The tandem repeat modules of Xist lncRNA: a swiss army knife for the control of X-chromosome inactivation

X-inactive-specific transcript (Xist) is a long non-coding RNA (lncRNA) essential for X-chromosome inactivation (XCI) in female placental mammals. Thirty years after its discovery, it is still puzzling how this lncRNA triggers major structural and transcriptional changes leading to the stable silencing of an entire chromosome. Recently, a series of studies in mouse cells have uncovered domains of functional specialization within Xist mapping to conserved tandem repeat regions, known as Repeats A-to-F. These functional domains interact with various RNA binding proteins (RBPs) and fold into distinct RNA structures to execute specific tasks in a synergistic and coordinated manner during the inactivation process. This modular organization of Xist is mostly conserved in humans, but recent data point towards differences regarding functional specialization of the tandem repeats between the two species. In this review, we summarize the recent progress on understanding the role of Xist repetitive blocks and their involvement in the molecular mechanisms underlying XCI. We also discuss these findings in the light of the similarities and differences between mouse and human Xist.

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Polycomb complexes in X chromosome inactivation

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2017.0021 ◽

2017 ◽

Vol 372 (1733) ◽

pp. 20170021 ◽

Cited By ~ 35

Author(s):

Neil Brockdorff

Keyword(s):

X Chromosome ◽

Rna Binding ◽

Rna Binding Proteins ◽

X Chromosome Inactivation ◽

Chromosome Inactivation ◽

Histone H2a ◽

Post Translational Modifications ◽

Link Type ◽

Polycomb Proteins ◽

Inactive X Chromosome

Identifying the critical RNA binding proteins (RBPs) that elicit Xist mediated silencing has been a key goal in X inactivation research. Early studies implicated the Polycomb proteins, a family of factors linked to one of two major multiprotein complexes, PRC1 and PRC2 (Wang 2001 Nat. Genet. 28 , 371–375 ( doi:10.1038/ng574 ); Silva 2003 Dev. Cell 4 , 481–495 ( doi:10.1016/S1534-5807(03)00068-6 ); de Napoles 2004 Dev. Cell 7 , 663–676 ( doi:10.1016/j.devcel.2004.10.005 ); Plath 2003 Science 300 , 131–135 ( doi:10.1126/science.1084274 )). PRC1 and PRC2 complexes catalyse specific histone post-translational modifications (PTMs), ubiquitylation of histone H2A at position lysine 119 (H2AK119u1) and methylation of histone H3 at position lysine 27 (H3K27me3), respectively, and accordingly, these modifications are highly enriched over the length of the inactive X chromosome (Xi). A key study proposed that PRC2 subunits bind directly to Xist RNA A-repeat element, a region located at the 5′ end of the transcript known to be required for Xist mediated silencing (Zhao 2008 Science 322 , 750–756 ( doi:10.1126/science.1163045 )). Subsequent recruitment of PRC1 was assumed to occur via recognition of PRC2 mediated H3K27me3 by the CBX subunit of PRC1, as has been shown to be the case at other Polycomb target loci (Cao 2002 Science 298 , 1039–1043 ( doi:10.1126/science.1076997 )). More recently, several reports have questioned aspects of the prevailing view, both in relation to the mechanism for Polycomb recruitment by Xist RNA and the contribution of the Polycomb pathway to Xist mediated silencing. In this article I provide an overview of our recent progress towards resolving these discrepancies. This article is part of the themed issue ‘X-chromosome inactivation: a tribute to Mary Lyon’.

