UMR3244 – Dynamique de l’information génétique

Publications de l’équipe

Année de publication : 2019

Emilia Puig Lombardi, Allyson Holmes, Daniela Verga, Marie-Paule Teulade-Fichou, Alain Nicolas, Arturo Londoño-Vallejo (2019 Jul 9)

Thermodynamically stable and genetically unstable G-quadruplexes are depleted in genomes across species.

Nucleic acids research : 47 : 6098-6113 : DOI : 10.1093/nar/gkz463 En savoir plus
Résumé

G-quadruplexes play various roles in multiple biological processes, which can be positive when a G4 is involved in the regulation of gene expression or detrimental when the folding of a stable G4 impairs DNA replication promoting genome instability. This duality interrogates the significance of their presence within genomes. To address the potential biased evolution of G4 motifs, we analyzed their occurrence, features and polymorphisms in a large spectrum of species. We found extreme bias of the short-looped G4 motifs, which are the most thermodynamically stable in vitro and thus carry the highest folding potential in vivo. In the human genome, there is an over-representation of single-nucleotide-loop G4 motifs (G4-L1), which are highly conserved among humans and show a striking excess of the thermodynamically least stable G4-L1A (G3AG3AG3AG3) sequences. Functional assays in yeast showed that G4-L1A caused the lowest levels of both spontaneous and G4-ligand-induced instability. Analyses across 600 species revealed the depletion of the most stable G4-L1C/T quadruplexes in most genomes in favor of G4-L1A in vertebrates or G4-L1G in other eukaryotes. We discuss how these trends might be the result of species-specific mutagenic processes associated to a negative selection against the most stable motifs, thus neutralizing their detrimental effects on genome stability while preserving positive G4-associated biological roles.

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Kevin Grosselin, Adeline Durand, Justine Marsolier, Adeline Poitou, Elisabetta Marangoni, Fariba Nemati, Ahmed Dahmani, Sonia Lameiras, Fabien Reyal, Olivia Frenoy, Yannick Pousse, Marcel Reichen, Adam Woolfe, Colin Brenan, Andrew D. Griffiths*, Céline Vallot* & Annabelle Gérard* (2019 May 31)

High-throughput single-cell ChIP-seq identifies heterogeneity of chromatin states in breast cancer

Nature Genetics : 1060–1066 : DOI : 10.1038/s41588-019-0424-9 En savoir plus
Résumé

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Année de publication : 2018

Antoine Hocher, Myriam Ruault, Petra Kaferle, Marc Descrimes, Mickaël Garnier, Antonin Morillon, Angela Taddei (2018 Oct 26)

Expanding heterochromatin reveals discrete subtelomeric domains delimited by chromatin landscape transitions.

Genome research : DOI : gr.236554.118 En savoir plus
Résumé

The eukaryotic genome is divided into chromosomal domains of heterochromatin and euchromatin. Transcriptionally silent heterochromatin is found at subtelomeric regions, leading to the telomeric position effect (TPE) in yeast fly and human. Heterochromatin generally initiates and spreads from defined loci, and diverse mechanisms prevent the ectopic spread of heterochromatin into euchromatin. Here, we overexpressed the silencing factor Sir3 at varying levels in yeast and found that Sir3 spreads into Extended Silent Domains (ESDs), eventually reaching saturation at subtelomeres. We observed the spread of Sir3 into subtelomeric domains associated with specific histone marks in wild-type cells and stopping at zones of histone mark transitions including H3K79 tri-methylation levels. Our study shows that the conserved H3K79 methyltransferase Dot1 is essential in restricting Sir3 spread beyond ESDs, thus ensuring viability upon overexpression of Sir3. Lastly, our analyses of published data demonstrate how ESDs unveil uncharacterized discrete domains isolating structural and functional subtelomeric features from the rest of the genome. Our work offers a new approach on how to separate subtelomeres from the core chromosome.

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Année de publication : 2017

Wery M, Gautier C, Descrimes M, Yoda M, Vennin-Rendos H, Migeot V, Gautheret D, Hermand D, Morillon A (2017 Nov 7)

Native Elongating Transcript Sequencing reveals global anti-correlation between sense and antisense nascent transcription in fission yeast

RNA : DOI : 10.1261/rna.063446.117 En savoir plus
Résumé

Antisense transcription can regulate sense gene expression. However, previous annotations of antisense transcription units have been based on detection of mature antisense long non-coding (aslnc)RNAs by RNA-Seq and/or micro-arrays, only giving a partial view of the antisense transcription landscape and incomplete molecular bases for antisense-mediated regulation. Here, we used Native Elongating Transcript sequencing to map genome-wide nascent antisense transcription in fission yeast. Strikingly, antisense transcription was detected for most protein-coding genes, correlating with low sense transcription, especially when overlapping the mRNA start site. RNA profiling revealed that the resulting aslncRNAs mainly correspond to cryptic Xrn1/Exo2-sensitive transcripts (XUTs). ChIP-Seq analyses showed that antisense (as)XUTs expression is associated with specific histone modifications patterns. Finally, we showed that asXUTs are controlled by the histone chaperone Spt6 and respond to meiosis induction, in both cases anti-correlating with levels of the paired-sense mRNAs, supporting physiological significance to antisense-mediated gene attenuation. Our work highlights that antisense transcription is much more extended than anticipated and might constitute an additional non-promoter determinant of gene regulation complexity.

