ARN non-codant, épigénétique et fluidité du génome

Publications de l’équipe

Année de publication : 2019

Aria Ronsmans, Maxime Wery, Ugo Szachnowski, Camille Gautier, Marc Descrimes, Evelyne Dubois, Antonin Morillon, Isabelle Georis (2019 Mar 1)

Transcription-dependent spreading of the Dal80 yeast GATA factor across the body of highly expressed genes.

PLoS genetics : e1007999 : DOI : 10.1371/journal.pgen.1007999 En savoir plus
Résumé

GATA transcription factors are highly conserved among eukaryotes and play roles in transcription of genes implicated in cancer progression and hematopoiesis. However, although their consensus binding sites have been well defined in vitro, the in vivo selectivity for recognition by GATA factors remains poorly characterized. Using ChIP-Seq, we identified the Dal80 GATA factor targets in yeast. Our data reveal Dal80 binding to a large set of promoters, sometimes independently of GATA sites, correlating with nitrogen- and/or Dal80-sensitive gene expression. Strikingly, Dal80 was also detected across the body of promoter-bound genes, correlating with high expression. Mechanistic single-gene experiments showed that Dal80 spreading across gene bodies requires active transcription. Consistently, Dal80 co-immunoprecipitated with the initiating and post-initiation forms of RNA Polymerase II. Our work suggests that GATA factors could play dual, synergistic roles during transcription initiation and post-initiation steps, promoting efficient remodeling of the gene expression program in response to environmental changes.

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Bingning Xie, Emmanuelle Becker, Igor Stuparevic, Maxime Wery, Ugo Szachnowski, Antonin Morillon, Michael Primig (2019 Feb 15)

The anti-cancer drug 5-fluorouracil affects cell cycle regulators and potential regulatory long non-coding RNAs in yeast.

RNA biology : 1-15 : DOI : 10.1080/15476286.2019.1581596 En savoir plus
Résumé

5-fluorouracil (5-FU) was isolated as an inhibitor of thymidylate synthase, which is important for DNA synthesis. The drug was later found to also affect the conserved 3′-5′ exoribonuclease EXOSC10/Rrp6, a catalytic subunit of the RNA exosome that degrades and processes protein-coding and non-coding transcripts. Work on 5-FU’s cytotoxicity has been focused on mRNAs and non-coding transcripts such as rRNAs, tRNAs and snoRNAs. However, the effect of 5-FU on long non-coding RNAs (lncRNAs), which include regulatory transcripts important for cell growth and differentiation, is poorly understood. RNA profiling of synchronized 5-FU treated yeast cells and protein assays reveal that the drug specifically inhibits a set of cell cycle regulated genes involved in mitotic division, by decreasing levels of the paralogous Swi5 and Ace2 transcriptional activators. We also observe widespread accumulation of different lncRNA types in treated cells, which are typically present at high levels in a strain lacking EXOSC10/Rrp6. 5-FU responsive lncRNAs include potential regulatory antisense transcripts that form double-stranded RNAs (dsRNAs) with overlapping sense mRNAs. Some of these transcripts encode proteins important for cell growth and division, such as the transcription factor Ace2, and the RNA exosome subunit EXOSC6/Mtr3. In addition to revealing a transcriptional effect of 5-FU action via DNA binding regulators involved in cell cycle progression, our results have implications for the function of putative regulatory lncRNAs in 5-FU mediated cytotoxicity. The data raise the intriguing possibility that the drug deregulates lncRNAs/dsRNAs involved in controlling eukaryotic cell division, thereby highlighting a new class of promising therapeutical targets.

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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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Watts BR, Wittmann S, Wery M, Gautier C, Kus K, Birot A, Heo DH, Kilchert C, Morillon A, Vasiljeva L (2018 Mar 26)

Histone deacetylation promotes transcriptional silencing at facultative heterochromatin

Nucleic Acid ResearchHistone deacetylation promotes transcriptional silencing at facultative heterochromatin : DOI : 10.1093/nar/gky232 En savoir plus
Résumé