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Local Tandem Repeat Expansion in Xist RNA as a Model for the Functionalisation of ncRNA

Non-Coding RNA ◽

10.3390/ncrna4040028 ◽

2018 ◽

Vol 4 (4) ◽

pp. 28 ◽

Cited By ~ 11

Author(s):

Neil Brockdorff

Keyword(s):

Tandem Repeat ◽

Tandem Repeats ◽

Rna Binding ◽

Rna Binding Proteins ◽

Repeat Expansion ◽

Chromosome Inactivation ◽

Rna Sequences ◽

Xist Rna ◽

Non Coding Rnas ◽

In Cis

Xist, the master regulator of the X chromosome inactivation in mammals, is a 17 kb lncRNA that acts in cis to silence the majority of genes along the chromosome from which it is transcribed. The two key processes required for Xist RNA function, localisation in cis and recruitment of silencing factors, are genetically separable, at least in part. Recent studies have identified Xist RNA sequences and associated RNA-binding proteins (RBPs) that are important for these processes. Notably, several of the key Xist RNA elements correspond to local tandem repeats. In this review, I use examples to illustrate different modes whereby tandem repeat amplification has been exploited to allow orthodox RBPs to confer new functions for Xist-mediated chromosome inactivation. I further discuss the potential generality of tandem repeat expansion in the evolution of functional long non-coding RNAs (lncRNAs).

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Unusual maintenance of X chromosome inactivation predisposes female lymphocytes for increased expression from the inactive X

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1520113113 ◽

2016 ◽

Vol 113 (14) ◽

pp. E2029-E2038 ◽

Cited By ~ 104

Author(s):

Jianle Wang ◽

Camille M. Syrett ◽

Marianne C. Kramer ◽

Arindam Basu ◽

Michael L. Atchison ◽

...

Keyword(s):

B Cells ◽

X Chromosome ◽

Lupus Erythematosus ◽

Rna Binding ◽

Rna Binding Proteins ◽

X Chromosome Inactivation ◽

Chromosome Inactivation ◽

Fish Analysis ◽

Biallelic Expression ◽

Xist Rna

Females have a greater immunological advantage than men, yet they are more prone to autoimmune disorders. The basis for this sex bias lies in the X chromosome, which contains many immunity-related genes. Female mammals use X chromosome inactivation (XCI) to generate a transcriptionally silent inactive X chromosome (Xi) enriched with heterochromatic modifications and XIST/Xist RNA, which equalizes gene expression between the sexes. Here, we examine the maintenance of XCI in lymphocytes from females in mice and humans. Strikingly, we find that mature naïve T and B cells have dispersed patterns of XIST/Xist RNA, and they lack the typical heterochromatic modifications of the Xi. In vitro activation of lymphocytes triggers the return of XIST/Xist RNA transcripts and some chromatin marks (H3K27me3, ubiquitin-H2A) to the Xi. Single-cell RNA FISH analysis of female T cells revealed that the X-linked immunity genes CD40LG and CXCR3 are biallelically expressed in some cells. Using knockout and knockdown approaches, we find that Xist RNA-binding proteins, YY1 and hnRNPU, are critical for recruitment of XIST/Xist RNA back to the Xi. Furthermore, we examined B cells from patients with systemic lupus erythematosus, an autoimmune disorder with a strong female bias, and observed different XIST RNA localization patterns, evidence of biallelic expression of immunity-related genes, and increased transcription of these genes. We propose that the Xi in female lymphocytes is predisposed to become partially reactivated and to overexpress immunity-related genes, providing the first mechanistic evidence to our knowledge for the enhanced immunity of females and their increased susceptibility for autoimmunity.

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X-Chromosome Inactivation and Autosomal Random Monoallelic Expression as “Faux Amis”

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.740937 ◽

2021 ◽

Vol 9 ◽

Author(s):

Vasco M. Barreto ◽

Nadiya Kubasova ◽

Clara F. Alves-Pereira ◽

Anne-Valerie Gendrel

Keyword(s):

X Chromosome ◽

Molecular Mechanisms ◽

Gene Expression Regulation ◽

Phenotypic Diversity ◽

X Chromosome Inactivation ◽

Cellular Level ◽

Chromosome Inactivation ◽

Monoallelic Expression ◽

Distinctive Features ◽

Faux Amis

X-chromosome inactivation (XCI) and random monoallelic expression of autosomal genes (RMAE) are two paradigms of gene expression regulation where, at the single cell level, genes can be expressed from either the maternal or paternal alleles. X-chromosome inactivation takes place in female marsupial and placental mammals, while RMAE has been described in mammals and also other species. Although the outcome of both processes results in random monoallelic expression and mosaicism at the cellular level, there are many important differences. We provide here a brief sketch of the history behind the discovery of XCI and RMAE. Moreover, we review some of the distinctive features of these two phenomena, with respect to when in development they are established, their roles in dosage compensation and cellular phenotypic diversity, and the molecular mechanisms underlying their initiation and stability.

