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Autoimmune diseases appear to have multiple contributing factors including genetics, epigenetics, environmental factors, and aging. The predominance of females among patients with autoimmune diseases suggests possible involvement of the X chromosome and X chromosome inactivation. X chromosome inactivation is an epigenetic event resulting in multiple levels of control for modulation of the expression of X-linked genes in normal female cells such that there remains only one active X chromosome in the cell. The extent of this control is unique among the chromosomes and has the potential for problems when regulation is disrupted. Here we discuss the X chromosome inactivation process and how the X chromosome and X chromosome inactivation may be involved in development of autoimmune disorders. [ABSTRACT FROM AUTHOR]

Mammals inactivate one of the two female X chromosomes to compensate for the unequal copy number of X-linked genes between males and females. This process of X inactivation entails the silencing of one X chromosome in a developmentally regulated manner. In this work, we review recent findings in X inactivation and discuss how these advance the mechanistic understanding. Recent results provide an insight how the cell counts and chooses the appropriate number of X chromosomes to inactivate, how chromosome-wide gene repression is coordinated and how a stable inactive X chromosome is established. Key components of this complex regulatory system have now been identified and provide entry points for understanding epigenetic regulation in mammals. A majority of the data has been obtained from studying mice. It is presently not clear how general these findings can be applied to other mammalian species. We try to assess this aspect from data, which has become available. [ABSTRACT FROM AUTHOR]

The DNA methylation status of human X chromosomes from male and female neutrophils was identified by high-throughput sequencing of HpaII and MspI digested fragments. In the intergenic and intragenic regions on the X chromosome, the sites outside CpG islands were heavily hypermethylated to the same degree in both genders. Nearly half of X chromosome promoters were either hypomethylated or hypermethylated in both females and males. Nearly one third of X chromosome promoters were a mixture of hypomethylated and heterogeneously methylated sites in females and were hypomethylated in males. Thus, a large fraction of genes that are silenced on the inactive X chromosome are hypomethylated in their promoter regions. These genes frequently belong to the evolutionarily younger strata of the X chromosome. The promoters that were hypomethylated at more than two sites contained most of the genes that escaped silencing on the inactive X chromosome. The overall levels of expression of X-linked genes were indistinguishable in females and males, regardless of the methylation state of the inactive X chromosome. Thus, in addition to DNA methylation, other factors are involved in the fine tuning of gene dosage compensation in neutrophils. [ABSTRACT FROM AUTHOR]

We report on the characterization of the Peromyscus melanophrys karyotype and sex chromosome system. Classic studies reported the sex chromosome system of this species may be as complex as an X1X1X2X2/X1X2Y1Y2 and provided conflicting identification of the X chromosome. Using Peromyscus maniculatus chromosome paints, we have positively identified the sex chromosomes and clarified the sex determining system that once perplexed Peromyscus researchers. The sex chromosomes are characterized by a unique autosomal translocation of DNA shared between both the X and Y chromosomes. The translocated material is late replicating and heterochromatic yet retains the active chromatin conformation. Thus, autosomal regions derived from translocations involving repeat-rich material may retain some epigenetic marks specific to the sex chromosomes despite loss of epigenetic silencing activity. [ABSTRACT FROM AUTHOR]

Epigenetic regulation is important for stable maintenance of cell identity. For continued function of organs and tissues, illegitimate changes in cell identity must be avoided. Failure to do so can trigger tumour development and disease. How epigenetic patterns are established during cell differentiation has been explored by studying model systems such as X inactivation. Mammals balance the X-linked gene dosage between the sexes by silencing of one of the two X chromosomes in females. This is initiated by expression of the non-coding X-inactive specific transcript (Xist) RNA and depends on specific cellular contexts, in which essential silencing factors are expressed. Normally X inactivation is initiated in early embryogenesis, but recent reports identified instances where Xist is expressed and can initiate gene repression. Here we describe the features that characterize the cellular permissivity to initiation of X inactivation and note that these can also occur in cancer cells and in specific haematopoietic progenitors. We propose that embryonic pathways for epigenetic regulation are re-established in adult progenitor cells and tumour cells. Understanding their reactivation will deepen our understanding of tumourigenesis and may be exploited for cancer therapy. [ABSTRACT FROM AUTHOR]

The article presents a study regarding the role of RNA interference (RNAi) in X inactivation process. It states that RNAi machinery was elaborately involved in yeast centrometric chromatin thru small RNA generated from antisense and pericentric sense non-coding RNA and RNA-induced transcriptional silencing (RITS) Furthermore, the study also suggests the involvement of RNAi pathway or small double-stranded RNA (dsRNA) in chromosome stability and heterochromatin formation in mammals.

X chromosome inactivation (XCI) is mediated by the non-coding RNA Xist, which directs chromatin modification and gene silencing in cis. The RNA binding protein SPEN and associated corepressors have a central role in Xist-mediated gene silencing. Other silencing factors, notably the Polycomb system, have been reported to function downstream of SPEN. In recent work, we found that SPEN has an additional role in correct localization of Xist RNA in cis , indicating that its contribution to chromatin-mediated gene silencing needs to be reappraised. Making use of a SPEN separation-of-function mutation, we show that SPEN and Polycomb pathways, in fact, function in parallel to establish gene silencing. We also find that differentiation-dependent recruitment of the chromosomal protein SmcHD1 is required for silencing many X-linked genes. Our results provide important insights into the mechanism of X inactivation and the coordination of chromatin-based gene regulation with cellular differentiation and development. [Display omitted] • SPEN and PRC1 pathways function in parallel to establish gene silencing by Xist RNA • Removal of both pathways abolishes gene silencing and disrupts Xist localization • Differentiation is required to achieve maximal X-linked gene silencing • Differentiation-dependent SmcHD1 recruitment contributes to silencing of many genes Bowness et al. examine molecular pathways harnessed by the long non-coding RNA Xist for gene silencing during X chromosome inactivation. They find that Xist establishes silencing via SPEN and Polycomb protein complexes acting in parallel and that maximal silencing requires cellular differentiation and recruitment of a third protein, SmcHD1. [ABSTRACT FROM AUTHOR]

