342 research outputs found
Efficiency of Xist-mediated silencing on autosomes is linked to chromosomal domain organisation
BACKGROUND: X chromosome inactivation, the mechanism used by mammals to equalise dosage of X-linked genes in XX females relative to XY males, is triggered by chromosome-wide localisation of a cis-acting non-coding RNA, Xist. The mechanism of Xist RNA spreading and Xist-dependent silencing is poorly understood. A large body of evidence indicates that silencing is more efficient on the X chromosome than on autosomes, leading to the idea that the X chromosome has acquired sequences that facilitate propagation of silencing. LINE-1 (L1) repeats are relatively enriched on the X chromosome and have been proposed as candidates for these sequences. To determine the requirements for efficient silencing we have analysed the relationship of chromosome features, including L1 repeats, and the extent of silencing in cell lines carrying inducible Xist transgenes located on one of three different autosomes. RESULTS: Our results show that the organisation of the chromosome into large gene-rich and L1-rich domains is a key determinant of silencing efficiency. Specifically genes located in large gene-rich domains with low L1 density are relatively resistant to Xist-mediated silencing whereas genes located in gene-poor domains with high L1 density are silenced more efficiently. These effects are observed shortly after induction of Xist RNA expression, suggesting that chromosomal domain organisation influences establishment rather than long-term maintenance of silencing. The X chromosome and some autosomes have only small gene-rich L1-depleted domains and we suggest that this could confer the capacity for relatively efficient chromosome-wide silencing. CONCLUSIONS: This study provides insight into the requirements for efficient Xist mediated silencing and specifically identifies organisation of the chromosome into gene-rich L1-depleted and gene-poor L1-dense domains as a major influence on the ability of Xist-mediated silencing to be propagated in a continuous manner in cis
T cell lineage choice and differentiation in the absence of the RNase III enzyme dicer
The ribonuclease III enzyme Dicer is essential for the processing of micro-RNAs (miRNAs) and small interfering RNAs (siRNAs) from double-stranded RNA precursors. miRNAs and siRNAs regulate chromatin structure, gene transcription, mRNA stability, and translation in a wide range of organisms. To provide a model system to explore the role of Dicer-generated RNAs in the differentiation of mammalian cells in vivo, we have generated a conditional Dicer allele. Deletion of Dicer at an early stage of T cell development compromised the survival of alphabeta lineage cells, whereas the numbers of gammadelta-expressing thymocytes were not affected. In developing thymocytes, Dicer was not required for the maintenance of transcriptional silencing at pericentromeric satellite sequences (constitutive heterochromatin), the maintenance of DNA methylation and X chromosome inactivation in female cells (facultative heterochromatin), and the stable shutdown of a developmentally regulated gene (developmentally regulated gene silencing). Most remarkably, given that one third of mammalian mRNAs are putative miRNA targets, Dicer seems to be dispensable for CD4/8 lineage commitment, a process in which epigenetic regulation of lineage choice has been well documented. Thus, although Dicer seems to be critical for the development of the early embryo, it may have limited impact on the implementation of some lineage-specific gene expression programs
RNF12 Activates Xist and Is Essential for X Chromosome Inactivation
In somatic cells of female placental mammals, one of the two X chromosomes is transcriptionally silenced to accomplish an equal dose of X-encoded gene products in males and females. Initiation of random X chromosome inactivation (XCI) is thought to be regulated by X-encoded activators and autosomally encoded suppressors controlling Xist. Spreading of Xist RNA leads to silencing of the X chromosome in cis. Here, we demonstrate that the dose dependent X-encoded XCI activator RNF12/RLIM acts in trans and activates Xist. We did not find evidence for RNF12-mediated regulation of XCI through Tsix or the Xist intron 1 region, which are both known to be involved in inhibition of Xist. In addition, we found that Xist intron 1, which contains a pluripotency factor binding site, is not required for suppression of Xist in undifferentiated ES cells. Analysis of female Rnf12−/− knockout ES cells showed that RNF12 is essential for initiation of XCI and is mainly involved in the regulation of Xist. We conclude that RNF12 is an indispensable factor in up-regulation of Xist transcription, thereby leading to initiation of random XCI
A Dual Origin of the Xist Gene from a Protein-Coding Gene and a Set of Transposable Elements
