UMR3348 – Intégrité du génome, ARN et cancer

Publications de l’unité

Année de publication : 2013

Sarah Lambert, Antony M Carr (2013 Mar 1)

Impediments to replication fork movement: stabilisation, reactivation and genome instability.

Chromosoma : 33-45 : DOI : 10.1007/s00412-013-0398-9 En savoir plus
Résumé

Maintaining genome stability is essential for the accurate transmission of genetic material. Genetic instability is associated with human genome disorders and is a near-universal hallmark of cancer cells. Genetic variation is also the driving force of evolution, and a genome must therefore display adequate plasticity to evolve while remaining sufficiently stable to prevent mutations and chromosome rearrangements leading to a fitness disadvantage. A primary source of genome instability are errors that occur during chromosome replication. More specifically, obstacles to the movement of replication forks are known to underlie many of the gross chromosomal rearrangements seen both in human cells and in model organisms. Obstacles to replication fork progression destabilize the replisome (replication protein complex) and impact on the integrity of forked DNA structures. Therefore, to ensure the successful progression of a replication fork along with its associated replisome, several distinct strategies have evolved. First, there are well-orchestrated mechanisms that promote continued movement of forks through potential obstacles. Second, dedicated replisome and fork DNA stabilization pathways prevent the dysfunction of the replisome if its progress is halted. Third, should stabilisation fail, there are mechanisms to ensure damaged forks are accurately fused with a converging fork or, when necessary, re-associated with the replication proteins to continue replication. Here, we review what is known about potential barriers to replication fork progression, how these are tolerated and their impact on genome instability.

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Andrea E Prota, Maria M Magiera, Marijn Kuijpers, Katja Bargsten, Daniel Frey, Mara Wieser, Rolf Jaussi, Casper C Hoogenraad, Richard A Kammerer, Carsten Janke, Michel O Steinmetz (2013 Feb 4)

Structural basis of tubulin tyrosination by tubulin tyrosine ligase.

The Journal of cell biology : 259-70 : DOI : 10.1083/jcb.201211017 En savoir plus
Résumé

Tubulin tyrosine ligase (TTL) catalyzes the post-translational retyrosination of detyrosinated α-tubulin. Despite the indispensable role of TTL in cell and organism development, its molecular mechanism of action is poorly understood. By solving crystal structures of TTL in complex with tubulin, we here demonstrate that TTL binds to the α and β subunits of tubulin and recognizes the curved conformation of the dimer. Biochemical and cellular assays revealed that specific tubulin dimer recognition controls the activity of the enzyme, and as a consequence, neuronal development. The TTL-tubulin structure further illustrates how the enzyme binds the functionally crucial C-terminal tail sequence of α-tubulin and how this interaction catalyzes the tyrosination reaction. It also reveals how TTL discriminates between α- and β-tubulin, and between different post-translationally modified forms of α-tubulin. Together, our data suggest that TTL has specifically evolved to recognize and modify tubulin, thus highlighting a fundamental role of the evolutionary conserved tubulin tyrosination cycle in regulating the microtubule cytoskeleton.

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Année de publication : 2012

Audrey Costes, Sarah A E Lambert (2012 Dec 27)

Homologous recombination as a replication fork escort: fork-protection and recovery.

Biomolecules : 39-71 : DOI : 10.3390/biom3010039 En savoir plus
Résumé

Homologous recombination is a universal mechanism that allows DNA repair and ensures the efficiency of DNA replication. The substrate initiating the process of homologous recombination is a single-stranded DNA that promotes a strand exchange reaction resulting in a genetic exchange that promotes genetic diversity and DNA repair. The molecular mechanisms by which homologous recombination repairs a double-strand break have been extensively studied and are now well characterized. However, the mechanisms by which homologous recombination contribute to DNA replication in eukaryotes remains poorly understood. Studies in bacteria have identified multiple roles for the machinery of homologous recombination at replication forks. Here, we review our understanding of the molecular pathways involving the homologous recombination machinery to support the robustness of DNA replication. In addition to its role in fork-recovery and in rebuilding a functional replication fork apparatus, homologous recombination may also act as a fork-protection mechanism. We discuss that some of the fork-escort functions of homologous recombination might be achieved by loading of the recombination machinery at inactivated forks without a need for a strand exchange step; as well as the consequence of such a model for the stability of eukaryotic genomes.