In mammals, X-chromosome dosage compensation is achieved by inactivating one of the two X chromosomes in females. In mice, X inactivation is initially imprinted, with inactivation of the paternal X (Xp) chromosome occurring during preimplantation development. One theory is that the Xp is preinactivated in female embryos, because of its previous silence during meiosis in the male germ line. The extent to which the Xp is active after fertilization and the exact time of onset of X-linked gene silencing have been the subject of debate. We performed a systematic, single-cell transcriptional analysis to examine the activity of the Xp chromosome for a panel of X-linked genes throughout early preimplantation development in the mouse. Rather than being preinactivated, we found the Xp to be fully active at the time of zygotic gene activation, with silencing beginning from the 4-cell stage onward. X-inactivation patterns were, however, surprisingly diverse between genes. Some loci showed early onset (4-8-cell stage) of X inactivation, and some showed extremely late onset (postblastocyst stage), whereas others were never fully inactivated. Thus, we show that silencing of some X-chromosomal regions occurs outside of the usual time window and that escape from X inactivation can be highly lineage specific. These results reveal that imprinted X inactivation in mice is far less concerted than previously thought and highlight the epigenetic diversity underlying the dosage compensation process during early mammalian development. [ABSTRACT FROM AUTHOR]

Sex determination varies greatly among animal species including mammals. Although two identical (XX) or distinct (XY) gonosomes are found in all species of the latter, the ratio of sex-specific content varies considerably ranging from a small minority in monotremes to more than 90% in primates. However, the similarities are strong enough to allow us to study those mechanisms in comparison with each other and obtain useful information on the evolution of sex determination. The paper focuses on the basic principles of sex determination in mammals, stressing the common evolutionary background behind the striking similarities observed even among phenotypically and geographically distinct species. Sequencing and comparative genetic data seem to suggest that a common evolutionary origin for all mammalian sex determination strategies is most probable. Nevertheless, original research data is still required for the understanding of the complex underlying mechanisms. [ABSTRACT FROM AUTHOR]

The mammalian olfactory system utilizes three large receptor families: the olfactory receptors (ORs) of the main nose and the vomeronasal type-1 and type-2 receptor genes (V1Rs and V2Rs) of the vomeronasal organ. We find that these loci are among the most long interspersed nuclear element (LINE)-dense regions of mammalian genomes. We investigate two evolutionary models to account for this cohabitation. First, we investigate an adaptive selection model, in which LINEs have contributed to expansions of mouse V1R repertoires. We find that even evolutionarily stable V1R loci are exceptionally LINE-rich compared to other genome loci, including loci containing other large gene clusters. Also, a more detailed analysis of specific V1R duplications does not reveal LINE patterns predicted by common LINE-mediated duplication mechanisms. Next, we investigate neutral models, in which LINEs were tolerated by, but not advantageous for, surrounding V1R genes. We find that V1R loci are exceptionally LINE-rich compared to other regions of similar AT base composition, and that duplicated V1R gene blocks are generally depleted of LINE elements, suggesting that these loci did not become densely populated with LINEs simply as a consequence of targeted integration or passive multiplication along with the genes. Finally, we show that individual LINE repeats of a given age at V1R, V2R, and OR loci exhibit a significantly longer average length than at other autosomal loci, suggesting a reduced tendency for these LINEs to be disrupted. We speculate that LINEs at V1R, V2R, and OR loci might be selectively retained because they contribute to allelic regulation of these three gene families. [ABSTRACT FROM AUTHOR]

Abstract: We studied a child with apparent monosomy of chromosome 21. Cytogenetic, FISH and microsatellite analyses revealed a 45,X,−21,+der(X)t(X;21)(q25;q21.1) karyotype resulting from a de novo, unbalanced, X;21 non-reciprocal translocation of paternal origin, with partial monosomy of chromosomes 21 and X. An extreme, skewed X-inactivation pattern of the der(X) chromosome was demonstrated. Skewed inactivation probably accounted for a mild phenotype with respect to Xq25→qter deletion while propagation of inactivation to the adjacent 21q region may account for mild clinical features associated to distal 21q monosomy. [Copyright &y& Elsevier]

Abstract: Sex differences in behavior can be attributed to differences in steroid hormones. Sex chromosome complement can also influence behavior, independent of gonadal differentiation. The mice used for this work combined a spontaneous mutation of the Sry gene with a transgene for Sry that is incorporated into an autosome thus disassociating gonad differentiation from sex chromosome complement. The resulting genotypes are XX and XY− females (ovary-bearing) along with XXSry and XY− Sry males (testes-bearing). Here we report results of basic behavioral phenotyping conducted with these mice. Motor coordination, use of olfactory cues to find a food item, general activity, foot shock threshold, and behavior in an elevated plus maze were not affected by gonadal sex or sex chromosome complement. In a one-way active avoidance learning task females were faster to escape an electric shock than males. In addition, sex chromosome complement differences were noted during social interactions with submissive intruders. Female XY− mice were faster to follow an intruder than XX female mice. All XY− mice spent more time sniffing and grooming the intruder than the XX mice, with XY− females spending the most amount of time in this activity. Finally, XX females were faster to display an asocial behavior, digging, and engaged in more digging than XXSry male mice. All of these behaviors were tested in gonadectomized adults, thus, differences in circulating levels of gonadal steroids cannot account for these effects. Taken together, these data show that sex chromosome complement affects social interaction style in mice. [Copyright &y& Elsevier]

Abstract: Fabry disease is an under-recognized X-linked lysosomal disorder, due to α-galactosidase A deficiency. Most of the mutations in the GLA gene are detectable using genomic sequencing analysis. However, deletions of one or more exons or deletion encompassing the entire gene are undetectable, especially in heterozygous females. The Multiplex Ligation-dependent Probe Amplification (MLPA) is an efficient tool for discovering these rearrangements. In this study two novel different deletions were detected using MLPA assay on two Fabry patients, both resulted mutation negative by sequencing analysis. These data suggest that this screening should be systematically included in genetic testing surveys of patients with Fabry disease. [Copyright &y& Elsevier]

