Increased skewing of X chromosome inactivation with age in both blood and buccal cells

The X chromosome inactivation pattern in peripheral blood cells becomes more skewed after age 55, and a genetic effect on this age-related skewing has been reported. We investigated the effect of age on X inactivation phenotype in blood, buccal cells and tissue from duodenal biopsies in 80 females aged 19-90 years. The X inactivation pattern correlated positively with age in blood (r = 0.238, P = 0.034) and buccal cells (r = 0.260, P = 0.02). The mean degree of skewing was higher in the elderly (>/=55 years) than in the young (<55 years) in blood (70.1 and 63.5%, respectively, P = 0.013) and in buccal cells (64.7 and 59.0%, respectively, P = 0.004). Correlation of X inactivation between the different tissues was high in all tissues with a tendency to increase with age for blood and buccal cells (P = 0.082). None of the duodenal biopsies had a skewed X inactivation, and the mean degree of skewing was similar in the two age groups. The tendency for the same X chromosome to be the preferentially active X in both blood and buccal cells with advancing age is in agreement with a genetic effect on age-related skewing and indicates that genes other than those involved in hematopoiesis should be investigated in the search for genes contributing to age related skewing.     

X chromosome inactivation mosaicism in the mouse

A cytologically detectable mosaicism resulting from X-chromosome inactivation occurring in mice heterozygous for Cattanach's translocation has been used to examine the time of X chromosome inactivation, and the sizes of primordial precursor pools for lung, thymus, spleen, fascia, and melanocytes. The extent of covariance in mosaic composition among tissues within individuals suggests that, if X inactivation occurs randomly, it must occur after determination of embryoblast cells, at some time immediately before or after implantation, and that it must occur before divergence of mesoderm from ectoderm. The extent of independent variance among the various tissues is such as to suggest that none of them arise from primordial precursor pools smaller than 20 to 30 cells.     

Non-random X chromosome inactivation in Aicardi syndrome

Most females have random X-chromosome inactivation (XCI), defined as an equal likelihood for inactivation of the maternally- or paternally-derived X chromosome in each cell. Several X-linked disorders have been associated with a higher prevalence of non-random XCI patterns, but previous studies on XCI patterns in Aicardi syndrome were limited by small numbers and older methodologies, and have yielded conflicting results. We studied XCI patterns in DNA extracted from peripheral blood leukocytes of 35 girls with typical Aicardi syndrome (AIC) from 0.25 to 16.42 years of age, using the human androgen receptor assay. Data on 33 informative samples showed non-random XCI in 11 (33%), defined as a >80:20% skewed ratio of one versus the other X chromosome being active. In six (18%) of these, there was a >95:5% extremely skewed ratio of one versus the other X chromosome being active. XCI patterns on maternal samples were not excessively skewed. The prevalence of non-random XCI in Aicardi syndrome is significantly different from that in the general population (p < 0.0001) and provides additional support for the hypothesis that Aicardi syndrome is an X-linked disorder. We also investigated the correlation between X-inactivation patterns and clinical severity and found that non-random XCI is associated with a high neurological composite severity score. Conversely, a statistically significant association was found between random XCI and the skeletal composite score. Correlations between X-inactivation patterns and individual features were made and we found a significant association between vertebral anomalies and random XCI.     

Stochastic models for X chromosome inactivation

Summary 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.     

X chromosome inactivation during Drosophila spermatogenesis

Genes with male- and testis-enriched expression are under-represented on the Drosophila melanogaster X chromosome. There is also an excess of retrotransposed genes, many of which are expressed in testis, that have “escaped” the X chromosome and moved to the autosomes. It has been proposed that inactivation of the X chromosome during spermatogenesis contributes to these patterns: genes with a beneficial function late in spermatogenesis should be selectively favored to be autosomal in order to avoid inactivation. However, conclusive evidence for X inactivation in the male germline has been lacking. To test for such inactivation, we used a transgenic construct in which expression of a lacZ reporter gene was driven by the promoter sequence of the autosomal, testis-specific ocnus gene. Autosomal insertions of this transgene showed the expected pattern of male- and testis-specific expression. X-linked insertions, in contrast, showed only very low levels of reporter gene expression. Thus, we find that X linkage inhibits the activity of a testis-specific promoter. We obtained the same result using a vector in which the transgene was flanked by chromosomal insulator sequences. These results are consistent with global inactivation of the X chromosome in the male germline and support a selective explanation for X chromosome avoidance of genes with beneficial effects late in spermatogenesis.     