X-chromosome inactivation, which occurs in female eutherian mammals is controlled by a complex X-linked locus termed the X-inactivation center (XIC). Previously it was proposed that genes of the XIC evolved, at least in part, as a result of pseudogenization of protein-coding genes. In this study we show that the key XIC gene Xist, which displays fragmentary homology to a protein-coding gene Lnx3, emerged de novo in early eutherians by integration of mobile elements which gave rise to simple tandem repeats. The Xist gene promoter region and four out of ten exons found in eutherians retain homology to exons of the Lnx3 gene. The remaining six Xist exons including those with simple tandem repeats detectable in their structure have similarity to different transposable elements. Integration of mobile elements into Xist accompanies the overall evolution of the gene and presumably continues in contemporary eutherian species. Additionally we showed that the combination of remnants of protein-coding sequences and mobile elements is not unique to the Xist gene and is found in other XIC genes producing non-coding nuclear RNA
Further examination of the Xist promoter-switch hypothesis in X inactivation: Evidence against the existence and function of a P0 promoter
Dicer regulates Xist promoter methylation in ES cells indirectly through transcriptional control of Dnmt3a
<p>Abstract</p> <p>Background</p> <p>X chromosome inactivation is the mechanism used in mammals to achieve dosage compensation of X-linked genes in XX females relative to XY males. Chromosome silencing is triggered in <it>cis </it>by expression of the non-coding RNA <it>Xist</it>. As such, correct regulation of the <it>Xist </it>gene promoter is required to establish appropriate X chromosome activity both in males and females. Studies to date have demonstrated co-transcription of an antisense RNA <it>Tsix </it>and low-level sense transcription prior to onset of X inactivation. The balance of sense and antisense RNA is important in determining the probability that a given <it>Xist </it>allele will be expressed, termed the X inactivation choice, when X inactivation commences.</p> <p>Results</p> <p>Here we investigate further the mechanism of <it>Xist </it>promoter regulation. We demonstrate that both sense and antisense transcription modulate <it>Xist </it>promoter DNA methylation in undifferentiated embryonic stem (ES) cells, suggesting a possible mechanistic basis for influencing X chromosome choice. Given the involvement of sense and antisense RNAs in promoter methylation, we investigate a possible role for the RNA interference (RNAi) pathway. We show that the <it>Xist </it>promoter is hypomethylated in ES cells deficient for the essential RNAi enzyme Dicer, but that this effect is probably a secondary consequence of reduced levels of <it>de novo </it>DNA methyltransferases in these cells. Consistent with this we find that Dicer-deficient XY and XX embryos show appropriate <it>Xist </it>expression patterns, indicating that Xist gene regulation has not been perturbed.</p> <p>Conclusion</p> <p>We conclude that <it>Xist </it>promoter methylation prior to the onset of random X chromosome inactivation is influenced by relative levels of sense and antisense transcription but that this probably occurs independent of the RNAi pathway. We discuss the implications for this data in terms of understanding <it>Xist </it>gene regulation and X chromosome choice in random X chromosome inactivation.</p
Relative judgement is relatively difficult: evidence against the role of relative judgement in absolute identification
A variety of processes have been put forward to explain absolute identification performance. One difference between current models of absolute identification is the extent to which the task involves accessing stored representations in long-term memory (e.g. exemplars in memory, Kent & Lamberts, Journal of Experimental Psychology: Learning Memory and Cognition, 31, 289–305, 2005) or relative judgement (comparison of the current stimulus to the stimulus on the previous trial, Stewart, Brown & Chater, Psychological Review, 112, 881–911, 2005). In two experiments we explored this by tapping into these processes. In Experiment 1 participants completed an absolute identification task using eight line lengths whereby a single stimulus was presented on each trial for identification. They also completed a matching task aimed at mirroring exemplar comparison in which eight line lengths were presented in a circular array and the task was to report which of these matched a target presented centrally. Experiment 2 was a relative judgement task and was similar to Experiment 1 except that the task was to report the difference (jump-size) between the current stimulus and that on the previous trial. The absolute identification and matching data showed clear similarities (faster and more accurate responding for stimuli near the edges of the range and similar stimulus-response confusions). In contrast, relative judgment performance was poor suggesting relative judgement is not straightforward. Moreover, performance as a function of jump-size differed considerably between the relative judgement and absolute identification tasks. Similarly, in the relative judgement task, predicting correct stimulus identification based on successful relative judgement yielded the reverse pattern of performance observed in the absolute identification task. Overall, the data suggest that relative judgement does not underlie absolute identification and that the task is more likely reliant on an exemplar comparison process