Using computer-based methods, we determined the global distribution of short interspersed nuclear elements ( SINEs) on human and mouse X chromosomes. It was shown that this distribution was similar to the distribution of CpG islands and genes, but was different from the distribution of LINE1 elements. Since SINEs (human Alu and mouse B2) may have binding sites for Polycomb protein YY1, we suggest that these repeats can serve as additional signals (“boosters”) in Polycomb-dependent silencing of gene-rich segments during X inactivation. [ABSTRACT FROM AUTHOR]

Neuroblastic tumours are composed of variable proportions of neuroblasts and Schwann cells. Whether both components share a common neoplastic origin is highly debated and discrepant results have been reported about the presence of tumour-related genetic alterations in Schwann cells. We have used X-methylation analysis and array-CGH to investigate contiguous Schwannian and neuroblastic areas in tumours with a nodular pattern. A skewed X inactivation was observed in four out of five stromal components. Interestingly, in these four cases, the X-inactivation profiles of the neuroblastic components were identical to the matched stromal areas. However, whereas all neuroblastic areas displayed chromosomal imbalances, no alteration was found in any Schwann cell components. Similarly, no alteration was observed in a series of 19 tumours with a single stroma-rich component, which occasionally exhibited a skewed X-inactivation pattern (3/17 informative tumours). Altogether, this indicates that most stroma-rich tumours display a polyclonal proliferation and that Schwann cells do not derive from neuroblasts. However, in tumours with both stroma-rich and -poor components, our results suggest that cells from both areas share a common progenitor.Oncogene (2008) 27, 3066–3071; doi:10.1038/sj.onc.1210965; published online 10 December 2007 [ABSTRACT FROM AUTHOR]

Focal dermal hypoplasia (FDH) is an X-linked dominant disorder characterized by patchy dermal hypoplasia with digital, ocular and dental abnormalities. Very recently, mutations in the PORCN gene were demonstrated to cause FDH. Here, we described three unrelated Thai girls who were sporadic cases of FDH. One of them had unilateral athelia, which has never been described in FDH. Mutation analysis by polymerase chain reaction sequencing the entire coding regions of PORCN successfully revealed three potentially pathogenic mutations, c.373+1G>A, c.737_738insA and c.1094G>A (p.R365Q). One was found in each of three patients. In addition, another sequence variant c.682C>T (p.R228C) with an inconclusive role was found in one patient and her unaffected mother. The two missense mutations were not detected in at least 100 ethnic-matched control chromosomes, and all four mutations had never been previously described. X chromosome inactivation studies showed random patterns in all of them. This study demonstrates that PORCN is the gene responsible for FDH across different populations and extends the total number of confirmed mutations to 26. [ABSTRACT FROM AUTHOR]

Only one X chromosome functions in diploid human cells irrespective of the sex of the individual and the number of X chromosomes. Yet, as we show, more than one X is active in the majority of human triploid cells. Therefore, we suggest that (i) the active X is chosen by repression of its XIST locus, (ii) the repressor is encoded by an autosome and is dosage sensitive, and (iii) the extra dose of this key repressor enables the expression of more than one X in triploid cells. Because autosomal trisomies might help locate the putative dosage sensitive trans-acting factor, we looked for two active X chromosomes in such cells. Previously, we reported that females trisomic for 18 different human autosomes had only one active X and a normal inactive X chromosome. Now we report the effect of triplication of the four autosomes not studied previously; data about these rare trisomies – full or partial – were used to identify autosomal regions relevant to the choice of active X. We find that triplication of the entire chromosomes 5 and 11 and parts of chromosomes 1 and 19 is associated with normal patterns of X inactivation, excluding these as candidate regions. However, females with inherited triplications of 1p21.3–q25.3, 1p31 and 19p13.2–q13.33 were not ascertained. Thus, if a single key dose-sensitive gene induces XIST repression, it could reside in one of these locations. Alternatively, more than one dosage-sensitive autosomal locus is required to form the repressor complex.European Journal of Human Genetics (2008) 16, 153–162; doi:10.1038/sj.ejhg.5201944; published online 31 October 2007 [ABSTRACT FROM AUTHOR]

Chromosome territory (CT) organization and chromatin condensation have been linked to gene expression, Although individual genes can be transcribed from inside CTs, some regions that have constitutively high expression or are coordinately activated loop out from CTs and decondense. The relationship between epigenetic marks, such as DNA methylation, and higher-order chromatin structures is largely unexplored, DNMT3B mutations in immuno-deficiency centromeric instability facial anomalies (ICF) syndrome result in loss of DNA methylation at particular sites, including CpG islands on the inactive X chromosome (Xi). This allows the specific effects of DNA methylation on CTs to be examined. Using fluorescence in situ hybridization, we reveal a differential organization of the human pseudoautosomal region (PAR)2 between the CTs of the X and Y in normal males and the active X (Xa) and the Xi in females. There is also a more condensed chromatin structure on Xi compared with Xa in this region. PAR2 genes are relocalized toward the outside of the Y and Xi CTs in ICF, and on the Xi, we show that this can extend to genes distant from the site of DNA hypomethylation itself. This reorganization is not simply a reflection of the transcriptional activation of the relocalized genes. This report of altered CT organization in a human genetic disease illustrates that DNA hypomethylation at restricted sites in the genome can lead to more extensive changes in nuclear organization away from the original site of eplgenetic change. [ABSTRACT FROM AUTHOR]

