lines have abnormal X-chromosomeinactivation (XCI)-states where the Xi no longer expresses XIST-RNA and has transcriptionally active regions (eroded Xi=Xe). The fate of each XCI-state upon differentiation is unclear because individual lines often contain a mixture of XCI-states. Here, we established homogeneous XiXa, XeXa, and XaXa ESC-lines. Employing RNA-FISH, RNA-sequencing and DNA methylation analyses, we found that these lines were unable to initiate XIST-expression and X-chromosome-wide silencing upon differentiation indicating that the ESC XCI-state is maintained in differentiated cells. Consequently, differentiated XeXa and XaXa cells displayed higher levels of X-linked gene-expression than XiXa cells. Although global transcriptional compensation between X-chromosomes and autosomes is not required for female ESC-differentiation, the degree of X-chromosome-silencing influences differentiation efficiencies. Our data suggest that the XiXIST+Xa state is inherent to human ESCs and that all other XCI-states, including XaXa, are abnormal and arise during ESC-derivation or maintenance.
2017-01-26 | GSE88931 | GEO
Project description:Female human ESC-lines can carry active X-chromosomes (Xa) or an XIST-RNA-coated inactive X-chromosome (XiXIST+). Additionally, many ESC lines have abnormal X-chromosomeinactivation (XCI)-states where the Xi no longer expresses XIST-RNA and has transcriptionally active regions (eroded Xi=Xe). The fate of each XCI-state upon differentiation is unclear because individual lines often contain a mixture of XCI-states. Here, we established homogeneous XiXa, XeXa, and XaXa ESC-lines. Employing RNA-FISH, RNA-sequencing and DNA methylation analyses, we found that these lines were unable to initiate XIST-expression and X-chromosome-wide silencing upon differentiation indicating that the ESC XCI-state is maintained in differentiated cells. Consequently, differentiated XeXa and XaXa cells displayed higher levels of X-linked gene-expression than XiXa cells. Although global transcriptional compensation between X-chromosomes and autosomes is not required for female ESC-differentiation, the degree of X-chromosome-silencing influences differentiation efficiencies. Our data suggest that the XiXIST+Xa state is inherent to human ESCs and that all other XCI-states, including XaXa, are abnormal and arise during ESC-derivation or maintenance.
2017-01-26 | GSE88932 | GEO
Project description:In mammals, chromosome-wide regulatory mechanisms ensure a balance of X-linked gene dosage between males (XY) and females (XX). In female cells, expression of genes from one of the two X-chromosomes is curtailed, with selective accumulation of Xist-RNA, Xist-associated proteins, specific histone modifications (eg. H3K27me3) and Barr body formation observed throughout interphase. Using chromosome flow-sorting, we show that during mitosis, Xist-associated proteins dissociate from inactive X (Xi) chromosomes, while high levels of H3K27me3 and increased compaction of the Xi relative to active X (Xa), are retained. Proteomic comparison of mitotic Xi and Xa revealed, unexpectedly, that components of Hbo1 and Msl/Mof histone acetyltransferase complexes co-enrich with Xa, while inhibitors of histone acetylation co-enrich with Xi. Furthermore, inhibition of Hbo1 or deletion of Msl/Mof components functionally abolishes mitotic differences in H3K27me3 marking and chromosome compaction. These data uncover critical roles for acetylation pathways in preserving X chromosome properties during mitosis.
2025-05-23 | PXD054014 | Pride
Project description:Erasure of epigenetic memory is required to convert somatic cells towards pluripotency. Reactivation of the inactive X chromosome (Xi) has been used to model epigenetic reprogramming in mouse, but human studies are hampered by Xi epigenetic instability and difficulties in tracking partially reprogrammed iPSCs. Here we used cell fusion to examine the earliest events in the reprogramming-induced Xi reactivation of human female fibroblasts. We show a rapid and widespread loss of Xi-associated H3K27me3 and XIST in fused cells that precedes the bi-allelic expression of selected Xi-genes by many heterokaryons (30-50%). After cell division, RNA-FISH and RNA-Seq analysis confirmed that Xi reactivation remained partial and showed that induction of human pluripotency-specific XACT transcripts occurred, but was rare (1%). These data effectively separate pre- and post-mitotic events in reprogramming-induced Xi reactivation, and suggest a hierarchy where early events such as XIST-delocalisation, are required but are insufficient to establish stable human X reactivation. We performed RNA-sequencing of human fibroblast clones derived from TERT-immortalised NHDF17914 (Lonza) before and at 5 days after fusion with mouse ESC (E14Tg2a:puroR). Two biological replicates were performed and sequenced.
2016-07-01 | E-GEOD-77443 | biostudies-arrayexpress
Project description:The inactive X chromosome (Xi) is folded in a stepwise process into a compartment-less structure. Here we investigate if the Xi can be unfolded in post-XCI cells. We began with ablating structural maintenance of chromosomes hinge domain containing 1 (Smchd1) in female mouse embryonic fibroblasts (MEFs), a cell type in which XCI is completed. We then performed in situ Hi-C on one Smchd1+/+ (wild-type, WT) and one Smchd1-/- (knockout, KO) MEF clone to probe the role of SMCHD1 in maintaining the higher-order structure of the Xi. To determine if the change in chromosome structure is accompanied by altered gene expression, we performed RNA-seq on two WT and two independently generated Smchd1-/- clones treated with either DMSO or 5-aza-2'-deoxycytidine (Aza), a DNA demethylating agent. In addition, we performed H3K27me3 and H2AK119ub ChIP-seq and Xist Capture Hybridization Analysis of RNA Targets with deep sequencing (CHART-seq) on one WT and one Smchd1-/- MEF clone to determine if there is an alteration in the epigenetic state of the Smchd1-/- Xi. Finally, we examined the role of Xist, Polycomb repressive complex 1 (PRC1), and Heterogeneous Nuclear Ribonucleoprotein K (HNRNPK) in regulating the Xi structure in post-XCI cells. To do so we performed in situ Hi-C on WT and Smchd1-/- MEFs depleted with either PRC1 or HNRNPK, as well as on fibroblasts in which Xist is deleted on the Xi (XidelXist).
