Telomeres in ICF syndrome cells are vulnerable to DNA damage due to elevated DNA:RNA hybrids.

DNA:RNA hybrids, nucleic acid structures with diverse physiological functions, can disrupt genome integrity when dysregulated. Human telomeres were shown to form hybrids with the lncRNA TERRA, yet the formation and distribution of these hybrids among telomeres, their regulation and their cellular effects remain elusive. Here we predict and confirm in several human cell types that DNA:RNA hybrids form at many subtelomeric and telomeric regions. We demonstrate that ICF syndrome cells, which exhibit short telomeres and elevated TERRA levels, are enriched for hybrids at telomeric regions throughout the cell cycle. Telomeric hybrids are associated with high levels of DNA damage at chromosome ends in ICF cells, which are significantly reduced with overexpression of RNase H1. Our findings suggest that abnormally high TERRA levels in ICF syndrome lead to accumulation of telomeric hybrids that, in turn, can result in telomeric dysfunction

Germline genes hypomethylation and expression define a molecular signature in peripheral blood of ICF patients: implications for diagnosis and etiology.

International audienceBACKGROUND: Immunodeficiency Centromeric Instability and Facial anomalies (ICF) is a rare autosomal recessive disease characterized by reduction in serum immunoglobulins with severe recurrent infections, facial dysmorphism, and more variable symptoms including mental retardation. ICF is directly related to a genomic methylation defect that mainly affects juxtacentromeric heterochromatin regions of certain chromosomes, leading to chromosomal rearrangements that constitute a hallmark of this syndrome upon cytogenetic testing. Mutations in the de novo DNA methyltransferase DNMT3B, the protein ZBTB24 of unknown function, or loci that remain to be identified, lie at its origin. Despite unifying features, common or distinguishing molecular signatures are still missing for this disease. METHOD: We used the molecular signature that we identified in a mouse model for ICF1 to establish transcriptional biomarkers to facilitate diagnosis and understanding of etiology of the disease. We assayed the expression and methylation status of a set of genes whose expression is normally restricted to germ cells, directly in whole blood samples and epithelial cells of ICF patients. RESULTS: We report that DNA hypomethylation and expression of MAEL and SYCE1 represent robust biomarkers, easily testable directly from uncultured cells to diagnose the most prevalent sub-type of the syndrome. In addition, we identified the first unifying molecular signatures for ICF patients. Of importance, we validated the use of our biomarkers to diagnose a baby born to a family with a sick child. Finally, our analysis revealed unsuspected complex molecular signatures in two ICF patients suggestive of a novel genetic etiology for the disease. CONCLUSIONS: Early diagnosis of ICF syndrome is crucial since early immunoglobulin supplementation can improve the course of disease. However, ICF is probably underdiagnosed, especially in patients that present with incomplete phenotype or born to families with no affected relatives. The specific and robust biomarkers identified in this study could be introduced into routine clinical immunology or neurology departments to facilitate testing of patients with suspected ICF syndrome. In addition, as exemplified by two patients with a combination of molecular defects never described before, our data support the search for new types of mutations at the origin of ICF syndrome

Non-coding murine centromeric transcripts associate with and potentiate Aurora B kinase

Non-coding RNAs are emerging as key players in many fundamental biological processes, including specification of higher-order chromatin structure. We examined the implication of RNA transcribed from mouse centromeric minor satellite repeats in the formation and function of centromere-associated complexes. Here we show that the levels of minor satellite RNA vary during cell-cycle progression, peaking in G2/M phase, concomitant with accumulation of proteins of the chromosomal passenger complex near the centromere. Consistent with this, we describe that murine minor satellite RNA are components of CENP-A-associated centromeric fractions and associate with proteins of the chromosomal passenger complex Aurora B and Survivin at the onset of mitosis. Interactions of endogenous Aurora B with CENP-A and Survivin are sensitive to RNaseA. Likewise, the kinase activity of Aurora B requires an RNA component. More importantly, Aurora B kinase activity can be potentiated by minor satellite RNA. In addition, decreased Aurora B activity after RNA depletion can be specifically rescued by restitution of these transcripts. Together, our data provide new functional evidence for minor satellite transcripts as key partners and regulators of the mitotic kinase Aurora B

