A primate-specific retroviral enhancer wires the XACT lncRNA into the core pluripotency network in human

AbstractTransposable elements (TEs) have been of paramount importance in shaping genomic and epigenomic landscapes of their hosts and in driving the expansion of gene regulatory networks during mammalian evolution. They are found in nearly all long non-coding RNAs (lncRNAs) and have promoted their evolution and function, often in a species- and tissue-specific manner. X-chromosome inactivation (XCI) is an essential process that relies on several TE-enriched lncRNAs. While XCI is conserved across species, one striking difference between human and mouse is the existence of XACT (X active coating transcript), a human-specific lncRNA that coats active X chromosomes in pluripotent cells and may oppose X chromosome silencing in this context. Here, we explore how different families of TEs have contributed to shaping the XACT locus and how they couple its expression to pluripotency in humans. Through a combination of sequence analysis across primates, transcriptional interference and genome editing in human embryonic stem cells (hESCs), we identify a critical enhancer for the transcriptional regulation of the XACT locus that evolved from an ancestral group (LTR48B/ERV1) of mammalian endogenous retroviruses (ERVs), prior to the emergence of XACT. Furthermore, we show that this ancient ERV was hijacked by evolutionarily younger hominoid-specific ERVs that gave rise to the promoter of XACT, thus wiring its expression to the pluripotency network. This work illustrates how retroviral-derived sequences may intervene in species-specific regulatory pathways.

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A primate-specific retroviral enhancer wires the XACT lncRNA into the core pluripotency network in humans

Nature Communications ◽

10.1038/s41467-019-13551-1 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 5

Author(s):

Miguel Casanova ◽

Madeleine Moscatelli ◽

Louis Édouard Chauvière ◽

Christophe Huret ◽

Julia Samson ◽

...

Keyword(s):

Gene Regulatory Networks ◽

Regulatory Networks ◽

Endogenous Retroviruses ◽

X Chromosomes ◽

Regulatory Pathways ◽

Gene Regulatory ◽

Non Coding Rnas ◽

Specific Regulation ◽

Species Specific ◽

And Function

AbstractTransposable elements (TEs) have been proposed to play an important role in driving the expansion of gene regulatory networks during mammalian evolution, notably by contributing to the evolution and function of long non-coding RNAs (lncRNAs). XACT is a primate-specific TE-derived lncRNA that coats active X chromosomes in pluripotent cells and may contribute to species-specific regulation of X-chromosome inactivation. Here we explore how different families of TEs have contributed to shaping the XACT locus and coupling its expression to pluripotency. Through a combination of sequence analysis across primates, transcriptional interference, and genome editing, we identify a critical enhancer for the regulation of the XACT locus that evolved from an ancestral group of mammalian endogenous retroviruses (ERVs), prior to the emergence of XACT. This ERV was hijacked by younger hominoid-specific ERVs that gave rise to the promoter of XACT, thus wiring its expression to the pluripotency network. This work illustrates how retroviral-derived sequences may intervene in species-specific regulatory pathways.

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Species-specific differences in X chromosome inactivation in mammals

Reproduction ◽

10.1530/rep-13-0173 ◽

2013 ◽

Vol 146 (4) ◽

pp. R131-R139 ◽

Cited By ~ 18

Author(s):

Takashi Sado ◽

Takehisa Sakaguchi

Keyword(s):

X Chromosome ◽

Molecular Basis ◽

Embryonic Stem ◽

X Chromosome Inactivation ◽

X Inactivation ◽

Chromosome Inactivation ◽

X Chromosomes ◽

Inactive X Chromosome ◽

Species Specific ◽

Genetically Manipulated

In female mammals, the dosage difference in X-linked genes between XX females and XY males is compensated for by inactivating one of the two X chromosomes during early development. Since the discovery of the X inactive-specific transcript (XIST) gene in humans and its subsequent isolation of the mouse homolog, Xist, in the early 1990s, the molecular basis of X chromosome inactivation (X-inactivation) has been more fully elucidated using genetically manipulated mouse embryos and embryonic stem cells. Studies on X-inactivation in other mammals, although limited when compared with those in the mice, have revealed that, while their inactive X chromosome shares many features with those in the mice, there are marked differences in not only some epigenetic modifications of the inactive X chromosome but also when and how X-inactivation is initiated during early embryonic development. Such differences raise the issue about what extent of the molecular basis of X-inactivation in the mice is commonly shared among others. Recognizing similarities and differences in X-inactivation among mammals may provide further insight into our understanding of not only the evolutionary but also the molecular aspects for the mechanism of X-inactivation. Here, we reviewed species-specific differences in X-inactivation and discussed what these differences may reveal.