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Jarroux J, Morillon A, Pinskaya M. (2017 Oct 1)

History, Discovery, and Classification of lncRNAs

Adv Exp Med BiolLong non coding RNA biology : 1008 : 1-46 : DOI : 10.1007/978-981-10-5203-3_1 En savoir plus
Résumé

The RNA World Hypothesis suggests that prebiotic life revolved around RNA instead of DNA and proteins. Although modern cells have changed significantly in 4 billion years, RNA has maintained its central role in cell biology. Since the discovery of DNA at the end of the nineteenth century, RNA has been extensively studied. Many discoveries such as housekeeping RNAs (rRNA, tRNA, etc.) supported the messenger RNA model that is the pillar of the central dogma of molecular biology, which was first devised in the late 1950s. Thirty years later, the first regulatory non-coding RNAs (ncRNAs) were initially identified in bacteria and then in most eukaryotic organisms. A few long ncRNAs (lncRNAs) such as H19 and Xist were characterized in the pre-genomic era but remained exceptions until the early 2000s. Indeed, when the sequence of the human genome was published in 2001, studies showed that only about 1.2% encodes proteins, the rest being deemed « non-coding. » It was later shown that the genome is pervasively transcribed into many ncRNAs, but their functionality remained controversial. Since then, regulatory lncRNAs have been characterized in many species and were shown to be involved in processes such as development and pathologies, revealing a new layer of regulation in eukaryotic cells. This newly found focus on lncRNAs, together with the advent of high-throughput sequencing, was accompanied by the rapid discovery of many novel transcripts which were further characterized and classified according to specific transcript traits.In this review, we will discuss the many discoveries that led to the study of lncRNAs, from Friedrich Miescher’s « nuclein » in 1869 to the elucidation of the human genome and transcriptome in the early 2000s. We will then focus on the biological relevance during lncRNA evolution and describe their basic features as genes and transcripts. Finally, we will present a non-exhaustive catalogue of lncRNA classes, thus illustrating the vast complexity of eukaryotic transcriptomes

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Dan Filipescu, Monica Naughtin, Katrina Podsypanina, Vincent Lejour, Laurence Wilson, Zachary A Gurard-Levin, Guillermo A Orsi, Iva Simeonova, Eleonore Toufektchan, Laura D Attardi, Franck Toledo, Geneviève Almouzni (2017 Mar 31)

Essential role for centromeric factors following p53 loss and oncogenic transformation.

Genes & development : 463-480 : DOI : 10.1101/gad.290924.116 En savoir plus
Résumé

In mammals, centromere definition involves the histone variant CENP-A (centromere protein A), deposited by its chaperone, HJURP (Holliday junction recognition protein). Alterations in this process impair chromosome segregation and genome stability, which are also compromised by p53 inactivation in cancer. Here we found that CENP-A and HJURP are transcriptionally up-regulated in p53-null human tumors. Using an established mouse embryonic fibroblast (MEF) model combining p53 inactivation with E1A or HRas-V12 oncogene expression, we reproduced a similar up-regulation of HJURP and CENP-A. We delineate functional CDE/CHR motifs within the Hjurp and Cenpa promoters and demonstrate their roles in p53-mediated repression. To assess the importance of HJURP up-regulation in transformed murine and human cells, we used a CRISPR/Cas9 approach. Remarkably, depletion of HJURP leads to distinct outcomes depending on their p53 status. Functional p53 elicits a cell cycle arrest response, whereas, in p53-null transformed cells, the absence of arrest enables the loss of HJURP to induce severe aneuploidy and, ultimately, apoptotic cell death. We thus tested the impact of HJURP depletion in pre-established allograft tumors in mice and revealed a major block of tumor progression in vivo. We discuss a model in which an « epigenetic addiction » to the HJURP chaperone represents an Achilles’ heel in p53-deficient transformed cells.

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Boris Bardot, Franck Toledo (2017 Feb 24)

Targeting MDM4 Splicing in Cancers.