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Allyson Holmes, Laura Roseaulin, Catherine Schurra, Herve Waxin, Sarah Lambert, Mikel Zaratiegui, Robert A Martienssen, Benoit Arcangioli (2012 Dec 27)

Lsd1 and lsd2 control programmed replication fork pauses and imprinting in fission yeast.

Cell reports : 1513-20 : DOI : 10.1016/j.celrep.2012.10.011 En savoir plus
Résumé

In the fission yeast Schizosaccharomyces pombe, a chromosomal imprinting event controls the asymmetric pattern of mating-type switching. The orientation of DNA replication at the mating-type locus is instrumental in this process. However, the factors leading to imprinting are not fully identified and the mechanism is poorly understood. Here, we show that the replication fork pause at the mat1 locus (MPS1), essential for imprint formation, depends on the lysine-specific demethylase Lsd1. We demonstrate that either Lsd1 or Lsd2 amine oxidase activity is required for these processes, working upstream of the imprinting factors Swi1 and Swi3 (homologs of mammalian Timeless and Tipin, respectively). We also show that the Lsd1/2 complex controls the replication fork terminators, within the rDNA repeats. These findings reveal a role for the Lsd1/2 demethylases in controlling polar replication fork progression, imprint formation, and subsequent asymmetric cell divisions.

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Antonio Castro-Castro, Carsten Janke, Guillaume Montagnac, Perrine Paul-Gilloteaux, Philippe Chavrier (2012 Dec 9)

ATAT1/MEC-17 acetyltransferase and HDAC6 deacetylase control a balance of acetylation of alpha-tubulin and cortactin and regulate MT1-MMP trafficking and breast tumor cell invasion.

European journal of cell biology : 950-60 : DOI : 10.1016/j.ejcb.2012.07.001 En savoir plus
Résumé

Invasive tumor cells use proteases to degrade and migrate through the stromal environment consisting of a 3D network of extracellular matrix macromolecules. In particular, MT1-MMP, a membrane-anchored metalloproteinase, is critical during cancer cell invasion. MT1-MMP is stored in endosomal compartments and then delivered to invadopodia, the specialized plasma membrane domains of invasive cancer cells endowed with extracellular matrix-degradation capacity. In macrophages, traffic of MT1-MMP vesicles to invadopodia-related podosomes requires microtubules. We previously found that in breast tumor MDA-MB-231 cells an increase of microtubule and cortactin acetylation upon inhibition of HDAC6 correlates with a decrease of matrix degradation and invasion in three-dimensional collagen I gel. Here, we investigated the role of the recently identified α-tubulin N-acetyltransferase 1 ATAT1 in invasive MDA-MB-231 cells. We found that the dynamics and distribution of MT1-MMP-positive endosomes require regulation of acetylation levels. We observed that ATAT1 tubulin acetyltransferase binds and regulates cortactin acetylation levels. In addition, ATAT1 colocalizes with cortactin at the adherent surface of the cells and it is required for 2D migration and invasive migration of MDA-MB-231 cells in collagen matrix. All together, our data indicate that a balance of acetylation and deaceylation by ATAT1/HDAC6 enzymes with opposite activities regulates the migratory and invasive capacities of breast tumor cells.

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Etienne Dardenne, Sandra Pierredon, Keltouma Driouch, Lise Gratadou, Magali Lacroix-Triki, Micaela Polay Espinoza, Eleonora Zonta, Sophie Germann, Hussein Mortada, Jean-Philippe Villemin, Martin Dutertre, Rosette Lidereau, Stéphan Vagner, Didier Auboeuf (2012 Nov 1)

Splicing switch of an epigenetic regulator by RNA helicases promotes tumor-cell invasiveness.