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Zscan4c activates endogenous retrovirus MERVL and cleavage embryo genes

Nucleic Acids Research ◽

10.1093/nar/gkz594 ◽

2019 ◽

Cited By ~ 4

Author(s):

Weiyu Zhang ◽

Fuquan Chen ◽

Ruiqing Chen ◽

Dan Xie ◽

Jiao Yang ◽

...

Keyword(s):

Developmental Stages ◽

Embryonic Stem ◽

Endogenous Retrovirus ◽

Endogenous Retroviruses ◽

Enhancer Activity ◽

Cell Cleavage ◽

Cell Embryo ◽

And Function ◽

Scan Domain

AbstractEndogenous retroviruses (ERVs) contribute to ∼10 percent of the mouse genome. They are often silenced in differentiated somatic cells but differentially expressed at various embryonic developmental stages. A minority of mouse embryonic stem cells (ESCs), like 2-cell cleavage embryos, highly express ERV MERVL. However, the role of ERVs and mechanism of their activation in these cells are still poorly understood. In this study, we investigated the regulation and function of the stage-specific expressed ERVs, with a particular focus on the totipotency marker MT2/MERVL. We show that the transcription factor Zscan4c functions as an activator of MT2/MERVL and 2-cell/4-cell embryo genes. Zinc finger domains of Zscan4c play an important role in this process. In addition, Zscan4c interacts with MT2 and regulates MT2-nearby 2-cell/4-cell genes through promoting enhancer activity of MT2. Furthermore, MT2 activation is accompanied by enhanced H3K4me1, H3K27ac, and H3K14ac deposition on MT2. Zscan4c also interacts with GBAF chromatin remodelling complex through SCAN domain to further activate MT2 enhancer activity. Taken together, we delineate a previously unrecognized regulatory axis that Zscan4c interacts with and activates MT2/MERVL loci and their nearby genes through epigenetic regulation.

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Retroviral Transcriptional Regulation and Embryonic Stem Cells: War and Peace

Molecular and Cellular Biology ◽

10.1128/mcb.01293-14 ◽

2014 ◽

Vol 35 (5) ◽

pp. 770-777 ◽

Cited By ~ 63

Author(s):

Sharon Schlesinger ◽

Stephen P. Goff

Keyword(s):

Dna Sequences ◽

Regulatory Networks ◽

Es Cells ◽

Expression Patterns ◽

Embryonic Stem ◽

Cell Types ◽

Endogenous Retroviruses ◽

Regulatory Sequences ◽

Cellular Genes ◽

Wide Range

Retroviruses have evolved complex transcriptional enhancers and promoters that allow their replication in a wide range of tissue and cell types. Embryonic stem (ES) cells, however, characteristically suppress transcription of proviruses formed after infection by exogenous retroviruses and also of most members of the vast array of endogenous retroviruses in the genome. These cells have unusual profiles of transcribed genes and are poised to make rapid changes in those profiles upon induction of differentiation. Many of the transcription factors in ES cells control both host and retroviral genes coordinately, such that retroviral expression patterns can serve as markers of ES cell pluripotency. This overlap is not coincidental; retrovirus-derived regulatory sequences are often used to control cellular genes important for pluripotency. These sequences specify the temporal control and perhaps “noisy” control of cellular genes that direct proper cell gene expression in primitive cells and their differentiating progeny. The evidence suggests that the viral elements have been domesticated for host needs, reflecting the wide-ranging exploitation of any and all available DNA sequences in assembling regulatory networks.

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Histone H3 Lysine 36 Trimethylation Is Established over theXistPromoter by AntisenseTsixTranscription and Contributes to RepressingXistExpression

Molecular and Cellular Biology ◽

10.1128/mcb.00561-15 ◽

2015 ◽

Vol 35 (22) ◽

pp. 3909-3920 ◽

Cited By ~ 15

Author(s):

Tatsuya Ohhata ◽

Mika Matsumoto ◽

Martin Leeb ◽

Shinwa Shibata ◽

Satoshi Sakai ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

X Chromosome ◽

Antisense Rna ◽

Histone H3 ◽

Embryonic Stem ◽

X Chromosome Inactivation ◽

Chromosome Inactivation ◽

X Chromosomes ◽

Histone H3.3

One of the two X chromosomes in female mammals is inactivated by the noncodingXistRNA. In mice, X chromosome inactivation (XCI) is regulated by the antisense RNATsix, which repressesXiston the active X chromosome. In the absence ofTsix, PRC2-mediated histone H3 lysine 27 trimethylation (H3K27me3) is established over theXistpromoter. Simultaneous disruption ofTsixand PRC2 leads to derepression ofXistand in turn silencing of the single X chromosome in male embryonic stem cells. Here, we identified histone H3 lysine 36 trimethylation (H3K36me3) as a modification that is recruited byTsixcotranscriptionally and extends over theXistpromoter. Reduction of H3K36me3 by expression of a mutated histone H3.3 with a substitution of methionine for lysine at position 36 causes a significant derepression ofXist. Moreover, depletion of the H3K36 methylaseSetd2leads to upregulation ofXist, suggesting H3K36me3 as a modification that contributes to the mechanism ofTsixfunction in regulating XCI. Furthermore, we found that reduction of H3K36me3 does not facilitate an increase in H3K27me3 over theXistpromoter, indicating that additional mechanisms exist by whichTsixblocks PRC2 recruitment to theXistpromoter.