Genes : DOI : E82 En savoir plus
Résumé

MDM4, an essential negative regulator of the P53 tumor suppressor, is frequently overexpressed in cancer cells that harbor a wild-type P53. By a mechanism based on alternative splicing, the MDM4 gene generates two mutually exclusive isoforms: MDM4-FL, which encodes the full-length MDM4 protein, and a shorter splice variant called MDM4-S. Previous results suggested that the MDM4-S isoform could be an important driver of tumor development. In this short review, we discuss a recent set of data indicating that MDM4-S is more likely a passenger isoform during tumorigenesis and that targeting MDM4 splicing to prevent MDM4-FL protein expression appears as a promising strategy to reactivate p53 in cancer cells. The benefits and risks associated with this strategy are also discussed.

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Eléonore Toufektchan, Sara Jaber, Franck Toledo (2017 Jan 26)

[Dangerous liaisons: p53, dyskeratosis congenita and Fanconi anemia].

Medecine sciences : M/S : 95-98 : DOI : 10.1051/medsci/20173301018 En savoir plus
Résumé

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Bruno Teste, Jerome Champ, Arturo Londono-Vallejo, Stéphanie Descroix, Laurent Malaquin, Jean-Louis Viovy, Irena Draskovic, Guillaume Mottet (2017 Jan 17)

Chromatin immunoprecipitation in microfluidic droplets: towards fast and cheap analyses.

Lab on a chip : 530-537 : DOI : 10.1039/c6lc01535b En savoir plus
Résumé

Genetic organization is governed by the interaction of DNA with histone proteins, and differential modifications of these proteins is a fundamental mechanism of gene regulation. Histone modifications are primarily studied through chromatin immunoprecipitation (ChIP) assays, however conventional ChIP procedures are time consuming, laborious and require a large number of cells. Here we report for the first time the development of ChIP in droplets based on a microfluidic platform combining nanoliter droplets, magnetic beads (MB) and magnetic tweezers (MT). The droplet approach enabled compartmentalization and improved mixing, while reducing the consumption of samples and reagents in an integrated workflow. Anti-histone antibodies grafted to MB were used as a solid support to capture and transfer the target chromatin from droplets to droplets in order to perform chromatin immunoprecipitation, washing, elution and purification of DNA. We designed a new ChIP protocol to investigate four different types of modified histones with known roles in gene activation or repression. We evaluated the performances of this new ChIP in droplet assay in comparison with conventional methods. The proposed technology dramatically reduces analytical time from a few days to 7 hours, simplifies the ChIP protocol and decreases the number of cells required by 100 fold while maintaining a high degree of sensitivity and specificity. Therefore this droplet-based ChIP assay represents a new, highly advantageous and convenient approach to epigenetic analyses.

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Yann Duroc, Rajeev Kumar, Lepakshi Ranjha, Céline Adam, Raphaël Guérois, Khan Md Muntaz, Marie-Claude Marsolier-Kergoat, Florent Dingli, Raphaëlle Laureau, Damarys Loew, Bertrand Llorente, Jean-Baptiste Charbonnier, Petr Cejka, Valérie Borde (2017 Jan 5)

Concerted action of the MutLβ heterodimer and Mer3 helicase regulates the global extent of meiotic gene conversion.

eLife : DOI : 10.7554/eLife.21900 En savoir plus
Résumé

Gene conversions resulting from meiotic recombination are critical in shaping genome diversification and evolution. How the extent of gene conversions is regulated is unknown. Here we show that the budding yeast mismatch repair related MutLβ complex, Mlh1-Mlh2, specifically interacts with the conserved meiotic Mer3 helicase, which recruits it to recombination hotspots, independently of mismatch recognition. This recruitment is essential to limit gene conversion tract lengths genome-wide, without affecting crossover formation. Contrary to expectations, Mer3 helicase activity, proposed to extend the displacement loop (D-loop) recombination intermediate, does not influence the length of gene conversion events, revealing non-catalytical roles of Mer3. In addition, both purified Mer3 and MutLβ preferentially recognize D-loops, providing a mechanism for limiting gene conversion in vivo. These findings show that MutLβ is an integral part of a new regulatory step of meiotic recombination, which has implications to prevent rapid allele fixation and hotspot erosion in populations.

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Céline Vallot, Catherine Patrat, Amanda J Collier, Christophe Huret, Miguel Casanova, Tharvesh M Liyakat Ali, Matteo Tosolini, Nelly Frydman, Edith Heard, Peter J Rugg-Gunn, Claire Rougeulle (2017 Jan 5)

XACT Noncoding RNA Competes with XIST in the Control of X Chromosome Activity during Human Early Development.