Abstract: Histone variants replace the core histones in a substantial fraction of nucleosomes, affecting chromatin structure and impacting chromatin-templated processes. In many instances incorporation of histone variants results in formation of specialized regions of chromatin. Proper localization of histone variants to distinct regions of the genome is critical for their function, yet how this specific localization is achieved remains unclear. macroH2A1 is enriched on the inactive X chromosome in female mammalian cells, where it functions to maintain gene silencing. macroH2A1 consists of a histone H2A-like histone domain and a large, globular C-terminal macro domain that is not present in other histone proteins. The histone domain of macroH2A1 is alone sufficient to direct enrichment on the inactive X chromosome when expressed in female cells, indicating that sequences important for correct localization lie in this domain. Here we investigate whether divergent sequences of the H2A variant macroH2A1 contribute to its correct localization. We mapped the regions of the macroH2A1 histone domain that are sufficient for localization to the inactive X chromosome using chimeras between H2A and the histone domain of macroH2A1. Multiple short sequences dispersed along the macroH2A1 histone domain individually supported enrichment on the inactive X chromosome when introduced into H2A. These sequences map to the surface of the macroH2A1/H2B dimer, but are buried in the crystal structure of the macroH2A1 containing nucleosome, suggesting that they may contribute to recognition by macroH2A1/H2B deposition factors. [Copyright &y& Elsevier]

Abstract: Objective: X inactivation pattern in X chromosome rearrangements usually favor the less unbalanced cells. It is correlated to a normal phenotype, small size or infertility. We studied the correlation between phenotype and X inactivation ratio in patients with X structural anomalies. Patients and methods: During the 1999–2005 period, 12 X chromosome rearrangements, including three prenatal cases, were diagnosed in the Laboratoire de Cytogénétique of Strasbourg. In seven cases, X inactivation ratio could be assessed by late replication or methylation assay. Results: In three of seven cases (del Xp, dup Xp, t(X;A)), X inactivation ratio and phenotype were consistent. The four other cases showed discrepancies between phenotype and X inactivation pattern: mental retardation and dysmorphism in a case of balanced X-autosome translocation, schizophrenia and autism in two cases of XX maleness and MLS syndrome (microphthalmia with linear skin defects) in a case of Xp(21.3-pter) deletion. Conclusion: Discrepancies between X inactivation ratio and phenotype are not rare and can be due to gene disruption, position effect, complex microrearrangements, variable pattern of X inactivation in different tissues or fortuitous association. In this context, the prognostic value of X inactivation study in prenatal diagnosis will be discussed. [Copyright &y& Elsevier]

Abstract: About 25% of X-linked genes may escape inactivation at least to some degree. However, in vitro results from somatic cell hybrids may not reflect what happens in vivo. Therefore, we analyzed the female/male (F/M) gene fold expression ratio for 299 X-linked and 7795 autosomal genes from 11 different tissues from an existing in vivo microarray database. On average 5.1 and 4.9% of genes showed higher expression in females compared with 7.4 and 7.9% in males, respectively, for X-linked and autosomal genes. A trend was found for F/M gene fold ratios greater than 1.5 for several X-linked genes indicating overexpression in females among multiple tissues. Nine X-linked genes showed overexpression in females in at least 3 of the 11 studied tissues. Of the 9 genes, 6 were located on the short arm and 3 on the long arm of the X chromosome. Six of the 9 genes have previously been reported to escape X inactivation. However, in general, no consistent pattern was seen for the expression of X-linked genes between in vitro and in vivo systems. This study indicates that factors other than the X-inactivation process may impact on the expression of X-linked genes resulting in an overall similar gender expression for both X-linked and autosomal genes. [Copyright &y& Elsevier]

Rett syndrome is a largely sporadic, X-linked neurological disorder with a characteristic phenotype, but which exhibits substantial phenotypic variability. This variability has been partly attributed to an effect of X chromosome inactivation (XCI). There have been conflicting reports regarding incidence of skewed X inactivation in Rett syndrome. In rare familial cases of Rett syndrome, favourably skewed X inactivation has been found in phenotypically normal carrier mothers. We have investigated the X inactivation pattern in DNA from blood and buccal cells of sporadic Rett patients (n=96) and their mothers (n=84). The mean degree of skewing in blood was higher in patients (70.7%) than controls (64.9%). Unexpectedly, the mothers of these patients also had a higher mean degree of skewing in blood (70.8%) than controls. In accordance with these findings, the frequency of skewed (XCI ≥80%) X inactivation in blood was also higher in both patients (25%) and mothers (30%) than in controls (11%). To test whether the Rett patients with skewed X inactivation were daughters of skewed mothers, 49 mother–daughter pairs were analysed. Of 14 patients with skewed X inactivation, only three had a mother with skewed X inactivation. Among patients, mildly affected cases were shown to be more skewed than more severely affected cases, and there was a trend towards preferential inactivation of the paternally inherited X chromosome in skewed cases. These findings, particularly the greater degree of X inactivation skewing in Rett syndrome patients, are of potential significance in the analysis of genotype–phenotype correlations in Rett syndrome.European Journal of Human Genetics (2006) 14, 1189–1194. doi:10.1038/sj.ejhg.5201682; published online 5 July 2006 [ABSTRACT FROM AUTHOR]

Abstract: X chromosome inactivation (XCI) is the phenomenon through which one of the two X chromosomes in female mammals is silenced to achieve dosage compensation with males. XCI is a highly complex, tightly controlled and developmentally regulated process. The mouse undergoes two forms of XCI: imprinted, which occurs in all cells of the preimplantation embryo and in the extraembryonic lineage, and random, which occurs in somatic cells after implantation. This review presents results and hypotheses that have recently been proposed concerning important aspects of both imprinted and random XCI in mice. We focus on how imprinted XCI occurs during preimplantation development, including a brief discussion of the debate as to when silencing initiates. We also discuss regulation of random XCI, focusing on the requirement for Tsix antisense transcription through the Xist locus, on the regulation of Xist chromatin structure by Tsix and on the effect of Tsix regulatory elements on choice and counting. Finally, we review exciting new data revealing that X chromosomes co-localize during random XCI. To conclude, we highlight other aspects of X-linked gene regulation that make it a suitable model for epigenetics at work. [Copyright &y& Elsevier]

