Dynamique de la Chromatine

Publications de l’équipe

Année de publication : 2015

Carlos Rivera, Francisco Saavedra, Francisca Alvarez, César Díaz-Celis, Valentina Ugalde, Jianhua Li, Ignasi Forné, Zachary A Gurard-Levin, Geneviève Almouzni, Axel Imhof, Alejandra Loyola (2015 Sep 7)

Methylation of histone H3 lysine 9 occurs during translation.

Nucleic acids research : 9097-106 : DOI : 10.1093/nar/gkv929 En savoir plus
Résumé

Histone post-translational modifications are key contributors to chromatin structure and function, and participate in the maintenance of genome stability. Understanding the establishment and maintenance of these marks, along with their misregulation in pathologies is thus a major focus in the field. While we have learned a great deal about the enzymes regulating histone modifications on nucleosomal histones, much less is known about the mechanisms establishing modifications on soluble newly synthesized histones. This includes methylation of lysine 9 on histone H3 (H3K9), a mark that primes the formation of heterochromatin, a critical chromatin landmark for genome stability. Here, we report that H3K9 mono- and dimethylation is imposed during translation by the methyltransferase SetDB1. We discuss the importance of these results in the context of heterochromatin establishment and maintenance and new therapeutic opportunities in pathologies where heterochromatin is perturbed.

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Camille Clément, Geneviève Almouzni (2015 Aug 6)

MCM2 binding to histones H3-H4 and ASF1 supports a tetramer-to-dimer model for histone inheritance at the replication fork.

Nature structural & molecular biology : 587-9 : DOI : 10.1038/nsmb.3067 En savoir plus
Résumé

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Guillermo A Orsi, Sivakanthan Kasinathan, Gabriel E Zentner, Steven Henikoff, Kami Ahmad (2015 Apr 2)

Mapping regulatory factors by immunoprecipitation from native chromatin.

Current protocols in molecular biology / edited by Frederick M. Ausubel ... [et al.] : 21.31.1-25 : DOI : 10.1002/0471142727.mb2131s110 En savoir plus
Résumé

Occupied Regions of Genomes from Affinity-purified Naturally Isolated Chromatin (ORGANIC) is a high-resolution method that can be used to quantitatively map protein-DNA interactions with high specificity and sensitivity. This method uses micrococcal nuclease (MNase) digestion of chromatin and low-salt solubilization to preserve protein-DNA complexes, followed by immunoprecipitation and paired-end sequencing for genome-wide mapping of binding sites. In this unit, we describe methods for isolation of nuclei and MNase digestion of unfixed chromatin, immunoprecipitation of protein-DNA complexes, and high-throughput sequencing to map sites of bound factors.

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Hiroaki Tachiwana, Sebastian Müller, Julia Blümer, Kerstin Klare, Andrea Musacchio, Geneviève Almouzni (2015 Apr 1)

HJURP involvement in de novo CenH3(CENP-A) and CENP-C recruitment.

Cell reports : 22-32 : DOI : 10.1016/j.celrep.2015.03.013 En savoir plus
Résumé

Although our understanding of centromere maintenance, marked by the histone H3 variant CenH3(CENP-A) in most eukaryotes, has progressed, the mechanism underlying the de novo formation of centromeres remains unclear. We used a synthetic system to dissect how CenH3(CENP-A) contributes to the accumulation of CENP-C and CENP-T, two key components that are necessary for the formation of functional kinetochores. We find that de novo CENP-T accumulation depends on CENP-C and that recruitment of these factors requires two domains in CenH3(CENP-A): the HJURP-binding region (CATD) and the CENP-C-binding region (CAC). Notably, HJURP interacts directly with CENP-C and is critical for de novo accumulation of CENP-C at synthetic centromeres. On the basis of our findings, we propose that HJURP serves a dual chaperone function in coordinating CenH3(CENP-A) and CENP-C recruitment.

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Sihem Cheloufi, Ulrich Elling, Barbara Hopfgartner, Youngsook L Jung, Jernej Murn, Maria Ninova, Maria Hubmann, Aimee I Badeaux, Cheen Euong Ang, Danielle Tenen, Daniel J Wesche, Nadezhda Abazova, Max Hogue, Nilgun Tasdemir, Justin Brumbaugh, Philipp Rathert, Julian Jude, Francesco Ferrari, Andres Blanco, Michaela Fellner, Daniel Wenzel, Marietta Zinner, Simon E Vidal, Oliver Bell, Matthias Stadtfeld, Howard Y Chang, Genevieve Almouzni, Scott W Lowe, John Rinn, Marius Wernig, Alexei Aravin, Yang Shi, Peter J Park, Josef M Penninger, Johannes Zuber, Konrad Hochedlinger (2015 Feb 23)

The histone chaperone CAF-1 safeguards somatic cell identity.