It is important to accurately regulate the expression of genes involved in development and environmental response. In the fission yeast Schizosaccharomyces pombe, meiotic genes are tightly repressed during vegetative growth. Despite being embedded in heterochromatin these genes are transcribed and believed to be repressed primarily at the level of RNA. However, the mechanism of facultative heterochromatin formation and the interplay with transcription regulation is not understood. We show genome-wide that HDAC-dependent histone deacetylation is a major determinant in transcriptional silencing of facultative heterochromatin domains. Indeed, mutation of class I/II HDACs leads to increased transcription of meiotic genes and accumulation of their mRNAs. Mechanistic dissection of the pho1 gene where, in response to phosphate, transient facultative heterochromatin is established by overlapping lncRNA transcription shows that the Clr3 HDAC contributes to silencing independently of SHREC, but in an lncRNA-dependent manner. We propose that HDACs promote facultative heterochromatin by establishing alternative transcriptional silencing.

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Alculumbre SG, Saint-André V, Di Domizio J, Vargas P, Sirven P, Bost P, Maurin M, Maiuri P, Wery M, Roman MS, Savey L, Touzot M, Terrier B, Saadoun D, Conrad C, Gilliet M, Morillon A, Soumelis V (2018 Jan 1)

Diversification of human plasmacytoid predendritic cells in response to a single stimulus

Nature Immunology : 19(1) : 63-75 : DOI : 10.1038/s41590-017-0012-z En savoir plus
Résumé

Innate immune cells adjust to microbial and inflammatory stimuli through a process termed environmental plasticity, which links a given individual stimulus to a unique activated state. Here, we report that activation of human plasmacytoid predendritic cells (pDCs) with a single microbial or cytokine stimulus triggers cell diversification into three stable subpopulations (P1-P3). P1-pDCs (PD-L1+CD80-) displayed a plasmacytoid morphology and specialization for type I interferon production. P3-pDCs (PD-L1-CD80+) adopted a dendritic morphology and adaptive immune functions. P2-pDCs (PD-L1+CD80+) displayed both innate and adaptive functions. Each subpopulation expressed a specific coding- and long-noncoding-RNA signature and was stable after secondary stimulation. P1-pDCs were detected in samples from patients with lupus or psoriasis. pDC diversification was independent of cell divisions or preexisting heterogeneity within steady-state pDCs but was controlled by a TNF autocrine and/or paracrine communication loop. Our findings reveal a novel mechanism for diversity and division of labor in innate immune cells

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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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Année de publication : 2016

Alvaro de Andres-Pablo, Antonin Morillon, Maxime Wery (2016 May 28)

LncRNAs, lost in translation or licence to regulate?

Current genetics En savoir plus
Résumé

Over the last decade, advances in transcriptomics have revealed that the pervasive transcription of eukaryotic genomes produces plethora of long noncoding RNAs (lncRNAs), which are now recognized as major regulators of multiple cellular processes. Although they have been thought to lack any protein-coding potential, recent ribosome-profiling data indicate that lncRNAs can interact with the translation machinery, leading to the production of functional peptides in some cases. In this perspective, we have explored the idea that translation can be part of the fate of cytoplasmic lncRNAs, raising the possibility for them to work as bifunctional RNAs, endowed with dual coding and regulatory functions.

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Maxime Wery, Marc Descrimes, Nicolas Vogt, Anne-Sophie Dallongeville, Daniel Gautheret, Antonin Morillon (2016 Jan 26)

Nonsense-Mediated Decay Restricts LncRNA Levels in Yeast Unless Blocked by Double-Stranded RNA Structure.

Molecular cell : 379-92 : DOI : 10.1016/j.molcel.2015.12.020 En savoir plus
Résumé

Antisense long non-coding (aslnc)RNAs represent a substantial part of eukaryotic transcriptomes that are, in yeast, controlled by the Xrn1 exonuclease. Nonsense-Mediated Decay (NMD) destabilizes the Xrn1-sensitive aslncRNAs (XUT), but what determines their sensitivity remains unclear. We report that 3′ single-stranded (3′-ss) extension mediates XUTs degradation by NMD, assisted by the Mtr4 and Dbp2 helicases. Single-gene investigation, genome-wide RNA analyses, and double-stranded (ds)RNA mapping revealed that 3′-ss extensions discriminate the NMD-targeted XUTs from stable lncRNAs. Ribosome profiling showed that XUT are translated, locking them for NMD activity. Interestingly, mutants of the Mtr4 and Dbp2 helicases accumulated XUTs, suggesting that dsRNA unwinding is a critical step for degradation. Indeed, expression of anticomplementary transcripts protects cryptic intergenic lncRNAs from NMD. Our results indicate that aslncRNAs form dsRNA that are only translated and targeted to NMD if dissociated by Mtr4 and Dbp2. We propose that NMD buffers genome expression by discarding pervasive regulatory transcripts.