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From teratocarcinomas to embryonic stem cells

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2002.1058 ◽

2002 ◽

Vol 357 (1420) ◽

pp. 405-417 ◽

Cited By ~ 174

Author(s):

Peter W. Andrews

Keyword(s):

Stem Cells ◽

Regulatory Networks ◽

Adult Stem Cells ◽

Es Cells ◽

Embryonic Stem ◽

Early Embryo ◽

Clear Understanding ◽

Regulatory Pathways ◽

Wide Range ◽

Clinical Potential

The recent derivation of human embryonic stem (ES) cell lines, together with results suggesting an unexpected degree of plasticity in later, seemingly more restricted, stem cells (so–called adult stem cells), have combined to focus attention on new opportunities for regenerative medicine, as well as for understanding basic aspects of embryonic development and diseases such as cancer. Many of the ideas that are now discussed have a long history and much has been underpinned by the earlier studies of teratocarcinomas, and their embryonal carcinoma (EC) stem cells, which present a malignant surrogate for the normal stem cells of the early embryo. Nevertheless, although the potential of EC and ES cells to differentiate into a wide range of tissues is now well attested, little is understood of the key regulatory mechanisms that control their differentiation. Apart from the intrinsic biological interest in elucidating these mechanisms, a clear understanding of the molecular process involved will be essential if the clinical potential of these cells is to be realized. The recent observations of stem–cell plasticity suggest that perhaps our current concepts about the operation of cell regulatory pathways are inadequate, and that new approaches for analysing complex regulatory networks will be essential.

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X-chromosome upregulation is dynamically linked to the X-inactivation state

10.1101/2020.07.06.189787 ◽

2020 ◽

Cited By ~ 1

Author(s):

Antonio Lentini ◽

Christos Coucoravas ◽

Nathanael Andrews ◽

Martin Enge ◽

Qiaolin Deng ◽

...

Keyword(s):

Stem Cells ◽

X Chromosome ◽

Embryonic Stem ◽

Mrna Levels ◽

X Chromosomes ◽

Dosage Balance ◽

Mechanistic Basis ◽

Early Mouse ◽

Key Events

AbstractMammalian X-chromosome dosage balance is regulated by X-chromosome inactivation (XCI) and X-chromosome upregulation (XCU), but the dynamics of XCU as well as the interplay between the two mechanisms remain poorly understood. Here, we mapped XCU throughout early mouse embryonic development at cellular and allelic resolution, revealing sex- and lineage-specific dynamics along key events in X-chromosome regulation. Our data show that XCU is linearly proportional to the degree of XCI, indicating that dosage compensation ensues based on mRNA levels rather than number of active X chromosomes. In line with this, we reveal that the two active X chromosomes in female naïve embryonic stem cells are not hyperactive as previously thought. In all lineages, XCU was underlain by increased transcriptional burst frequencies, providing a mechanistic basis in vivo. Together, our results demonstrate unappreciated flexibility of XCU in balancing X-chromosome expression, and we propose a general model for allelic dosage balance, applicable for wider mechanisms of transcriptional regulation.

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A system for imaging the regulatory noncoding Xist RNA in living mouse embryonic stem cells

Molecular Biology of the Cell ◽

10.1091/mbc.e11-02-0146 ◽

2011 ◽

Vol 22 (14) ◽

pp. 2634-2645 ◽

Cited By ~ 33

Author(s):

Karen Ng ◽

Nathalie Daigle ◽

Aurélien Bancaud ◽

Tatsuya Ohhata ◽

Peter Humphreys ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

X Chromosome ◽

Single Point ◽

Embryonic Stem ◽

Mouse Embryonic Stem Cells ◽

Chromatin Domain ◽

Dynamic Binding ◽

X Chromosomes ◽

Xist Rna

In mammals, silencing of one of the two X chromosomes in female cells compensates for the different number of X chromosomes between the sexes. The noncoding Xist RNA initiates X chromosome inactivation. Xist spreads from its transcription site over the X chromosome territory and triggers the formation of a repressive chromatin domain. To understand localization of Xist over one X chromosome we aimed to develop a system for investigating Xist in living cells. Here we report successful visualization of transgenically expressed MS2‑tagged Xist in mouse embryonic stem cells. Imaging of Xist during an entire cell cycle shows that Xist spreads from a single point to a steady state when the chromosome is covered with a constant amount of Xist. Photobleaching experiments of the established Xist cluster indicate that chromosome‑bound Xist is dynamic and turns over on the fully Xist covered chromosome. It appears that in interphase the loss of bound Xist and newly produced Xist are in equilibrium. We also show that the turnover of bound Xist requires transcription, and Xist binding becomes stable when transcription is inhibited. Our data reveal a strategy for visualizing Xist and indicate that spreading over the chromosome might involve dynamic binding and displacement.