Nature : 218-24 : DOI : 10.1038/nature15749 En savoir plus
Résumé

Cellular differentiation involves profound remodelling of chromatic landscapes, yet the mechanisms by which somatic cell identity is subsequently maintained remain incompletely understood. To further elucidate regulatory pathways that safeguard the somatic state, we performed two comprehensive RNA interference (RNAi) screens targeting chromatin factors during transcription-factor-mediated reprogramming of mouse fibroblasts to induced pluripotent stem cells (iPS cells). Subunits of the chromatin assembly factor-1 (CAF-1) complex, including Chaf1a and Chaf1b, emerged as the most prominent hits from both screens, followed by modulators of lysine sumoylation and heterochromatin maintenance. Optimal modulation of both CAF-1 and transcription factor levels increased reprogramming efficiency by several orders of magnitude and facilitated iPS cell formation in as little as 4 days. Mechanistically, CAF-1 suppression led to a more accessible chromatin structure at enhancer elements early during reprogramming. These changes were accompanied by a decrease in somatic heterochromatin domains, increased binding of Sox2 to pluripotency-specific targets and activation of associated genes. Notably, suppression of CAF-1 also enhanced the direct conversion of B cells into macrophages and fibroblasts into neurons. Together, our findings reveal the histone chaperone CAF-1 to be a novel regulator of somatic cell identity during transcription-factor-induced cell-fate transitions and provide a potential strategy to modulate cellular plasticity in a regenerative setting.

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Casey E Romanoski, Christopher K Glass, Hendrik G Stunnenberg, Laurence Wilson, Genevieve Almouzni (2015 Feb 20)

Epigenomics: Roadmap for regulation.

Nature : 314-6 : DOI : 10.1038/518314a En savoir plus
Résumé

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Nicolas Richet, Danni Liu, Pierre Legrand, Christophe Velours, Armelle Corpet, Albane Gaubert, May Bakail, Gwenaelle Moal-Raisin, Raphael Guerois, Christel Compper, Arthur Besle, Berengère Guichard, Genevieve Almouzni, Françoise Ochsenbein (2015 Jan 23)

Structural insight into how the human helicase subunit MCM2 may act as a histone chaperone together with ASF1 at the replication fork.

Nucleic acids research : 1905-17 : DOI : 10.1093/nar/gkv021 En savoir plus
Résumé

MCM2 is a subunit of the replicative helicase machinery shown to interact with histones H3 and H4 during the replication process through its N-terminal domain. During replication, this interaction has been proposed to assist disassembly and assembly of nucleosomes on DNA. However, how this interaction participates in crosstalk with histone chaperones at the replication fork remains to be elucidated. Here, we solved the crystal structure of the ternary complex between the histone-binding domain of Mcm2 and the histones H3-H4 at 2.9 Å resolution. Histones H3 and H4 assemble as a tetramer in the crystal structure, but MCM2 interacts only with a single molecule of H3-H4. The latter interaction exploits binding surfaces that contact either DNA or H2B when H3-H4 dimers are incorporated in the nucleosome core particle. Upon binding of the ternary complex with the histone chaperone ASF1, the histone tetramer dissociates and both MCM2 and ASF1 interact simultaneously with the histones forming a 1:1:1:1 heteromeric complex. Thermodynamic analysis of the quaternary complex together with structural modeling support that ASF1 and MCM2 could form a chaperoning module for histones H3 and H4 protecting them from promiscuous interactions. This suggests an additional function for MCM2 outside its helicase function as a proper histone chaperone connected to the replication pathway.

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Année de publication : 2014

Yasuhiro Arimura, Kazuyoshi Shirayama, Naoki Horikoshi, Risa Fujita, Hiroyuki Taguchi, Wataru Kagawa, Tatsuo Fukagawa, Geneviève Almouzni, Hitoshi Kurumizaka (2014 Oct 30)

Crystal structure and stable property of the cancer-associated heterotypic nucleosome containing CENP-A and H3.3.

Scientific reports : 7115 : DOI : 10.1038/srep07115 En savoir plus
Résumé

The centromere-specific histone H3 variant, CENP-A, is overexpressed in particular aggressive cancer cells, where it can be mislocalized ectopically in the form of heterotypic nucleosomes containing H3.3. In the present study, we report the crystal structure of the heterotypic CENP-A/H3.3 particle and reveal its « hybrid structure », in which the physical characteristics of CENP-A and H3.3 are conserved independently within the same particle. The CENP-A/H3.3 nucleosome forms an unexpectedly stable structure as compared to the CENP-A nucleosome, and allows the binding of the essential centromeric protein, CENP-C, which is ectopically mislocalized in the chromosomes of CENP-A overexpressing cells.