Steroid receptor RNA activator protein binds to and counteracts SRA RNA-mediated activation of MyoD and muscle differentiation

The steroid receptor RNA activator (SRA) has the unusual property to function as both a non-coding RNA (ncRNA) and a protein SRAP. SRA ncRNA is known to increase the activity of a range of nuclear receptors as well as the master regulator of muscle differentiation MyoD. The contribution of SRA to either a ncRNA or a protein is influenced by alternative splicing of the first intron, the retention of which disrupts the SRAP open reading frame. We reported here that the ratio between non-coding and coding SRA isoforms increased during myogenic differentiation of human satellite cells but not myotonic dystrophy patient satellite cells, in which differentiation capacity is affected. Using constructs that exclusively produce SRA ncRNA or SRAP, we demonstrated that whereas SRA ncRNA was indeed an enhancer of myogenic differentiation and myogenic conversion of non-muscle cells through the co-activation of MyoD activity, SRAP prevented this SRA RNA-dependant co-activation. Interestingly, the SRAP inhibitory effect is mediated through the interaction of SRAP with its RNA counterpart via its RRM-like domain interacting with the functional sub-structure of SRA RNA, STR7. This study thus provides a new model for SRA-mediated regulation of MyoD transcriptional activity in the promotion of normal muscle differentiation, which takes into account the nature of SRA molecules present

Evaluation of DNA Methylation Episignatures for Diagnosis and Phenotype Correlations in 42 Mendelian Neurodevelopmental Disorders.

Genetic syndromes frequently present with overlapping clinical features and inconclusive or ambiguous genetic findings which can confound accurate diagnosis and clinical management. An expanding number of genetic syndromes have been shown to have unique genomic DNA methylation patterns (called episignatures ). Peripheral blood episignatures can be used for diagnostic testing as well as for the interpretation of ambiguous genetic test results. We present here an approach to episignature mapping in 42 genetic syndromes, which has allowed the identification of 34 robust disease-specific episignatures. We examine emerging patterns of overlap, as well as similarities and hierarchical relationships across these episignatures, to highlight their key features as they are related to genetic heterogeneity, dosage effect, unaffected carrier status, and incomplete penetrance. We demonstrate the necessity of multiclass modeling for accurate genetic variant classification and show how disease classification using a single episignature at a time can sometimes lead to classification errors in closely related episignatures. We demonstrate the utility of this tool in resolving ambiguous clinical cases and identification of previously undiagnosed cases through mass screening of a large cohort of subjects with developmental delays and congenital anomalies. This study more than doubles the number of published syndromes with DNA methylation episignatures and, most significantly, opens new avenues for accurate diagnosis and clinical assessment in individuals affected by these disorders

Compartiments nucléaires d’hétérochromatine et répression génique : le modèle de la différenciation hématopoïétique humaine

Il est classiquement admis que l’expression des gènes est déterminée par l’interaction de facteurs de transcription avec la structure répressive de la chromatine. Plus récemment, le contexte originel de la transcription, c’est-à-dire le noyau, a été pris en considération. De nombreuses observations soutiennent que l’organisation nucléaire, définie par le positionnement non aléatoire de gènes et de protéines régulatrices, influence l’expression du génome. La relation entre structure chromatinienne, mouvements chromosomiques et destin cellulaire est encore mal comprise. Néanmoins, l’étude de l’implication de ces aspects épigénétiques dans les changements héritables des programmes d’expression génique qui accompagnent la différentiation cellulaire est un domaine en plein essor. Le compartiment d’hétérochromatine péricentromérique sera présenté comme un des exemples les mieux étudiés pour comprendre l’impact de l’organisation nucléaire et du positionnement de gènes sur leur transcription. Nous replacerons l’influence des compartiments d’hétérochromatine dans le contexte de la différenciation de progéniteurs hématopoïétiques multipotents humains

DNA methylation in satellite repeats disorders

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Coding and Non-coding RNAs, the Frontier Has Never Been So Blurred

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