In mammals X inactivation is initiated by expression of Xist RNA and involves the recruitment of Polycomb repressive complex 1 (PRC1) and 2 (PRC2), which mediate chromosome-wide ubiquitination of histone H2A and methylation of histone H3, respectively. Here, we show that PRC1 recruitment by Xist RNA is independent of gene silencing. We find that Eed is required for the recruitment of the canonical PRC1 proteins Mph1 and Mph2 by Xist. However, functional Ring1b is recruited by Xist and mediates ubiquitination of histone H2A in Eed deficient embryonic stem (ES) cells, which lack histone H3 lysine 27 tri-methylation. Xist expression early in ES cell differentiation establishes a chromosomal memory, which allows efficient H2A ubiquitination in differentiated cells and is independent of silencing and PRC2. Our data show that Xist recruits PRC1 components by both PRC2 dependent and independent modes and in the absence of PRC2 function is sufficient for the establishment of Polycomb-based memory systems in X inactivation. [ABSTRACT FROM AUTHOR]

X inactivation is associated with chromosome-wide establishment of inactive chromatin. Although this is classically regarded as facultative heterochromatin that is uniform in nature, the exact distribution of associated epigenetic marks is not well defined. Here we have analysed histone modifications in human somatic cells within two selected regions of the X chromosome. Intergenic, coding and promoter regions are segregated into differentially marked chromatin. H3K27me3 is most prominent in intergenic and silenced coding regions, but is associated with some active coding regions as well. Histone H3/H4 acetylation and H3K4me3 are locally enriched at promoter regions but do not necessarily mark continuing transcription. Remarkably, H3K9me3 is predominant in coding regions of active genes, a phenomenon that is not restricted to the X chromosome. These results argue against the exclusiveness of individual marks to heterochromatin or euchromatin, but rather suggest that composite patterns of interdependent or mutually exclusive modifications together signal the gene expression status. [ABSTRACT FROM AUTHOR]

Aim: Fabry disease is an X-linked lysosomal storage disorder characterized by an accumulation of neutral glycosphingolipids in multiple organ systems caused by α-galactosidase A deficiency due to mutations in the GLA gene. The majority of heterozygous females show the characteristic signs and symptoms of the disease, and some of them are severely affected. The current hypothesis for the occurrence of disease manifestations in females is skewed X inactivation favouring the mutant GLA allele. Method: We analyzed the patterns of X inactivation in the leukocytes of 28 biochemically and genetically characterized symptomatic Fabry disease heterozygotes and their correlation with clinical and biochemical disease expression. Results: X inactivation patterns in symptomatic females who are heterozygous for Fabry disease did not differ from those of female controls of the same age ( p = 0.669). Thirteen (46%) of the 28 females with Fabry disease showed random X inactivation, ten (36%) moderate skewing, and five (18%) highly skewed X inactivation. Segregation analysis was performed in the families of six females who had highly or moderately skewed X inactivation. In four of these females, skewing favoured the wild-type GLA allele and in the other two skewing favoured the mutant allele. Patterns of X inactivation or the extent of skewing were not related to the severity of clinical manifestations or to residual enzyme activity. Conclusion: In this study we provide evidence that heterozygous females with Fabry disease show random X inactivation. Our data do not support the hypothesis that the occurrence and severity of disease manifestations in the majority of Fabry heterozygotes are related to skewed X inactivation. [ABSTRACT FROM AUTHOR]

In female mammalian cells, one of the two X chromosomes is inactivated in early embryonic life. Females are mosaics for two cell populations, one with the maternal and one with the paternal X as the active chromosome. Skewed X inactivation is arbitrarily defined, often as a pattern where 80% or more of the cells show a preferential inactivation of one X chromosome. Inactivation is presumed to be permanent for all descendants of a cell; however, after about 55 years of age, the frequency of skewed X inactivation in peripheral blood cells increases, probably through selection. Unfavourable skewing of X inactivation, where the X chromosome carrying a mutant allele is the predominantly active X, has been found in affected female carriers of several X-linked disorders; however, for many X-linked disorders, a consistent relationship between the pattern of X inactivation and clinical phenotype has been difficult to demonstrate. One reason for this may be that peripheral blood cells are not a representative or relevant tissue in many disorders. In some severe X-linked disorders, post-inactivation selection takes place against the X chromosome carrying the mutant allele, leading to a completely skewed X-inactivation pattern. Skewed X inactivation has also been reported in young females with breast cancer, and may indicate an effect of X-linked genes on the development of this condition. Conclusion: The process of X inactivation and the resultant degree of skewing is clearly important for the expression of genetic diseases. It is also important to consider, however, that under normal conditions the frequency of skewed X inactivation increases with age in peripheral blood cells. Analysis of the expression of a large proportion of the genes on the X chromosome has revealed that X-chromosome inactivation is more heterogeneous than previously thought. [ABSTRACT FROM AUTHOR]

Abstract: Higher expression of X-linked genes in females might contribute to brain sexual differentiation. Although X-inactivation is thought to balance gene dosage between the two sexes, some X-linked genes escape X inactivation and therefore are expressed from both X chromosomes in females. Eif2s3x encodes subunit three of eukaryotic translation initiation factor 2, which regulates the rate of protein translation, and escapes X-inactivation in both humans and mice. By Northern blot analysis, we found Eif2s3x to be expressed higher in females than in males in developing and adult brains as well as adult liver. Gonadally intact XX mice had a higher level of Eif2s3x mRNA expression than XY mice regardless of whether they had testes or ovaries, suggesting that sexually dimorphic gene expression arises as a consequence of sex chromosome complement. In situ hybridization indicated that Eif2s3x mRNA was expressed preferentially in specific brain regions including the habenula, anterodorsal thalamic nucleus, hippocampus, hypothalamus, and cerebellum. Females had significantly higher levels of Eif2s3x mRNA expression than males in cortex, hippocampus and paraventricular nucleus but not in the habenula. The effect of a sex difference in Eif2s3x transcription, however, could potentially be offset by the additional expression in male brains of its Y-linked homologue Eif2s3y which was found in similar brain regions. The sex difference in Eif2s3x transcript appears not to be preserved at the protein level, since no difference in the levels of Eif2s3 protein was found between (1) males and females (2) XX and XY mice, or (3) XO and XX females. [Copyright &y& Elsevier]