X chromosome inactivation in X autosome translocation carrier cows.

The pattern of X chromosome inactivation in X autosome translocation carries in a herd of Limousin-Jersey crossbred cattle was studied using the reverse banding technique consisting of 5-bromodeoxyuridine incorporation and acridine orange staining and autoradiography on cultures of solid tissues and blood samples exposed to tritiated thymidine. The late-replicating X chromosome was noted to be the normal X in strikingly high proportions of cells in cultures of different tissues from all translocation carriers. It is suggested that the predominance of cells in which the normal X is inactivated may be the result of a post-inactivation selection process. Such a selection process during the prenatal life favouring cells in which the genes of the normal X chromosome remain unexpressed in translocation carrier females may be the mechanism that helps these conceptuses escape the adverse effects of functional aneuploidy. Based on the observation that the translocation carriers of this line of cattle are exclusively females and that there is a higher than expected rate of pregnancy loss, it is also postulated that the altered X chromosome may be lethal to all male conceptuses and to some of their female counterparts.     

X chromosome inactivation in the cycle of life

Female mammalian cells silence one of their two X chromosomes, resulting in equal expression levels of X-encoded genes in female XX and male XY cells. In mice, the X chromosomes in female cells go through sequential steps of inactivation and reactivation. Depending on the developmental time window, imprinted or random X chromosome inactivation (XCI) is initiated, and both processes lead to an inactive X chromosome that is clonally inherited. Here, we review new insights into the life cycle of XCI and provide an overview of the mechanisms regulating X inactivation and reactivation.     

Origin and evolution of X chromosome inactivation

Evolution of the mammalian sex chromosomes heavily impacts on the expression of X-encoded genes, both in marsupials and placental mammals. The loss of genes from the Y chromosome forced a two-fold upregulation of dose sensitive X-linked homologues. As a corollary, female cells would experience a lethal dose of X-linked genes, if this upregulation was not counteracted by evolution of X chromosome inactivation (XCI) that allows for only one active X chromosome per diploid genome. Marsupials rely on imprinted XCI, which inactivates always the paternally inherited X chromosome. In placental mammals, random XCI (rXCI) is the predominant form, inactivating either the maternal or paternal X. In this review, we discuss recent new insights in the regulation of XCI. Based on these findings, we propose an X inactivation center (Xic), composed of a cis-Xic and trans-Xic that encompass all elements and factors acting to control rXCI either in cis or in trans. We also highlight that XCI may have evolved from a very small nucleation site on the X chromosome in the vicinity of the Sox3 gene. Finally, we discuss the possible evolutionary road maps that resulted in imprinted XCI and rXCI as observed in present day mammals.     

X chromosome inactivation: heterogeneity of heterochromatin

The silent X chromosome in mammalian females is a classic example of facultative heterochromatin, the term highlighting the compacted and inactive nature of the chromosome. However, it is now clear that the heterochromatin of the inactive X is not homogeneous--as indeed, not all genes on the inactive X are silenced. We summarize known features and events of X inactivation in different mouse and human model systems, and highlight the heterogeneity of chromatin along the inactive X. Characterizing this heterogeneity is likely to provide insight into the cis-acting sequences involved in X chromosome inactivation.     

X chromosome inactivation in nuclear transfer ES cells

Nuclear transfer ES (ntES) cells are established from cloned blastocysts generated by somatic cell nuclear transfer and are expected to be an important resource for regenerative medicine. However, cloned mammals, generated by similar methods, show various abnormalities, which suggest disordered gene regulation. Random X chromosome inactivation (XCI) has been observed to take place in cloned female mouse embryos, but XCI does not necessarily occur according to Xce strength, a genetic element that determines the likelihood of each X chromosome to be inactivated. This observation suggests incomplete reprogramming of epigenetic marks related to XCI. Here, we investigated XCI in ntES cell lines, which were established using differentiated embryoid bodies that originated from a female mouse ES cell line. We examined Xist RNA localization, histone modifications in the Xist locus, and XCI choice. We did not find substantial differences between the ntES lines and their parental ES line. This suggests that the Xist locus and the epigenetic marks involved in XCI are reprogrammed by nuclear transfer and subsequent ntES cell establishment. In contrast to skewed XCI in cloned mice, our observations indicate that normal XCI choice takes place in ntES cells, which supports the goal of safe therapeutic cloning for clinical use.     