Cell stem cell : 102-111 : DOI : 10.1016/j.stem.2016.10.014 En savoir plus
Résumé

Sex chromosome dosage compensation is essential in most metazoans, but the developmental timing and underlying mechanisms vary significantly, even among placental mammals. Here we identify human-specific mechanisms regulating X chromosome activity in early embryonic development. Single-cell RNA sequencing and imaging revealed co-activation and accumulation of the long noncoding RNAs (lncRNAs) XACT and XIST on active X chromosomes in both early human pre-implantation embryos and naive human embryonic stem cells. In these contexts, the XIST RNA adopts an unusual, highly dispersed organization, which may explain why it does not trigger X chromosome inactivation at this stage. Functional studies in transgenic mouse cells show that XACT influences XIST accumulation in cis. Our findings therefore suggest a mechanism involving antagonistic activity of XIST and XACT in controlling X chromosome activity in early human embryos, and they highlight the contribution of rapidly evolving lncRNAs to species-specific developmental mechanisms.

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Année de publication : 2016

Robert Jackson, Bruce A Rosa, Sonia Lameiras, Sean Cuninghame, Josee Bernard, Wely B Floriano, Paul F Lambert, Alain Nicolas, Ingeborg Zehbe (2016 Nov 4)

Functional variants of human papillomavirus type 16 demonstrate host genome integration and transcriptional alterations corresponding to their unique cancer epidemiology.

BMC genomics : 851 En savoir plus
Résumé

Human papillomaviruses (HPVs) are a worldwide burden as they are a widespread group of tumour viruses in humans. Having a tropism for mucosal tissues, high-risk HPVs are detected in nearly all cervical cancers. HPV16 is the most common high-risk type but not all women infected with high-risk HPV develop a malignant tumour. Likely relevant, HPV genomes are polymorphic and some HPV16 single nucleotide polymorphisms (SNPs) are under evolutionary constraint instigating variable oncogenicity and immunogenicity in the infected host.

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Pierre Thouvenot, Lou Fourrière, Elodie Dardillac, Barbara Ben Yamin, Aurianne Lescure, Vincent Lejour, Xavier Heiligenstein, Jean-Baptiste Boulé, Maryse Romao, Graça Raposo-Benedetti, Bernard S Lopez, Alain Nicolas, Gaël A Millot (2016 Nov 2)

Yeast cells reveal the misfolding and the cellular mislocalisation of the human BRCA1 protein.

Journal of cell science : DOI : jcs.192880 En savoir plus
Résumé

Understanding the effect of an ever-growing number of human variants detected by genome sequencing is a medical challenge. The yeast Saccharomyces cerevisiae model has held attention for its capacity to monitor the functional impact of missense mutations found in human genes, including the BRCA1 breast/ovarian cancer susceptibility gene. When expressed in yeast, the wild-type full-length BRCA1 protein forms a single nuclear aggregate and induces a growth inhibition. Both events are modified by pathogenic mutations of BRCA1. However, the biological interpretation of these events remains to be determined. Here, we show that the BRCA1 nuclear aggregation and the growth inhibition are sensitive to misfolding effects induced by missense mutations. Moreover, misfolding mutations impair the nuclear targeting of BRCA1 in yeast cells and in a human cell line. In conclusion, we establish a connection between misfolding and nuclear transport impairment and we illustrate that yeast is a suitable model to decipher the effect of misfolding mutations.

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Loic Verlingue, Aurélien Dugourd, Gautier Stoll, Emmanuel Barillot, Laurence Calzone, Arturo Londoño-Vallejo (2016 Sep 11)

A comprehensive approach to the molecular determinants of lifespan using a Boolean model of geroconversion.

Aging cell : DOI : 10.1111/acel.12504 En savoir plus
Résumé

Altered molecular responses to insulin and growth factors (GF) are responsible for late-life shortening diseases such as type-2 diabetes mellitus (T2DM) and cancers. We have built a network of the signaling pathways that control S-phase entry and a specific type of senescence called geroconversion. We have translated this network into a Boolean model to study possible cell phenotype outcomes under diverse molecular signaling conditions. In the context of insulin resistance, the model was able to reproduce the variations of the senescence level observed in tissues related to T2DM’s main morbidity and mortality. Furthermore, by calibrating the pharmacodynamics of mTOR inhibitors, we have been able to reproduce the dose-dependent effect of rapamycin on liver degeneration and lifespan expansion in wild-type and HER2-neu mice. Using the model, we have finally performed an in silico prospective screen of the risk-benefit ratio of rapamycin dosage for healthy lifespan expansion strategies. We present here a comprehensive prognostic and predictive systems biology tool for human aging.

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Franck Toledo (2016 Aug 20)

p53: A two-faced regulator of telomere metabolism? (comment on DOI 10.1002/bies.201600078).

BioEssays : news and reviews in molecular, cellular and developmental biology : DOI : 10.1002/bies.201600149 En savoir plus
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