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Dan Filipescu, Sebastian Müller, Geneviève Almouzni (2014 Oct 8)

Histone H3 variants and their chaperones during development and disease: contributing to epigenetic control.

Annual review of cell and developmental biology : 615-46 : DOI : 10.1146/annurev-cellbio-100913-013311 En savoir plus
Résumé

Within the nucleus, the interplay between lineage-specific transcription factors and chromatin dynamics defines cellular identity. Control of this interplay is necessary to properly balance stability and plasticity during the development and entire life span of multicellular organisms. Here, we present our current knowledge of the contribution of histone H3 variants to chromatin dynamics during development. We review the network of histone chaperones that governs their deposition timing and sites of incorporation and highlight how their distinct distribution impacts genome organization and function. We integrate the importance of H3 variants in the context of nuclear reprogramming and cell differentiation, and, using the centromere as a paradigm, we describe a case in which the identity of a given genomic locus is propagated across different cell types. Finally, we compare development to changes in stress and disease. Both physiological and pathological settings underline the importance of H3 dynamics for genome and chromatin integrity.

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Zachary A Gurard-Levin, Geneviève Almouzni (2014 Sep 4)

Histone modifications and a choice of variant: a language that helps the genome express itself.

F1000prime reports : 76 : DOI : 10.12703/P6-76 En savoir plus
Résumé

Covalent post-translational modifications on histones impact chromatin structure and function. Their misfunction, along with perturbations or mutations in genes that regulate their dynamic status, has been observed in several diseases. Thus, targeting histone modifications represents attractive opportunities for therapeutic intervention and biomarker discovery. The best approach to address this challenge is to paint a comprehensive picture integrating the growing number of modifications on individual residues and their combinatorial association, the corresponding modifying enzymes, and effector proteins that bind modifications. Furthermore, how they are imposed in a distinct manner during the cell cycle and on specific histone variants are important dimensions to consider. Firstly, this report highlights innovative technologies used to characterize histone modifications, and the corresponding enzymes and effector proteins. Secondly, we examine the recent progress made in understanding the dynamics and maintenance of histone modifications on distinct variants. We also discuss their roles as potential carriers of epigenetic information. Finally, we provide examples of initiatives to exploit histone modifications in cancer management, with the potential for new therapeutic opportunities.

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Kelly Romeo, Yann Louault, Sylvain Cantaloube, Isabelle Loiodice, Geneviève Almouzni, Jean-Pierre Quivy (2014 Jul 31)

The SENP7 SUMO-Protease Presents a Module of Two HP1 Interaction Motifs that Locks HP1 Protein at Pericentric Heterochromatin.

Cell reports : DOI : S2211-1247(15)00005-4 En savoir plus
Résumé

HP1 enrichment at pericentric heterochromatin is essential for proper chromosome segregation. While H3K9me3 is thought to be a major contributor to HP1 enrichment at pericentric domains, in mouse cells, the SUMO-protease SENP7 is required in addition to H3K9me3. How this is achieved remains elusive. Here, we find that loss of SENP7 leads to an increased time spent in mitosis. Furthermore, we reveal that a short module comprising two consecutive HP1 interaction motifs on SENP7 is the determinant for HP1 enrichment and acts by restricting HP1 mobility at pericentric domains. We propose a mechanism for maintenance of HP1 enrichment in which this module functions on top of H3K9me3 to lock contiguous HP1 molecules already docked on H3K9me3-modified nucleosomes. H3K9me3 would thus promote HP1 enrichment only if a locking system is in place. This mechanism may apply to other nuclear domains to contribute to the control of genome plasticity and integrity.

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Roderick J O'Sullivan, Genevieve Almouzni (2014 Jul 2)

Assembly of telomeric chromatin to create ALTernative endings.

Trends in cell biology : 675-85 : DOI : 10.1016/j.tcb.2014.07.007 En savoir plus
Résumé

Circumvention of the telomere length-dependent mechanisms that control the upper boundaries of cellular proliferation is necessary for the unlimited growth of cancer. Most cancer cells achieve cellular immortality by up-regulating the expression of telomerase to extend and maintain their telomere length. However, a small but significant number of cancers do so via the exchange of telomeric DNA between chromosomes in a pathway termed alternative lengthening of telomeres, or ALT. Although it remains to be clarified why a cell chooses the ALT pathway and how ALT is initiated, recently identified mutations in factors that shape the chromatin and epigenetic landscape of ALT telomeres are shedding light on these mechanisms. In this review, we examine these recent findings and integrate them into the current models of the ALT mechanism.