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Low RNA Binding Strength of Human X Chromosome May Explain X Chromosome Inactivation

10.20944/preprints201811.0066.v1 ◽

2018 ◽

Author(s):

Ning Ji ◽

Lifang Yan ◽

Zhixue Song ◽

Shufeng Liu ◽

Chao Liu ◽

...

Keyword(s):

Gene Expression ◽

X Chromosome ◽

Repetitive Sequence ◽

Rna Binding ◽

Repetitive Sequences ◽

X Chromosome Inactivation ◽

Chromosome Inactivation ◽

X Chromosomes ◽

Binding Strength ◽

Egfp Reporter

Two X chromosomes of female mammals randomly inactivate one of paternal or maternal X chromosome in early embryonic development and all the daughter cells produced from these cells retain the same feature of X chromosome inactivation, which is called X chromosome inactivation (XCI). Studying the mechanisms of XCI is important for understanding epigenetic that plays an important role in age-associated diseases. The previous studies have demonstrated that binding of RNAs and DNAs may play a role in activating gene expression. In this paper, our study aims to explore whether the mechanisms of XCI involve the RNA binding strength to X chromosome DNAs. The bioinformatics analyses based on big data were used to analyze the simulated binding strength of RNAs (RNA binding strength) to 23 chromosomes (including X chromosome and 22 human autosomes) and the characteristics of repetitive sequences in the X-inactivation centre. The results revealed that RNA binding strength of the long arm of the X chromosome that is almost entirely inactivated in XCI was significantly lower than that of all autosomes and the short arm of X chromosome, meanwhile the RNA binding strengths of inactivation regions in X chromosome were significantly lower than that of regions escaping from XCI. Different repetitive sequence clusters within the center of XCI presented a cross distribution characteristic. To further prove whether the repetitive sequences in human X chromosome involve in XCI, we cloned long interspersed element (LINE-1, L1) and short interspersed element (Alu) from human Xq13, the center of XCI, and constructed expression vectors carrying sense-antisense combination repetitive sequences (L1s or Alus). Effects of combined L1 or combined Alu sequences on expression of EGFP reporter gene were examined in stably transfected HeLa cells, which simulates the effects of repetitive sequences located on chromosomes. The results of experiments revealed transcribed L1 repetitive sequences activated EGFP reporter gene expression, so did the Alus. The experiment results suggested repetitive sequences activated genes by interaction of transcribed RNAs and DNAs. Since the binding of RNAs and DNAs can activate gene, so the low RNA binding strength of human X chromosome may be one of reasons of XCI. The cross distribution characteristics of different repetitive sequence clusters leading to a cascade of gene activation or gene inactivation may be the reason of transcriptional silencing one of the X chromosomes in female mammals.

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Genes that Escape X Chromosome Inactivation Modulate Sex Differences in Valve Myofibroblasts

Circulation ◽

10.1161/circulationaha.121.054108 ◽

2022 ◽

Author(s):

Brian A. Aguado ◽

Cierra J. Walker ◽

Joseph C. Grim ◽

Megan E. Schroeder ◽

Dilara Batan ◽

...