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Année de publication : 2015

Jia Li, Marie-Anne Poursat, Damien Drubay, Arnaud Motz, Zohra Saci, Antonin Morillon, Stefan Michiels, Daniel Gautheret (2015 Nov 21)

A Dual Model for Prioritizing Cancer Mutations in the Non-coding Genome Based on Germline and Somatic Events.

PLoS computational biology : e1004583 : DOI : 10.1371/journal.pcbi.1004583 En savoir plus
Résumé

We address here the issue of prioritizing non-coding mutations in the tumoral genome. To this aim, we created two independent computational models. The first (germline) model estimates purifying selection based on population SNP data. The second (somatic) model estimates tumor mutation density based on whole genome tumor sequencing. We show that each model reflects a different set of constraints acting either on the normal or tumor genome, and we identify the specific genome features that most contribute to these constraints. Importantly, we show that the somatic mutation model carries independent functional information that can be used to narrow down the non-coding regions that may be relevant to cancer progression. On this basis, we identify positions in non-coding RNAs and the non-coding parts of mRNAs that are both under purifying selection in the germline and protected from mutation in tumors, thus introducing a new strategy for future detection of cancer driver elements in the expressed non-coding genome.

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Marc Descrimes, Yousra Ben Zouari, Maxime Wery, Rachel Legendre, Daniel Gautheret, Antonin Morillon (2015 Sep 9)

VING: a software for visualization of deep sequencing signals.

BMC research notes : 419 : DOI : 10.1186/s13104-015-1404-5 En savoir plus
Résumé

Next generation sequencing (NGS) data treatment often requires mapping sequenced reads onto a reference genome for further analysis. Mapped data are commonly visualized using genome browsers. However, such software are not suited for a publication-ready and versatile representation of NGS data coverage, especially when multiple experiments are simultaneously treated.

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Flore Sinturel, Albertas Navickas, Maxime Wery, Marc Descrimes, Antonin Morillon, Claire Torchet, Lionel Benard (2015 Sep 8)

Cytoplasmic Control of Sense-Antisense mRNA Pairs.

Cell reports : 1853-64 : DOI : 10.1016/j.celrep.2015.08.016 En savoir plus
Résumé

Transcriptome analyses have revealed that convergent gene transcription can produce many 3′-overlapping mRNAs in diverse organisms. Few studies have examined the fate of 3′-complementary mRNAs in double-stranded RNA-dependent nuclear phenomena, and nothing is known about the cytoplasmic destiny of 3′-overlapping messengers or their impact on gene expression. Here, we demonstrate that the complementary tails of 3′-overlapping mRNAs can interact in the cytoplasm and promote post-transcriptional regulatory events including no-go decay (NGD) in Saccharomyces cerevisiae. Genome-wide experiments confirm that these messenger-interacting mRNAs (mimRNAs) form RNA duplexes in wild-type cells and thus have potential roles in modulating the mRNA levels of their convergent gene pattern under different growth conditions. We show that the post-transcriptional fate of hundreds of mimRNAs is controlled by Xrn1, revealing the extent to which this conserved 5′-3′ cytoplasmic exoribonuclease plays an unexpected but key role in the post-transcriptional control of convergent gene expression.

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Nicola Manfrini, Michela Clerici, Maxime Wery, Chiara Vittoria Colombo, Marc Descrimes, Antonin Morillon, Fabrizio d'Adda di Fagagna, Maria Pia Longhese (2015 Aug 1)

Resection is responsible for loss of transcription around a double-strand break in Saccharomyces cerevisiae.