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Zachary A Gurard-Levin, Jean-Pierre Quivy, Geneviève Almouzni (2014 Jun 7)

Histone chaperones: assisting histone traffic and nucleosome dynamics.

Annual review of biochemistry : 487-517 : DOI : 10.1146/annurev-biochem-060713-035536 En savoir plus
Résumé

The functional organization of eukaryotic DNA into chromatin uses histones as components of its building block, the nucleosome. Histone chaperones, which are proteins that escort histones throughout their cellular life, are key actors in all facets of histone metabolism; they regulate the supply and dynamics of histones at chromatin for its assembly and disassembly. Histone chaperones can also participate in the distribution of histone variants, thereby defining distinct chromatin landscapes of importance for genome function, stability, and cell identity. Here, we discuss our current knowledge of the known histone chaperones and their histone partners, focusing on histone H3 and its variants. We then place them into an escort network that distributes these histones in various deposition pathways. Through their distinct interfaces, we show how they affect dynamics during DNA replication, DNA damage, and transcription, and how they maintain genome integrity. Finally, we discuss the importance of histone chaperones during development and describe how misregulation of the histone flow can link to disease.

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Ekaterina Boyarchuk, Dan Filipescu, Isabelle Vassias, Sylvain Cantaloube, Geneviève Almouzni (2014 Jun 6)

The histone variant composition of centromeres is controlled by the pericentric heterochromatin state during the cell cycle.

Journal of cell science : 3347-59 : DOI : 10.1242/jcs.148189 En savoir plus
Résumé

Correct chromosome segregation requires a unique chromatin environment at centromeres and in their vicinity. Here, we address how the deposition of canonical H2A and H2A.Z histone variants is controlled at pericentric heterochromatin (PHC). Whereas in euchromatin newly synthesized H2A and H2A.Z are deposited throughout the cell cycle, we reveal two discrete waves of deposition at PHC – during mid to late S phase in a replication-dependent manner for H2A and during G1 phase for H2A.Z. This G1 cell cycle restriction is lost when heterochromatin features are altered, leading to the accumulation of H2A.Z at the domain. Interestingly, compromising PHC integrity also impacts upon neighboring centric chromatin, increasing the amount of centromeric CENP-A without changing the timing of its deposition. We conclude that the higher-order chromatin structure at the pericentric domain influences dynamics at the nucleosomal level within centromeric chromatin. The two different modes of rearrangement of the PHC during the cell cycle provide distinct opportunities to replenish one or the other H2A variant, highlighting PHC integrity as a potential signal to regulate the deposition timing and stoichiometry of histone variants at the centromere.

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Catherine Dehainault, Alexandra Garancher, Laurent Castéra, Nathalie Cassoux, Isabelle Aerts, François Doz, Laurence Desjardins, Livia Lumbroso, Rocío Montes de Oca, Geneviève Almouzni, Dominique Stoppa-Lyonnet, Celio Pouponnot, Marion Gauthier-Villars, Claude Houdayer (2014 May 23)

The survival gene MED4 explains low penetrance retinoblastoma in patients with large RB1 deletion.

Human molecular genetics : 5243-50 : DOI : 10.1093/hmg/ddu245 En savoir plus
Résumé

Retinoblastoma is a non-hereditary as well as an inherited pediatric tumor of the developing retina resulting from the inactivation of both copies of the RB1 tumor suppressor gene. Familial retinoblastoma is a highly penetrant genetic disease that usually develops by carrying germline mutations that inactivate one allele of the RB1 gene, leading to multiple retinoblastomas. However, large and complete germline RB1 deletions are associated with low or no tumor risk for reasons that remain unknown. In this study, we define a minimal genomic region associated with this low penetrance. This region encompasses few genes including MED4 a subunit of the mediator complex. We further show that retinoblastoma RB1 -/- cells cannot survive in the absence of MED4, both in vitro and in orthotopic xenograft models in vivo, therefore identifying MED4 as a survival gene in retinoblastoma. We propose that the contiguous loss of the adjacent retinoblastoma gene, MED4, explains the low penetrance in patients with large deletions that include both RB1 and MED4. Our findings also point to another synthetic lethal target in tumors with inactivated RB1 and highlight the importance of collateral damage in carcinogenesis.

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