Abstract: Immunodysregulation, Polyendocrinopathy, Enteropathy, X-linked (IPEX) syndrome is a rare inborn error of immune regulation characterized by the early onset of one or more autoimmune diseases in boys. IPEX is caused by mutations in FOXP3, and is thus the homologue of the scurfy mutant mouse. The gene product, Scurfin, is required for the development of CD4+CD25+ T regulatory cells. In the absence of T regulatory cells, activated CD4+ T cells instigate multi-organ damage resulting in type 1 diabetes, enteropathy, eczema, hypothyroidism, and other autoimmune disorders. While effective therapies are currently limited, studies in the scurfy mouse are revealing aspects of pathophysiology and genetics that will lead to novel approaches for treating IPEX and other autoimmune disorders. Females carrying Foxp3 mutations are unaffected. In new experiments we show that female scurfy mice that are also heterozygous in trans for the X-linked recessive common γ chain knockout contract autoimmune disease, proving that murine Foxp3 is subject to X-inactivation and providing an example of gene–gene interaction causing autoimmune disease in females. One explanation for the lesser disease severity in these females is proposed. [Copyright &y& Elsevier]

Autism is a heterogeneous neurodevelopmental disorder with a 3–4 times higher sex ratio in males than females. X chromosome genes may contribute to this higher sex ratio through unusual skewing of X chromosome inactivation. We studied X chromosome skewness in 30 females with classical autism and 35 similarly aged unaffected female siblings as controls using the polymorphic androgen receptor (AR) gene. Significantly, increased X chromosome skewness (e.g., >80:20%) was detected in our autism group (33%) compared to unaffected females (11%). X chromosome skewness was also seen in 50% of the mothers with autistic daughters. No mutation was seen in the promoter region of the XIST gene reported to be involved in X chromosome inactivation in our subjects. X chromosome skewness has been reported in female carriers of other neurological disorders such as X-linked mental retardation, adrenoleukodystrophy and Rett syndrome. [ABSTRACT FROM AUTHOR]

The active and inactive X chromosomes have distinct epigenetic marks in somatic nuclei, which undergo reprogramming after transplantation into oocytes. We show that, despite the disappearance of Xist RNA coating in 30 min, the epigenetic memory of the inactive X persists with the precocious appearance of histone H3 trimethylation of lysine 27 (H3-3meK27), without the expected colocalization with Eed/Ezh2. Subsequently, Xist re-appears on the original inactive X, and the silent Xist on the active X undergoes re-activation, resulting in unusual biallelic Xist RNA domains. Despite this abnormal Xist expression pattern, colocalization of H3-3meK27 and Eed is thereafter confined to a single Xist domain, which is presumably on the original inactive X. These epigenetic events differ markedly from the kinetics of preferential paternal X inactivation in normal embryos. All the epigenetic marks on the X are apparently erased in the epiblast, suggesting that the oocyte and epiblast may have distinct properties for stepwise programming of the genome. [ABSTRACT FROM AUTHOR]

Abstract: Skewed X chromosome inactivation (XCI) has been implicated in modulating the severity of Rett syndrome (RTT), although studies by different groups have yielded conflicting results. In this study we have characterised the XCI pattern in various neuroanatomical regions of nine RTT brains and non-neural tissue in two of these patients to determine whether or not variable XCI patterns occur in different brain regions or somatic tissues of the same patient. The mean XCI patterns for frontal and occipital cortex were compared between RTT and control subjects, and showed no significant differences when comparing RTT frontal to control frontal cortex or RTT occipital to control occipital cortex. However, one RTT subject displayed variability across the different neuroanatomical regions of the brain and skewing in some non-neural tissues. This observation adds another dimension to the epigenetic factors that may contribute to the phenotype in RTT. It also mandates that caution should be exercised in factoring XCI, including assumptions based on the blood XCI pattern, into the development of phenotype–genotype correlations. [Copyright &y& Elsevier]

Abstract: Lesch–Nyhan disease (LND) is an inborn error of purine metabolism caused by defective activity of the enzyme hypoxanthine guanine phosphoribosyl transferase (HPRT, EC 2.4.2.8), resulting from mutation in the corresponding gene on the long arm of the X chromosome (Xq26). The classic phenotype occurs almost exclusively in males and is characterized by hyperuricemia, mental retardation, severe dystonia, and self-injurious behavior. Heterozygous carrier females are usually clinically normal. However, a small number of clinically affected females have been described. In all previous cases there was a mutation in one HPRT allele and non-random inactivation of the X chromosome carrying the normal HPRT gene. We have analyzed a female MZ twin pair discordant for Lesch–Nyhan disease. The mother and both twins are heterozygous carriers of a HPRT splicing mutation (IVS8+4A>G; c.609+4A>G) and all three express the mutant allele at similar frequencies in peripheral blood T cells. The mother and one sister are clinically normal. In the affected twin, the clinical phenotype is classical for Lesch–Nyhan disease, despite the fact that HPRT activity in the blood was also normal. X inactivation analysis showed a skewed pattern in the fibroblasts of the affected twin sister, with the X chromosome carrying the normal HPRT allele preferentially inactivated. As in many other reported cases of X-linked diseases, the discordant phenotype of the two monozygous twin sisters suggests that the process responsible for monozygotic twinning can trigger skewed X inactivation. [Copyright &y& Elsevier]