Keyword(s):

Sex Differences ◽

X Chromosome ◽

Molecular Mechanisms ◽

Smooth Muscle Actin ◽

X Chromosome Inactivation ◽

Chromosome Inactivation ◽

Matrix Stiffness ◽

Alpha Smooth Muscle Actin ◽

Male And Female ◽

Hydrogel Matrix

Background: Aortic valve stenosis (AVS) is a sexually dimorphic disease, with women often presenting with sustained fibrosis and men with more extensive calcification. However, the intracellular molecular mechanisms that drive these clinically important sex differences remain under explored. Methods: Hydrogel scaffolds were designed to recapitulate key aspects of the valve tissue microenvironment and serve as a culture platform for sex-specific valvular interstitial cells (VICs; precursors to pro-fibrotic myofibroblasts). The hydrogel culture system was used to interrogate intracellular pathways involved in sex-dependent VIC-to-myofibroblast activation and deactivation. RNA-sequencing was used to define pathways involved in driving sex-dependent activation. Interventions using small molecule inhibitors and small interfering RNA (siRNA) transfections were performed to provide mechanistic insight into sex-specific cellular responses to microenvironmental cues, including matrix stiffness and exogenously delivered biochemical factors. Results: In both healthy porcine and human aortic valves, female leaflets had higher baseline activation of the myofibroblast marker, alpha-smooth muscle actin (α-SMA), compared to male leaflets. When isolated and cultured, female porcine and human VICs had higher levels of basal α-SMA stress fibers that further increased in response to the hydrogel matrix stiffness, both of which were higher than male VICs. A transcriptomic analysis of male and female porcine VICs revealed Rho-associated protein kinase (RhoA/ROCK) signaling as a potential driver of this sex-dependent myofibroblast activation. Further, we found that genes that escape X-chromosome inactivation, such as BMX and STS (encoding for Bmx non-receptor tyrosine kinase and steroid sulfatase, respectively) partially regulate the elevated female myofibroblast activation via RhoA/ROCK signaling. This finding was confirmed by treating male and female VICs with endothelin-1 and plasminogen activator inhibitor-1, factors that are secreted by endothelial cells and known to drive myofibroblast activation via RhoA/ROCK signaling. Conclusions: Together, in vivo and in vitro results confirm sex-dependencies in myofibroblast activation pathways and implicate genes that escape X-chromosome inactivation in regulating sex differences in myofibroblast activation and subsequent AVS progression. Our results underscore the importance of considering sex as a biological variable to understand the molecular mechanisms of AVS and help guide sex-based precision therapies.

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X-chromosome inactivation: molecular mechanisms and genetic consequences

Trends in Genetics ◽

10.1016/0168-9525(94)90169-4 ◽

1994 ◽

Vol 10 (7) ◽

pp. 230-235 ◽

Cited By ~ 87

Author(s):

Barbara R. Migeon

Keyword(s):

X Chromosome ◽

Molecular Mechanisms ◽

X Chromosome Inactivation ◽

Chromosome Inactivation

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STATISTICAL MECHANICS MODELS FOR X-CHROMOSOME INACTIVATION

Advances in Complex Systems ◽

10.1142/s0219525910002566 ◽

2010 ◽

Vol 13 (03) ◽

pp. 367-376

Author(s):

ANTONIO SCIALDONE ◽

MARIO NICODEMI

Keyword(s):

Statistical Mechanics ◽

X Chromosome ◽

Molecular Mechanisms ◽

X Chromosome Inactivation ◽

Chromosome Inactivation ◽

Regulatory Mechanisms ◽

Random Choice ◽

X Chromosomes ◽

Cellular Processes ◽

Regulatory Processes

We present statistical mechanics models to understand the physical and molecular mechanisms of X-Chromosome Inactivation (XCI), the process whereby a female mammal cell inactivates one of its two X-chromosomes. During XCI, X-chromosomes undergo a series of complex regulatory processes. At the beginning of XCI, the X's recognize and pair, then only one X which is randomly chosen is inactivated. Afterwards, the two X's move to different positions in the cell nucleus according to their different status (active/silenced). Our models illustrate about the still mysterious physical bases underlying all these regulatory steps, i.e., X-chromosome pairing, random choice of inactive X, and "shuttling" of the X's to their post-XCI locations. Our models are based on general and robust thermodynamic roots, and their validity can go beyond XCI, to explain analogous regulatory mechanisms in a variety of cellular processes.

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