Xist exploits three-dimensional chromosome architecture to spread across the X-chromosome
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE46918 | GEO | 2013/05/30
SECONDARY ACCESSION(S): PRJNA203042
REPOSITORIES: GEO
Similar Datasets
Project description:Many large noncoding RNAs (lncRNAs) regulate chromatin, but the mechanisms by which they localize to genomic targets remain unexplored. Here we investigate the localization mechanisms of Xist during X-chromosome inactivation (XCI), a paradigm of lncRNA-mediated chromatin regulation. During the maintenance of XCI, Xist binds broadly across the X-chromosome. During initiation of XCI, Xist initially transfers to distal regions across the X-chromosome that are not defined by specific sequences. Instead, Xist identifies these regions by exploiting the three-dimensional conformation of the X-chromosome. Xist initially accumulates on the periphery of actively transcribed regions and requires its silencing domain to spread across active regions. This suggests a model where Xist coats the entire X-chromosome by searching in three dimensions, modifying chromosome structure, and spreading to newly accessible locations. We examined the genomic localization of the Xist lncRNA using RNA Antisense Purification (RAP) in multiple cell contexts: 1) differentiated female cells (MLFs); 2) a time-course of Xist localization in male embryonic stem (ES) cells where the endogenous Xist promoter is replaced by a tet-inducible one (pSM33); 3) a time-course of Xist localization in differentiating female ES cells (F1 2-1); and 4) wild-type (delXF6) and A-repeat deletion (delSXC9) Xist transgenes incorporated into the Hprt locus under the control of a tet-inducible promoter.
2013-05-30 | E-GEOD-46918 | biostudies-arrayexpress
Project description:During development, transcriptional and chromatin modification changes co-occur but the order and causality of events often remain unclear. We explore the interrelationship of these processes using the paradigm of X-chromosome inactivation (XCI). We initiate XCI in female, mouse embryonic stem cells by inducing Xist expression and monitor changes in transcription and chromatin by allele-specific TT-seq and ChIP-seq respectively. An unprecedented temporal resolution enabled identification of the earliest chromatin alterations during XCI. We demonstrate that HDAC3 interacts with both NCOR1 and NCOR2 and is pre-bound on the X chromosome where it deacetylates histones to promote efficient gene silencing. We also reveal the choreography of polycomb accumulation following Xist RNA coating, with PRC1-associated H2AK119Ub preceding PRC2-associated H3K27me3. Furthermore, polycomb-associated marks accumulate initially at large, intergenic domains and then spreads into genes but only in the context of gene silencing. Our results provide the hierarchy of chromatin events during XCI and demonstrate that some chromatin changes play key roles in mediating transcriptional silencing.
2019-01-17 | PXD011344 | Pride
Project description:The Xist long noncoding RNA (lncRNA) is essential for X-chromosome inactivation (XCI), the process by which mammals compensate for unequal numbers of sex chromosomes. During XCI, Xist coats the future inactive X (Xi) and recruits Polycomb Repressive Complex 2 (PRC2) to the X-inactivation center (Xic). Currently unclear is how Xist spreads silencing on a 150 Mb scale. Here we generate high-resolution maps of Xist binding across a developmental time course using CHART-seq. In female cells undergoing XCI de novo, Xist follows a two-step mechanism in which it initially targets gene-rich islands before spreading to intervening gene-poor domains. Xist is depleted from genes that escape XCI but frequently concentrates near escapee boundaries. Xist binding was linearly proportional to PRC2 density and H3 lysine 27 trimethylation (H3K27me3), suggesting co-migration of Xist and PRC2. Interestingly, when the Xi is acutely stripped of Xist in post-XCI cells, Xist recovers quickly within both gene-rich and -poor domains on a time scale of hours instead of days, suggesting a previously primed Xi chromatin state. We conclude that Xist spreading takes on distinct stage-specific forms: During initial establishment, Xist follows a two-step mechanism, but during maintenance, Xist spreads rapidly to both gene-rich and -poor regions.