eLife : DOI : 10.7554/eLife.08942 En savoir plus
Résumé

Emerging evidence indicate that the mammalian checkpoint kinase ATM induces transcriptional silencing in cis to DNA double-strand breaks (DSBs) through a poorly understood mechanism. Here we show that in Saccharomyces cerevisiae a single DSB causes transcriptional inhibition of proximal genes independently of Tel1/ATM and Mec1/ATR. Since the DSB ends undergo nucleolytic degradation (resection) of their 5′-ending strands, we investigated the contribution of resection in this DSB-induced transcriptional inhibition. We discovered that resection-defective mutants fail to stop transcription around a DSB, and the extent of this failure correlates with the severity of the resection defect. Furthermore, Rad9 and generation of γH2A reduce this DSB-induced transcriptional inhibition by counteracting DSB resection. Therefore, the conversion of the DSB ends from double-stranded to single-stranded DNA, which is necessary to initiate DSB repair by homologous recombination, is responsible for loss of transcription around a DSB in S. cerevisiae.

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Hung-Fu Liao, Chu-Fan Mo, Shinn-Chih Wu, Dai-Han Cheng, Chih-Yun Yu, Kai-Wei Chang, Tzu-Hao Kao, Chia-Wei Lu, Marina Pinskaya, Antonin Morillon, Shih-Shun Lin, Winston T K Cheng, Déborah Bourc'his, Timothy Bestor, Li-Ying Sung, Shau-Ping Lin (2015 Jul 11)

Dnmt3l-knockout donor cells improve somatic cell nuclear transfer reprogramming efficiency.

Reproduction (Cambridge, England) : 245-56 : DOI : 10.1530/REP-15-0031 En savoir plus
Résumé

Nuclear transfer (NT) is a technique used to investigate the development and reprogramming potential of a single cell. DNA methyltransferase-3-like, which has been characterized as a repressive transcriptional regulator, is expressed in naturally fertilized egg and morula/blastocyst at pre-implantation stages. In this study, we demonstrate that the use of Dnmt3l-knockout (Dnmt3l-KO) donor cells in combination with Trichostatin A treatment improved the developmental efficiency and quality of the cloned embryos. Compared with the WT group, Dnmt3l-KO donor cell-derived cloned embryos exhibited increased cell numbers as well as restricted OCT4 expression in the inner cell mass (ICM) and silencing of transposable elements at the blastocyst stage. In addition, our results indicate that zygotic Dnmt3l is dispensable for cloned embryo development at pre-implantation stages. In Dnmt3l-KO mouse embryonic fibroblasts, we observed reduced nuclear localization of HDAC1, increased levels of the active histone mark H3K27ac and decreased accumulation of the repressive histone marks H3K27me3 and H3K9me3, suggesting that Dnmt3l-KO donor cells may offer a more permissive epigenetic state that is beneficial for NT reprogramming.

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Marta Kwapisz, Myriam Ruault, Erwin van Dijk, Stephanie Gourvennec, Marc Descrimes, Angela Taddei and Antonin Morillon (2015 Jul 3)

Expression of Subtelomeric lncRNAs Links Telomeres Dynamics to RNA Decay in S. cerevisiae

Non-coding RNA : 1 : 94-126 : DOI : 10.3390/ncrna1020094 En savoir plus
Résumé

Long non-coding RNAs (lncRNAs) have been shown to regulate gene expression, chromatin domains and chromosome stability in eukaryotic cells. Recent observations have reported the existence of telomeric repeats containing long ncRNAs – TERRA in mammalian and yeast cells. However, their functions remain poorly characterized. Here, we report the existence in S. cerevisiae of several lncRNAs within Y′ subtelomeric regions. We have called them subTERRA. These belong to Cryptic Unstable Transcripts (CUTs) and Xrn1p-sensitive Unstable Transcripts (XUTs) family. subTERRA transcription, carried out mainly by RNAPII, is initiated within the subtelomeric Y’ element and occurs in both directions, towards telomeres as well as centromeres. We show that subTERRA are distinct from TERRA and are mainly degraded by the general cytoplasmic and nuclear 5′- and 3′- RNA decay pathways in a transcription-dependent manner. subTERRA accumulates preferentially during the G1/S transition and in C-terminal rap1 mutant but independently of Rap1p function in silencing. The accumulation of subTERRA in RNA decay mutants coincides with telomere misregulation: shortening of telomeres, loss of telomeric clustering in mitotic cells and changes in silencing of subtelomeric regions. Our data suggest that subtelomeric RNAs expression links telomere maintenance to RNA degradation pathways.

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