The XIST noncoding RNA functions independently of BRCA1 in X inactivation

Females with germline mutations in BRCA1 are predisposed to develop breast and ovarian cancers. A previous report indicated that BRCA1 colocalizes with and is necessary for the correct localization of XIST, a noncoding RNA that coats the inactive X chromosome (Xi) to mediate formation of facultative heterochromatin. A model emerged from this study suggesting that loss of BRCA1 in female cells could reactivate genes on the Xi through loss of the XIST RNA. However, our independent studies of BRCA1 and XIST RNA revealed little evidence to support this model. We report that BRCA1 is not enriched on XIST RNA-coated chromatin of the Xi. Neither mutation nor depletion of BRCA1 causes significant changes in XIST RNA localization or X-linked gene expression. Together, these results do not support a role for BRCA1 in promoting XIST RNA localization to the Xi or regulating XIST-dependent functions in maintaining the stability of facultative heterochromatin.     

Cytokinesis: Rho marks the spot

During cytokinesis of a eukaryotic cell, following the chromosome movements of anaphase, a contractile ring forms in the cortex midway between the segregating chromosomes and divides the cell into two daughters. Recent studies have provided new insights into the mechanism by which the site of contractile ring assembly is specified.     

mRNA decay: x (XRN1) marks the spot

Degradation of mRNA is a vital aspect of gene expression. In yeast, Dcp1p, Dcp2p, Lsm1-7p, and Xrn1p are required for mRNA decay and are localized within discrete cytoplasmic foci; in the May 2 issue of Science, Sheth and Parker provide compelling evidence that these foci represent sites for mRNA decay.     

The search for the mouse X-chromosome inactivation centre

The phenomenon of X-chromosome inactivation in female mammals, whereby one of the two X chromosome present in each cell of the female embryo is inactivated early in development, was first described by Mary Lyon in 1961. Nearly 30 years later, the mechanism of X-chromosome inactivation remains unknown. Strong evidence has accumulated over the years, however, for the involvement of a major switch or inactivation centre on the mouse X chromosome. Identification of the inactivation centre at the molecular level would be an important step in understanding the mechanism of X-inactivation. In this paper we review the evidence for the existence and location of the X-inactivation centre on the mouse X-chromosome, present data on the molecular genetic mapping of this region, and describe ongoing strategies we are using to attempt to identify the inactivation centre at the molecular level.     

Polymorphic X-chromosome inactivation of the human TIMP1 gene.

X inactivation silences most but not all of the genes on one of the two X chromosomes in mammalian females. The human X chromosome preserves its activation status when isolated in rodent/human somatic-cell hybrids, and hybrids retaining either the active or inactive X chromosome have been used to assess the inactivation status of many X-linked genes. Surprisingly, the X-linked gene for human tissue inhibitor of metalloproteinases (TIMP1) is expressed in some but not all inactive X-containing somatic-cell hybrids, suggesting that this gene is either prone to reactivation or variable in its inactivation. Since many genes that escape X inactivation are clustered, we examined the expression of four genes (ARAF1, ELK1, ZNF41, and ZNF157) within approximately 100 kb of TIMP1. All four genes were expressed only from the active X chromosome, demonstrating that the factors allowing TIMP1 expression from the inactive X chromosome are specific to the TIMP1 gene. To determine if this variable inactivation of TIMP1 is a function of the hybrid-cell environment or also is observed in human cells, we developed an allele-specific assay to assess TIMP1 expression in human females. Expression of two alleles was detected in some female cells with previously demonstrated extreme skewing of X inactivation, indicating TIMP1 expression from the inactive chromosome. However, in other cells, no expression of TIMP1 was observed from the inactive X chromosome, suggesting that TIMP1 inactivation is polymorphic in human females.     

X-chromosome inactivation and the search for chromosome-wide silencers

X-chromosome inactivation leads to divergent fates for two homologous chromosomes. Whether one X remains active or becomes silenced depends on the activity of Xist, a gene expressed only from the inactive X and whose RNA product 'paints' the X in cis. Recent work argues that Xist RNA itself is the acting agent for initiating the silencing step. Xist RNA contains separable domains for RNA localization and chromosome silencing. While no Xist RNA-interacting factors have been identified, a growing collection of chromatin alterations have been identified on the inactive X, including variant histone H2A composition and histone H3 methylation. Some or all of these changes may be critical for chromosome-wide silencing. As none of the silencing proteins identified so far is unique to X chromosome inactivation, the specificity must partly reside in Xist RNA whose spread along the X orchestrates general silencing factors for this specific task.     