2013-11-04 | GSE48649 | GEO
Project description:The Xist long noncoding RNA (lncRNA) is essential for X-chromosome inactivation (XCI), the process by which mammals compensate for unequal numbers of sex chromosomes. During XCI, Xist coats the future inactive X (Xi) and recruits Polycomb Repressive Complex 2 (PRC2) to the X-inactivation center (Xic). Currently unclear is how Xist spreads silencing on a 150 Mb scale. Here we generate high-resolution maps of Xist binding across a developmental time course using CHART-seq. In female cells undergoing XCI de novo, Xist follows a two-step mechanism in which it initially targets gene-rich islands before spreading to intervening gene-poor domains. Xist is depleted from genes that escape XCI but frequently concentrates near escapee boundaries. Xist binding was linearly proportional to PRC2 density and H3 lysine 27 trimethylation (H3K27me3), suggesting co-migration of Xist and PRC2. Interestingly, when the Xi is acutely stripped of Xist in post-XCI cells, Xist recovers quickly within both gene-rich and -poor domains on a time scale of hours instead of days, suggesting a previously primed Xi chromatin state. We conclude that Xist spreading takes on distinct stage-specific forms: During initial establishment, Xist follows a two-step mechanism, but during maintenance, Xist spreads rapidly to both gene-rich and -poor regions. Capture hybridization analysis of RNA targets (CHART) and input samples of (differentiating) mouse embryonic stem (ES) cells and immortalized mouse embryonic fibroblasts (MEF) using paired-end 75 nt reads on Illumina HiSeq2500, with 2 replicates per sample (# of samples). RNA-seq of the same cell lines with 50 nt reads on Illumina HiSeq2000, with 2 replicates per sample (2 samples, 4 datasets total). CHIP-seq: Data from GSE36905 was aligned and processed as CHART-seq samples. Resulting coverage tracks (EZH2/K27me3) are linked directly to GSE48649 (bedGraphs linked below).
2013-11-04 | E-GEOD-48649 | biostudies-arrayexpress
Project description:Xist represents a paradigm for long non-coding RNA function in epigenetic regulation, although how it mediates X-chromosome inactivation (XCI) remains largely unexplained. Multiple Xist-RNA binding proteins have recently been identified, including SPEN/SHARP, whose knockdown has been associated with deficient XCI at multiple loci. Here we demonstrate that SPEN is a key orchestrator of XCI in vivo and unravel its mechanism of action. We show that SPEN is essential for initiating gene silencing on the X chromosome in preimplantation mouse embryos and embryonic stem cells. On the other hand, SPEN is dispensable for maintenance of XCI in neural progenitor cells, although it significantly dampens expression of genes that escape from XCI. During initiation of XCI, we show by live-cell imaging and CUT&RUN approaches that SPEN is immediately recruited to the X chromosome upon Xist up-regulation, where it is targeted to enhancers and promoters of actively transcribed genes. SPEN rapidly disengages from chromatin once silencing is accomplished, implying a need for active transcription to tether it to chromatin. We define SPEN’s SPOC (SPEN paralog and ortholog C-terminal) domain as a major effector of SPEN’s gene silencing function, and show that artificial tethering of SPOC to Xist RNA is sufficient to mediate X-linked gene silencing. We identify SPOC’s protein partners which include NCOR/SMRT, the m6A RNA methylation machinery, the NuRD complex, RNA polymerase II and factors involved in regulation of transcription initiation and elongation. We propose that SPEN acts as a molecular integrator for initiation of XCI, bridging Xist RNA with the transcription machinery as well as nucleosome remodelers and histone deacetylases, at active enhancers and promoters.
2021-04-19 | PXD015699 | Pride
Project description:Nuclear compartments play diverse roles in regulating gene expression, yet the molecular forces and components driving compartment formation are not well understood. Studying how the lncRNA Xist establishes the inactive-X-chromosome (Xi)-compartment, we found that the Xist RNA-binding-proteins PTBP1, MATR3, TDP43, and CELF1 form a condensate to create an Xi-domain that can be sustained in the absence of Xist. The E-repeat-sequence of Xist serves a multivalent binding-platform for these proteins. Without the E-repeat, Xist initially coats the X-chromosome during XCI onset but subsequently disperses across the nucleus with loss of gene silencing. Recruitment of PTBP1, MATR3, TDP-43 or CELF1 to E-Xist rescues these phenotypes, and requires both self-association of MATR3 and TDP-43 and a heterotypic PTBP1-MATR3-interaction. Together, our data reveal that Xist sequesters itself within the Xi-territory and perpetuates gene silencing by seeding a protein-condensate. Our findings uncover an unanticipated mechanism for epigenetic memory and elucidate the interplay between RNA and RNA-binding-proteins in creating compartments for gene regulation.
2020-09-03 | GSE137305 | GEO
Project description:X-chromosome inactivation (XCI) serves as a paradigm for RNA-mediated regulation of gene expression, wherein the long non-coding RNA XIST spreads across the X-chromosome in cis to mediate chromosome-wide gene silencing. In female naïve human pluripotent stem cells (hPSCs), XIST is in a dispersed configuration and XCI does not occur, raising questions about XIST’s function. We found that XIST spreads across the X-chromosome and induces dampening of X-linked gene expression in naïve hPSCs. Surprisingly, XIST also targets specific autosomal regions, where it induces repressive chromatin changes and gene expression dampening. Thereby, XIST equalizes X-linked gene dosage between male and female cells while inducing differences on autosomes. The dispersed Xist configuration and autosomal localization also occur transiently during XCI initiation in mouse PSCs. Together, our study identifies XIST as the regulator of X-chromosome dampening, uncovers an evolutionarily conserved trans-acting role of XIST/Xist, and reveals a correlation between XIST/Xist dispersal and autosomal targeting. Surprisingly, XIST also targets specific autosomal regions, where it induces repressive chromatin changes and gene expression dampening. Thereby, XIST equalizes X-linked gene dosage between male and female cells while inducing differences on autosomes. Dispersed Xist configuration and autosomal localization also occur transiently during XCI initiation in mouse PSCs. Together, our results identify XIST as the regulator of X-chromosome dampening, uncover an evolutionarily conserved trans-acting role of XIST/Xist, and reveal a correlation between XIST/Xist dispersal and autosomal targeting.