Understanding the X chromosome inactivation cycle in mice

During mouse development, imprinted X chromosome inactivation (XCI) is observed in preimplantation embryos and is inherited to the placental lineage, whereas random XCI is initiated in the embryonic proper. Xist RNA, which triggers XCI, is expressed ectopically in cloned embryos produced by somatic cell nuclear transfer (SCNT). To understand these mechanisms, we undertook a large-scale nuclear transfer study using different donor cells throughout the life cycle. The Xist expression patterns in the reconstructed embryos suggested that the nature of imprinted XCI is the maternal Xist-repressing imprint established at the last stage of oogenesis. Contrary to the prevailing model, this maternal imprint is erased in both the embryonic and extraembryonic lineages. The lack of the Xist-repressing imprint in the postimplantation somatic cells clearly explains how the SCNT embryos undergo ectopic Xist expression. Our data provide a comprehensive view of the XCI cycle in mice, which is essential information for future investigations of XCI mechanisms.     

X chromosome inactivation: theme and variations

My contribution to this special issue on Vertebrate Sex Chromosomes deals with the theme of X chromosome inactivation and its variations. I will argue that the single active X--characteristic of mammalian X dosage compensation--is unique to mammals, and that the major underlying mechanism(s) must be the same for most of them. The variable features reflect modifications that do not interfere with the basic theme. These variations were acquired during mammalian evolution--to solve special needs for imprinting and locking in the inactive state. Some of the adaptations reinforce the basic theme, and were needed because of species differences in the timing of interacting developmental events. Elucidating the molecular basis for the single active X requires that we distinguish the mechanisms essential for the basic theme from those responsible for its variations.     

X chromosome inactivation in carriers of Barth syndrome.

Barth syndrome (BTHS) is a rare X-linked recessive disorder characterized by cardiac and skeletal myopathy, neutropenia, and short stature. A gene for BTHS, G4.5, was recently cloned and encodes several novel proteins, named "tafazzins." Unique mutations have been found. No correlation between the location or type of mutation and the phenotype of BTHS has been found. Female carriers of BTHS seem to be healthy. This could be due to a selection against cells that have the mutant allele on the active X chromosome. We therefore analyzed X chromosome inactivation in 16 obligate carriers of BTHS, from six families, using PCR in the androgen-receptor locus. An extremely skewed X-inactivation pattern (>=95:5), not found in 148 female controls, was found in six carriers. The skewed pattern in two carriers from one family was confirmed in DNA from cultured fibroblasts. Five carriers from two families had a skewed pattern (80:20-<95:5), a pattern that was found in only 11 of 148 female controls. Of the 11 carriers with a skewed pattern, the parental origin of the inactive X chromosome was maternal in all seven cases for which this could be determined. In two families, carriers with an extremely skewed pattern and carriers with a random pattern were found. The skewed X inactivation in 11 of 16 carriers is probably the result of a selection against cells with the mutated gene on the active X chromosome. Since BTHS also shows great clinical variation within families, additional factors are likely to influence the expression of the phenotype. Such factors may also influence the selection mechanism in carriers.     

X chromosome imprinting and inactivation in preimplantation mammalian embryos

Dosage compensation for the mammalian X chromosome involves the silencing of one X chromosome to achieve equal X-linked gene expression between males and females. X chromosome inactivation (XCI) is controlled by a complex set of genetic elements located in a region known as the X chromosome inactivation center, and is regulated by a combination of genomic imprinting, cell lineage-dependent erasure of imprinting, an unidentified mechanism of X chromosome counting, an incompletely understood means of selection of one X chromosome for inactivation and developmentally regulated changes in X chromosome chromatin. A detailed understanding of when and how these components of XCI occur is essential for elucidating the operative mechanisms. A model accounting for early events related to XCI, including observations in uniparental and aneuploid embryos, is presented.