To investigate the genetic influence on X chromosome inactivation and on age-related skewing of X inactivation, in particular, we analysed the X inactivation pattern (XIP) in peripheral blood cells from 118 young monozygotic (MZ) twin pairs (18-53 years), 82 elderly MZ twin pairs (55-94 years), 146 young dizygotic (DZ) twin pairs (20-54 years) and 112 elderly DZ twin pairs (64-95 years). Elderly twins had a higher frequency of skewed X inactivation (34%) than young twins (15%) (P<0.001). Our data suggest that the increase in skewing occurs after age 50-60 years. The intraclass correlation was 0.61 and 0.58 in young and elderly MZ twin pairs, and 0.08 and 0.09 in young and elderly DZ twin pairs. Biometric analysis showed that dominant genetic effects accounted for 63 and 58%of the variance of XIP in the young and elderly twin pairs, respectively. The dominant genetic effect and the shared environment for monochorionic MZ twins may explain the high intraclass correlation for the MZ twin pairs compared to the DZ twin pairs. We did not observe a significant decrease in the intraclass correlation in elderly MZ twins compared to young MZ twins, which would be expected if age-related skewing were due to stochastic factors. We conclude that the increased skewing with age implies that a genetically dependent selection of blood cells take place.European Journal of Human Genetics (2005) 13, 599-606. doi:10.1038/sj.ejhg.5201398 Published online 9 March 2005 [ABSTRACT FROM AUTHOR]

Abstract: Somatic cell nuclear transfer embryos exhibit extensive epigenetic abnormalities, including aberrant methylation and abnormal imprinted gene expression. In this study, a thorough analysis of X chromosome inactivation (XCI) was performed in both preimplantation and postimplantation nuclear transfer embryos. Cloned blastocysts reactivated the inactive somatic X chromosome, possibly in a gradient fashion. Analysis of XCI by Xist RNA and Eed protein localization revealed heterogeneity within cloned embryos, with some cells successfully inactivating an X chromosome and others failing to do so. Additionally, a significant proportion of cells contained more than two X chromosomes, which correlated with an increased incidence of tetraploidy. Imprinted XCI, normally found in preimplantation embryos and extraembryonic tissues, was not observed in blastocysts or placentae from later stage clones, although fetuses recapitulated the Xce effect. We conclude that, although SCNT embryos can reactivate, count, and inactivate X chromosomes, they are not able to regulate XCI consistently. These results illustrate the heterogeneity of epigenetic changes found in cloned embryos. [Copyright &y& Elsevier]

Based on the formation of the XY body at pachytene and expression studies of a few X-linked genes, the X and Y chromosomes seem to undergo transcriptional inactivation during mammalian spermatogenesis. However, the extent and the mechanism of X and Y inactivation are not known. Here, we show that both the X and Y chromosomes undergo sequential change in their histone modifications beginning at the pachytene stage of meiosis. These changes are usually are associated with transcriptional inactivation in somatic cells, and they coincide with the exclusion of the phos-phorylated (active) form of RNA polymerase II from the XY body. Both sex chromosomes undergo extensive deacetylation at histone H3 and H4 and (di)methylation of lysine (K)9 on histone H3; however, there are no changes in H3-K4 methylation. These changes priest even when the XY body disappears in late pachytene, and the X and Y chromosomes segregate from one another after the first meiotic division. By the spermatid stage, histone modifications of the X and Y chromosomes revert to those of active chromatin and RNA polymerase II reengages with both chromosomes. Our observations indicate that X and Y inactivation is extensive and persists even when the X and Y chromosomes are separated in secondary spermatocytes. These findings provide insights into epigenetic programming and chromatin dynamics in the male germ line. [ABSTRACT FROM AUTHOR]

Somatic cloning by nuclear transfer returns a differentiated cell to a totipotent stage, a process termed nuclear reprogramming. During this de-differentiation process, genes inactivated during tissue differentiation are re- activated in a temporal and spatial special manner. It is believed that tissue differentiation occurs through epigenetic mechanisms, genetic inheritance that does not involve changes in DNA sequences. Developmental abnormalities and a high mortality rate in cloned offspring have frequently been observed and probably result from incomplete nuclear reprogramming. In this review, the reprogramming of two epigenetic mechanisms, imprinting and X chromosome inactivation, as well as recent attempts to modify pre-existing epigenetic marks in donor cells to improve nuclear transfer efficacy, are discussed. [ABSTRACT FROM AUTHOR]

Nuclear transfer (NT) studies have provided insight into the functional importance of epigenetic alteration of the X chromosomes during X-inactivation. Uniparental embryos created by NT have been informative as to the time and location at which the imprint controlling extraembryonic X-inactivation is established. Experiments with female somatic cells, have demonstrated that the inactive X chromosome (Xi) is reactivated after NT, leading to random X-inactivation in the embryonic lineages of cloned embryos. However, in the extraembryonic lineages of clones, epigenetic information from the donor cell nucleus persists, leading to preferential inactivation of the donor cell’s inactive X in the placenta of cloned animals. These results suggest epigenetic information established during embryonic X-inactivation is functionally equivalent to the gametic imprint. [Copyright &y& Elsevier]

The first specimen of platypus (Ornithorhynchus anatinus) that reached Britain in the late 18th century was regarded a scientific hoax. Over decades the anatomical characteristics of these unique mammals, such as egg laying and the existence of mammary glands, were hotly debated before they were accepted. Within the last 40 years, more and more details of monotreme physiology, histology, reproduction and genetics have been revealed. Some show similarities with birds or reptiles, some with therian mammals, but many are very specific to monotremes. The genome is no exception to monotreme uniqueness. An early opinion was that the karyotype, composed of a few large chromosomes and many small ones, resembled bird and reptile macro- and micro-chromosomes. However, the platypus genome also features characteristics that are not present in other mammals, such as a complex translocation system. The sex chromosome system is still not resolved. Nothing is known about dosage compensation and, unlike in therian mammals, there seems to be no genomic imprinting. In this article we will recount the mysteries of the monotreme genome and describe how we are using recently developed technology to identify chromosomes in mitosis, meiosis and sperm, to map genes to chromosomes, to unravel the sex chromosome system and the translocation chain and investigate X inactivation and genomic imprinting in monotremes. [Copyright &y& Elsevier]