2023-12-15 | GSE241443 | GEO
Project description:X-chromosome inactivation (XCI) serves as a paradigm for RNA-mediated regulation of gene expression, wherein the long non-coding RNA XIST spreads across the X-chromosome in cis to mediate chromosome-wide gene silencing. In female naïve human pluripotent stem cells (hPSCs), XIST is in a dispersed configuration and XCI does not occur, raising questions about XIST’s function. We found that XIST spreads across the X-chromosome and induces dampening of X-linked gene expression in naïve hPSCs. Surprisingly, XIST also targets specific autosomal regions, where it induces repressive chromatin changes and gene expression dampening. Thereby, XIST equalizes X-linked gene dosage between male and female cells while inducing differences on autosomes. The dispersed Xist configuration and autosomal localization also occur transiently during XCI initiation in mouse PSCs. Together, our study identifies XIST as the regulator of X-chromosome dampening, uncovers an evolutionarily conserved trans-acting role of XIST/Xist, and reveals a correlation between XIST/Xist dispersal and autosomal targeting. Surprisingly, XIST also targets specific autosomal regions, where it induces repressive chromatin changes and gene expression dampening. Thereby, XIST equalizes X-linked gene dosage between male and female cells while inducing differences on autosomes. Dispersed Xist configuration and autosomal localization also occur transiently during XCI initiation in mouse PSCs. Together, our results identify XIST as the regulator of X-chromosome dampening, uncover an evolutionarily conserved trans-acting role of XIST/Xist, and reveal a correlation between XIST/Xist dispersal and autosomal targeting.
2023-12-15 | GSE241442 | GEO
Project description:X-chromosome inactivation (XCI) serves as a paradigm for RNA-mediated regulation of gene expression, wherein the long non-coding RNA XIST spreads across the X-chromosome in cis to mediate chromosome-wide gene silencing. In female naïve human pluripotent stem cells (hPSCs), XIST is in a dispersed configuration and XCI does not occur, raising questions about XIST’s function. We found that XIST spreads across the X-chromosome and induces dampening of X-linked gene expression in naïve hPSCs. Surprisingly, XIST also targets specific autosomal regions, where it induces repressive chromatin changes and gene expression dampening. Thereby, XIST equalizes X-linked gene dosage between male and female cells while inducing differences on autosomes. The dispersed Xist configuration and autosomal localization also occur transiently during XCI initiation in mouse PSCs. Together, our study identifies XIST as the regulator of X-chromosome dampening, uncovers an evolutionarily conserved trans-acting role of XIST/Xist, and reveals a correlation between XIST/Xist dispersal and autosomal targeting. Surprisingly, XIST also targets specific autosomal regions, where it induces repressive chromatin changes and gene expression dampening. Thereby, XIST equalizes X-linked gene dosage between male and female cells while inducing differences on autosomes. Dispersed Xist configuration and autosomal localization also occur transiently during XCI initiation in mouse PSCs. Together, our results identify XIST as the regulator of X-chromosome dampening, uncover an evolutionarily conserved trans-acting role of XIST/Xist, and reveal a correlation between XIST/Xist dispersal and autosomal targeting.
2023-12-15 | GSE241438 | GEO
Project description:At initiation of X chromosome inactivation (XCI), Xist is monoallelically upregulated from the future inactive X (Xi) chromosome, overcoming repression by its antisense transcript Tsix. Xist recruits various chromatin remodelers, amongst them SPEN, which are involved in silencing of X-linked genes in cis and establishment of the Xi. Here, we show that SPEN plays an important role in the initiation of XCI. Spen null female mouse embryonic stem cells (ESCs) are defective in Xist upregulation upon differentiation. We find that Xist-mediated SPEN recruitment to the Xi chromosome happens very early in XCI, and that SPEN-mediated silencing of the Tsix promoter is required for Xist upregulation. Accordingly, failed Xist upregulation in Spen-/- ESCs can be rescued by concomitant removal of Tsix. These findings indicate that SPEN is not only required for the establishment of the Xi, but is also crucial in the initiation of the XCI process.
2021-09-30 | GSE163320 | GEO