X-chromosome inactivation in the human cytotrophoblast

Preferential paternal X-chromosome inactivation occurs in the cell lineages that differentiate first within the female rodent blastula (trophectoderm and extraembryonic endoderm). The present studies were designed to test the nature of X-chromosome inactivation (XCI) in the earliest differentiating cell lineage of the human placenta, the cytotrophoblast. Using glucose-6-phosphate-dehydrogenase (G6PD) polymorphisms as a marker system, term placentae were obtained from 13 female heterozygotes where parental allelic contributions could be determined. Chorionic villi were enzymatically digested and centrifuged in a Percoll density gradient to isolate a pure population of cytotrophoblasts, which was ascertained by cell culture, differentiation to syncytiotrophoblasts, and histochemical staining for alpha-human chorionic gonadotrophin (alpha-HCG). On electrophoresis, all 13 samples exhibited exclusive or near exclusive expression of the maternally derived X-linked enzyme variant, regardless of whether it was G6PD A or G6PD B. No intermediate bands were seen, indicating a single active G6PD locus per cell. The stromal cells of the villi, which derive from the mesoderm and differentiate later than the cytotrophoblast, exhibit random XCI. These findings establish preferential paternal XCI in the cytotrophoblast, the cell type first to differentiate within the human blastula.     

X-chromosome inactivation: molecular mechanisms from the human perspective

X-chromosome inactivation is an epigenetic process whereby one X chromosome is silenced in mammalian female cells. Since it was first proposed by Lyon in 1961, mouse models have been valuable tools to uncover the molecular mechanisms underlying X inactivation. However, there are also inherent differences between mouse and human X inactivation, ranging from sequence content of the X inactivation center to the phenotypic outcomes of X-chromosome abnormalities. X-linked gene dosage in males, females, and individuals with X aneuploidies and X/autosome translocations has demonstrated that many human genes escape X inactivation, implicating cis-regulatory elements in the spread of silencing. We discuss the potential nature of these elements and also review the elements in the X inactivation center involved in the early events in X-chromosome inactivation.     

X-marks the spot: X-chromosome identification during dosage compensation

Dosage compensation is the essential process that equalizes the dosage of X-linked genes between the sexes in heterogametic species. Because all of the genes along the length of a single chromosome are co-regulated, dosage compensation serves as a model system for understanding how domains of coordinate gene regulation are established. Dosage compensation has been best studied in mammals, flies and worms. Although dosage compensation systems are seemingly diverse across species, there are key shared principles of nucleation and spreading that are critical for accurate targeting of the dosage compensation complex to the X-chromosome(s). We will highlight the mechanisms by which long non-coding RNAs function together with DNA sequence elements to tether dosage compensation complexes to the X-chromosome. This article is part of a Special Issue entitled: Chromatin and epigenetic regulation of animal development.     

Random X-chromosome inactivation: skewing lessons for mice and men

The mammalian X-chromosome exists in two flavors, active and inactive, in each cell of the adult female. This phenomenon originates from the process of random choice occurring early in development in a small number of progenitor cells in which the decision is made to inactivate either one or the other X chromosome on a cell-autonomous basis. Once made, this initial decision is irreversible, although exceptions exist in specific chromosomal territories and cell lineages. Recent findings implicate various factors, including non-coding RNAs and chromatin modification complexes, as effectors in the initiation and maintenance of X-chromosome inactivation. The functional redundancy of such factors almost certainly plays an important role in the stability of the inactive X. Studying skewing or bias opens an important opportunity for understanding facets of the random choice process.     

A YY1 bridge for X inactivation

A landmark genomics project is taking shape in Africa that shifts the power and prominence to local scientists. If successful, the program will offer valuable insights into the inheritance of common diseases and reshape the paradigm of foreign-funded research.