GYNOCARE Update: Modern Strategies to Improve Diagnosis and Treatment of Rare Gynecologic Tumors—Current Challenges and Future Directions
More than 50% of all gynecologic tumors can be classified as rare (defined as an incidence of ≤6 per 100,000 women) and usually have a poor prognosis owing to delayed diagnosis and treatment. In contrast to almost all other common solid tumors, the treatment of rare gynecologic tumors (RGT) is often based on expert opinion, retrospective studies, or extrapolation from other tumor sites with similar histology, leading to difficulty in developing guidelines for clinical practice. Currently, gynecologic cancer research, due to distinct scientific and technological challenges, is lagging behind. Moreover, the overall efforts for addressing these challenges are fragmented across different European countries and indeed, worldwide. The GYNOCARE, COST Action CA18117 (European Network for Gynecological Rare Cancer Research) programme aims to address these challenges through the creation of a unique network between key stakeholders covering distinct domains from concept to cure: basic research on RGT, biobanking, bridging with industry, and setting up the legal and regulatory requirements for international innovative clinical trials. On this basis, members of this COST Action, (Working Group 1, “Basic and Translational Research on Rare Gynecological Cancer”) have decided to focus their future efforts on the development of new approaches to improve the diagnosis and treatment of RGT. Here, we provide a brief overview of the current state-of-the-art and describe the goals of this COST Action and its future challenges with the aim to stimulate discussion and promote synergy across scientists engaged in the fight against this rare cancer worldwide
HLA-DR Alpha 2 Mediates Negative Signalling via Binding to Tirc7 Leading to Anti-Inflammatory and Apoptotic Effects in Lymphocytes In Vitro and In Vivo
Classically, HLA-DR expressed on antigen presenting cells (APC) initiates lymphocyte activation via presentation of peptides to TCR bearing CD4+ T-Cells. Here we demonstrate that HLA-DR alpha 2 domain (sHLA-DRα2) also induces negative signals by engaging TIRC7 on lymphocytes. This interaction inhibits proliferation and induces apoptosis in CD4+ and CD8+ T-cells via activation of the intrinsic pathway. Proliferation inhibition is associated with SHP-1 recruitment by TIRC7, decreased phosphorylation of STAT4, TCR-ζ chain & ZAP70, and inhibition of IFN-γ and FasL expression. HLA-DRα2 and TIRC7 co-localize at the APC-T cell interaction site. Triggering HLA-DR - TIRC7 pathway demonstrates that sHLA-DRα2 treatment inhibits proinflammatory-inflammatory cytokine expression in APC & T cells after lipopolysaccaride (LPS) stimulation in vitro and induces apoptosis in vivo. These results suggest a novel antiproliferative role for HLA-DR mediated via TIRC7, revise the notion of an exclusive stimulatory interaction of HLA-DR with CD4+ T cells and highlights a novel physiologically relevant regulatory pathway
Histone acetylation controls the inactive X chromosome replication dynamics
In mammals, dosage compensation between male and female cells is achieved by inactivating one female X chromosome (Xi). Late replication of Xi was proposed to be involved in the maintenance of its silenced state. Here, we show a highly synchronous replication of the Xi within 1 to 2 h during early-mid S-phase by following DNA replication in living mammalian cells with green fluorescent protein-tagged replication proteins. The Xi was replicated before or concomitant with perinuclear or perinucleolar facultative heterochromatin and before constitutive heterochromatin. Ectopic expression of the X-inactive-specific transcript (Xist) gene from an autosome imposed the same synchronous replication pattern. We used mutations and chemical inhibition affecting different epigenetic marks as well as inducible Xist expression and we demonstrate that histone hypoacetylation has a key role in controlling Xi replication. The epigenetically controlled, highly coordinated replication of the Xi is reminiscent of embryonic genome replication in flies and frogs before genome activation and might be a common feature of transcriptionally silent chromatin
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