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Integrated analysis of Xist upregulation and X-chromosome inactivation with single-cell and single-allele resolution

Nature Communications ◽

10.1038/s41467-021-23643-6 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Guido Pacini ◽

Ilona Dunkel ◽

Norbert Mages ◽

Verena Mutzel ◽

Bernd Timmermann ◽

...

Keyword(s):

Gene Silencing ◽

Single Cell ◽

X Chromosome ◽

Embryonic Stem ◽

X Chromosome Inactivation ◽

Chromosome Inactivation ◽

Cellular Heterogeneity ◽

Integrated Analysis ◽

X Chromosomes ◽

Multiple Levels

AbstractTo ensure dosage compensation between the sexes, one randomly chosen X chromosome is silenced in each female cell in the process of X-chromosome inactivation (XCI). XCI is initiated during early development through upregulation of the long non-coding RNA Xist, which mediates chromosome-wide gene silencing. Cell differentiation, Xist upregulation and gene silencing are thought to be coupled at multiple levels to ensure inactivation of exactly one out of two X chromosomes. Here we perform an integrated analysis of all three processes through allele-specific single-cell RNA-sequencing. Specifically, we assess the onset of random XCI in differentiating mouse embryonic stem cells, and develop dedicated analysis approaches. By exploiting the inter-cellular heterogeneity of XCI onset, we identify putative Xist regulators. Moreover, we show that transient Xist upregulation from both X chromosomes results in biallelic gene silencing right before transitioning to the monoallelic state, confirming a prediction of the stochastic model of XCI. Finally, we show that genetic variation modulates the XCI process at multiple levels, providing a potential explanation for the long-known X-controlling element (Xce) effect, which leads to preferential inactivation of a specific X chromosome in inter-strain crosses. We thus draw a detailed picture of the different levels of regulation that govern the initiation of XCI. The experimental and computational strategies we have developed here will allow us to profile random XCI in more physiological contexts, including primary human cells in vivo.

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A RIF1 and KAP1-based, double-bookmarking system generates a toggle switch that stabilises the identities of the active and inactive X chromosomes during random X inactivation in mouse

10.1101/2020.06.04.133512 ◽

2020 ◽

Author(s):

Elin Enervald ◽

Rossana Foti ◽

Lynn Marie Powell ◽

Agnieszka Piszczek ◽

Sara C.B. Buonomo

Keyword(s):

X Chromosome ◽

Embryonic Stem ◽

X Inactivation ◽

Allelic Association ◽

Toggle Switch ◽

X Chromosomes ◽

Non Coding Rna ◽

The Future ◽

Long Non Coding Rna ◽

Stochastic Event

ABSTRACTDosage compensation for the X chromosome-linked genes in female placental mammals is achieved through the random silencing of one of the two X chromosomes. The onset of random X inactivation in mouse embryos and in differentiating embryonic stem cells requires the switch from a symmetric state, where both X chromosomes are equivalent, to an asymmetric state, where the identity of the future inactive and active X chromosomes are assigned. This “choice”, initiated by a stochastic event, needs to evolve into a stable and transmissible state. The transition from bi- to mono-allelic expression of the long non-coding RNA Tsix is thought to be one of the initial events breaking the symmetry of the two X chromosomes. Here we show that the asymmetric expression of Tsix triggers in turn the switch of RIF1 association with the Xist promoter from dynamic and symmetric to stable and asymmetric (on the future inactive X). On the future inactive X, RIF1 then plays an essential role in the upregulation of Xist, thus initiating the consolidation and stable transmission of the identity of the inactive X. Tsix-dependent exclusion of RIF1 from the future active X chromosome in turn permits the association of KAP1 with the Xist promoter, thus marking the future active X chromosome. Timely mono-allelic association of KAP1 is important for a stable choice and for X inactivation. We present here a double-bookmarking system, based on the mutually exclusive relationships of Tsix and RIF1, and RIF1 and KAP1. This system coordinates the identification of the active and inactive X chromosomes and initiates a self-sustaining loop that transforms an initially stochastic event into a stably inherited asymmetric X chromosome state.

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