Oral–facial–digital type 1 (OFD1) syndrome is an X-linked dominant condition characterized by malformations of the face, oral cavity, and digits. The responsible gene, OFD1, maps to human Xp22 and has an unknown function. We isolated and characterized the mouse Ofd1 gene and showed that it is subject to X-inactivation, in contrast to the human gene. Furthermore, we excluded a role for Ofd1 in the pathogenesis of the spontaneous mouse mutant Xpl, which had been proposed as a mouse model for this condition. Comparative sequence analysis demonstrated that OFD1 is conserved among vertebrates and absent in invertebrates. This analysis allowed the identification of evolutionarily conserved domains in the protein. Finally, we report the identification of 18 apparently nonfunctional OFD1 copies, organized in repeat units on the human Y chromosome. These degenerate OFD1-Y genes probably derived from the ancestral Y homologue of the X-linked gene. The high level of sequence identity among the different units suggests that duplication events have recently occurred during evolution. [Copyright &y& Elsevier]

Abstract The molecular mechanism underlying X chromosome inactivation in female mammals involves the non-coding RNAs Xist and its antisense partner Tsix . Prior to X inactivation, these RNAs are transcribed in an unstable form from all X chromosomes, both in the early embryo and in undifferentiated embryonic stem (ES) cells. Upon differentiation, the expression of these unstable transcripts from all alleles is silenced, and Xist RNA becomes stabilised specifically on the inactivating X chromosome. This pattern of expression is then maintained throughout subsequent somatic cell divisions. Once established, the inactive state of the X chromosome is remarkably stable, the only natural case of reactivation occurring in XX primordial germ cells (PGCs) when they enter the genital ridge. To gain insight into the X reactivation process, we have analysed Xist gene expression using RNA FISH in PGCs and also in PGC-derived embryonic germ (EG) cells. XX EG cells were shown to express unstable Xist/Tsix from both X chromosomes. In contrast, no unstable Xist/Tsix transcripts were detected in XX PGCs at any stage. Instead, a proportion of XX PGCs isolated from the genital ridge between 11.5 and 13.5 dpc (the period during which X chromosome reactivation occurs) showed an accumulation of stable Xist RNA on one X. The number of these cells decreased progressively and was nearly extinguished by 13.5 dpc. As a late marker for the inactive state, we analysed localisation of the histone H2A variant macroH2A1.2. Although macroH2A1.2 expression was observed in PGCs, no significant localisation to the inactive X was detected at any stage. We discuss these results in the context of understanding X chromosome reactivation. [ABSTRACT FROM AUTHOR]

Hardikar syndrome (HS) is a MED12-related ultra-rare multiple congenital malformation syndrome known to affect the gastrointestinal, cardiac, and genitourinary systems among other features including cleft lip/palate and pigmentary retinopathy. Only 10 patients affected with HS have been previously described in literature, of which seven were molecularly confirmed. We report a 20-year-old and a 13-month-old patient with HS diagnosed by exome sequencing bringing the total number of clinically diagnosed cases to 12 and MED12 associated to 9. We describe previously unreported molecular and clinical findings associated with HS and review all reported cases to permit prompt diagnosis, appropriate management, and genetic counseling of HS patients., (© 2022 The Authors. American Journal of Medical Genetics Part A published by Wiley Periodicals LLC.)

Long non-coding RNAs (lncRNAs) play an important role in genome regulation. Specifically, many lncRNAs interact with chromatin, recruit epigenetic complexes and in this way affect large-scale gene expression programs. However, the experimental data about lncRNA-chromatin interactions is still limited. The majority of experimental protocols do not provide any insight into the mechanics of lncRNA-based genome-wide epigenetic regulation. Here we present the HiMoRNA (Histone-Modifying RNA) database, a resource containing correlated lncRNA–epigenetic changes in specific genomic locations genome-wide. HiMoRNA integrates a large amount of multi-omics data to characterize the effects of lncRNA on epigenetic modifications and gene expression. The current release of HiMoRNA includes more than five million associations in humans for ten histone modifications in multiple genomic loci and 4145 lncRNAs. HiMoRNA provides a user-friendly interface to facilitate browsing, searching and retrieving of lncRNAs associated with epigenetic profiles of various chromatin loci. Analysis of the HiMoRNA data suggests that several lncRNA including JPX might be involved not only in regulation of XIST locus but also in direct establishment or maintenance of X-chromosome inactivation. We believe that HiMoRNA is a convenient and valuable resource that can provide valuable biological insights and greatly facilitate functional annotation of lncRNAs. [ABSTRACT FROM AUTHOR]

X chromosome inactivation is a mammalian dosage compensation mechanism, where one of two X chromosomes is randomly inactivated in female cells. Previous studies have suggested that primed human embryonic stem cells (hESCs) maintain an eroded state of the X chromosome and do not express XIST , while in naïve transition, both XIST and the eroded X chromosome are reactivated. However, the pattern of chromosome X reactivation in naïve hESCs remains mainly unknown. In this study, we examine the variations in the status of X chromosome between primed and naïve hESCs by analyzing RNA sequencing samples from different studies. We show that most samples of naïve hESCs indeed reactivate XIST and there is an increase in gene expression levels on chromosome X. However, most of the naïve samples do not fully activate chromosome X in a uniform manner and present a distinct eroded pattern, probably as a result of XIST reactivation and initiation of re-inactivation of chromosome X. This large-scale analysis provides a higher-resolution description of the changes occurring in chromosome X during primed-to-naïve transition and emphasizes the importance of taking these variations into consideration when studying X inactivation in embryonic development.

A multivariate Gaussian model for mammalian development is presented with the associated biological and mathematical assumptions. Many biological investigations use the female mammal X chromosome to test hypotheses and to estimate parameters of the developmental system. In particular, Lyon's (1961) hypotheses are used as a basis of the mathematical model. Experimental mouse data and three sets of human experimental data are analyzed using the hypothesized Gaussian model. The estimated biological parameters are consistent with some current biological theories. [ABSTRACT FROM AUTHOR]