2019-05-03 | GSE116413 | GEO
Project description:JARID2 ChIP-seq profile mediated by Xist was investigated using two separate approaches: In the first approach, we use the TX1072 female embryonic stem cell (ESC) line; this is a genetically polymorphic ESC line derived from a mouse with one X-chromosome (X-chr) from the Mus musculus castaneus (Cast) origin and the other of Mus musculus domesticus, C57BL/6 (BL6) strain origin, containing an inducible promoter on the BL6 Xist allele; differentiation and simultaneous induction of Xist for 4 days results in the inactivation of ONLY the BL6-derived X-chr and coating by JARID2 protein. We analysed this ESC line in the undifferentiated state, in which both X-chrs are active (and not coated by JARID2), and also after 4 days of differentiation under Xist inducible conditions, in which cells display one inactive X-chr from BL6 origin and one Xa of the Cast origin; the presence of SNPs allows for the comparison of the degree and localisation of JARID2 enrichment on the inactive X-chr versus the active X-chr; our results suggests that JARID2 is enriched on the inactive X-chr chromosome-wide and not in confined peaks as elsewhere in the genome; This pattern resembles patterns of Xist enrichment published before (Engreitz et al., 2013). In the second approach, we use a male ESC line that carries an inducible Xist transgene (TG) on chromosome 11 (chr11) (Wutz et al., 2000) in the presence (36:11 WT) or the absence of Eed (36:11 Eed-/-) Induction of Xist TG results in Xist coating and enrichment of JARID2 across the chr11, regardless of the presence of Eed when assess by IF/Xist RNA FISH experiments. We compare inducible and uninducible condition in both the 36:11 WT and Eed-/- ESCs; this way, we could assess the Xist-mediated JARID2 recruitment and the effect of the absence of Eed-/-; Our results shows that Xist induction result in a chromosome-wide enrichment of JARID2 on chr11, in a manner that resembles enrichment on the inactive X-chr; this pattern seem to be independent of EED, showing that in contrast to other regions in the genome where JARID2 occupancy is abrogated in the absence of EED, Xist-mediated JARID2 recruitment is not affected. JARID2 ChIP-seq analysis in differentiating female ESCs (TX1072) and in a male ESC harbouring a Xist transgene on chromosome 11 in WT and Eed-/- genetics settings
2014-01-23 | E-GEOD-52073 | biostudies-arrayexpress
Project description:The inactive X chromosome (Xi) serves as a model to understand gene silencing on a global scale. Here, we perform identification of direct RNA interacting proteins? (iDRiP) to isolate a comprehensive protein interactome for Xist, an RNA required for Xi silencing. We discover multiple classes of interactors, including cohesins, condensins, topoisomerases, RNA helicases, chromatin remodelers and modifiers, which synergistically repress Xi transcription. Inhibiting two or three interactors destabilizes silencing. While Xist attracts some interactors, it repels architectural factors. Xist evicts cohesins from the Xi and directs an Xi-specific chromosome conformation. Upon deleting Xist, the Xi acquires the cohesin-binding and chromosomal architecture of the active X. Our study unveils many layers of Xi repression and demonstrates a central role for RNA in the topological organization of mammalian chromosomes. The RNA-seq data sets generated in this study provide a resource for examining the effects of knockdowns of various Xist-interacting proteins on gene expression. The ChIP-seq data sets provide a comprehensive set of data examining CTCF and cohesion binding the X-chromosome, and the effects of deleting Xist on CTCF and cohesion binding. The Hi-C data is an allele-specific contact map of the X-chromosome higher-order chromatin structure in mouse. RNA-seq in 3 control and 10 knockdown cell lines. ChIP-seq for CTCF, Rad21 and Smc1a in wild-type fibroblasts and Xist-deletion fibroblasts. Hi-C in wild-type and Xist-deletion fibroblasts.
2015-06-19 | E-GEOD-67516 | biostudies-arrayexpress
Project description:3D-topology of DNA in the cell nucleus provides a level of transcription regulation beyond the sequence of the linear DNA. To study the relationship between transcriptional activity and spatial environment of a gene, we have used allele-specific 4C-technology to produce high-resolution topology maps of the active and inactive X-chromosomes in female cells. We found that loci on the active X form multiple long-range interactions, with spatial segregation of active and inactive chromatin. On the inactive X, silenced loci lack preferred interactions, suggesting a unique random organization inside the inactive territory. However, escapees, among which is Xist, are engaged in long-range contacts with each other, enabling identification of novel escapees. Deletion of Xist results in partial re-folding of the inactive X into a conformation resembling the active X, without affecting gene silencing or DNA methylation. Our data point to a role for Xist RNA in shaping the conformation of the inactive X-chromosome independently of transcription. Five or six 4C viewpoints were applied on the mouse female wild type active X-chromosome, the wild type inactive X-chromosome, the conditional Xist X-chromosome and the Xist knock out X-chromosome
2011-06-24 | E-GEOD-29509 | biostudies-arrayexpress