Nature structural & molecular biology : 1139-46 : DOI : 10.1038/nsmb.2390 En savoir plus
Résumé

Both epigenetic and splicing regulation contribute to tumor progression, but the potential links between these two levels of gene-expression regulation in pathogenesis are not well understood. Here, we report that the mouse and human RNA helicases Ddx17 and Ddx5 contribute to tumor-cell invasiveness by regulating alternative splicing of several DNA- and chromatin-binding factors, including the macroH2A1 histone. We show that macroH2A1 splicing isoforms differentially regulate the transcription of a set of genes involved in redox metabolism. In particular, the SOD3 gene that encodes the extracellular superoxide dismutase and plays a part in cell migration is regulated in an opposite manner by macroH2A1 splicing isoforms. These findings reveal a new regulatory pathway in which splicing factors control the expression of histone variant isoforms that in turn drive a transcription program to switch tumor cells to an invasive phenotype.

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Angela Bellini, Pierre-Marie Girard, Sarah Lambert, Ludovic Tessier, Evelyne Sage, Stefania Francesconi (2012 Oct 31)

Stress activated protein kinase pathway modulates homologous recombination in fission yeast.

PloS one : e47987 : DOI : 10.1371/journal.pone.0047987 En savoir plus
Résumé

Rad52 is a key player in homologous recombination (HR), a DNA repair pathway that is dedicated to double strand breaks repair and recovery of perturbed replication forks. Here we show that fission yeast Rad52 homologue is phosphorylated when S phase cells are exposed to ROS inducers such as ultraviolet A radiation or hydrogen peroxide, but not to ultraviolet C or camptothecin. Phosphorylation does not depend on kinases Chk1, Rad3, Tel1 or Cdc2, but depends on a functional stress activated protein kinase (SAPK) pathway and can be partially prevented by anti-oxidant treatment. Indeed, cells lacking Sty1, the major fission yeast MAP kinase of the SAPK pathway, do not display Rad52 phosphorylation and have UVA induced Rad52 foci that persist longer if compared to wild type cells. In addition, spontaneous intrachromosomal HR is diminished in cells lacking Sty1 and, more precisely, gene conversion is affected. Moreover, HR induced by site-specific arrest of replication forks is twice less efficient in cells that do not express Sty1. Importantly, impairing HR by deletion of the gene encoding the recombinase Rhp51 leads to Sty1 dependent Rad52 phosphorylation. Thus, SAPK pathway impinges on early step of HR through phosphorylation of Rad52 in cells challenged by oxidative stress or lacking Rhp51 and is required to promote spontaneous gene conversion and recovery from blocked replication forks.

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Ismail Iraqui, Yasmina Chekkal, Nada Jmari, Violena Pietrobon, Karine Fréon, Audrey Costes, Sarah A E Lambert (2012 Oct 18)

Recovery of arrested replication forks by homologous recombination is error-prone.

PLoS genetics : e1002976 : DOI : 10.1371/journal.pgen.1002976 En savoir plus
Résumé

Homologous recombination is a universal mechanism that allows repair of DNA and provides support for DNA replication. Homologous recombination is therefore a major pathway that suppresses non-homology-mediated genome instability. Here, we report that recovery of impeded replication forks by homologous recombination is error-prone. Using a fork-arrest-based assay in fission yeast, we demonstrate that a single collapsed fork can cause mutations and large-scale genomic changes, including deletions and translocations. Fork-arrest-induced gross chromosomal rearrangements are mediated by inappropriate ectopic recombination events at the site of collapsed forks. Inverted repeats near the site of fork collapse stimulate large-scale genomic changes up to 1,500 times over spontaneous events. We also show that the high accuracy of DNA replication during S-phase is impaired by impediments to fork progression, since fork-arrest-induced mutation is due to erroneous DNA synthesis during recovery of replication forks. The mutations caused are small insertions/duplications between short tandem repeats (micro-homology) indicative of replication slippage. Our data establish that collapsed forks, but not stalled forks, recovered by homologous recombination are prone to replication slippage. The inaccuracy of DNA synthesis does not rely on PCNA ubiquitination or trans-lesion-synthesis DNA polymerases, and it is not counteracted by mismatch repair. We propose that deletions/insertions, mediated by micro-homology, leading to copy number variations during replication stress may arise by progression of error-prone replication forks restarted by homologous recombination.

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Adrien Decorsière, Christine Toulas, Françoise Fouque, Anne-Françoise Tilkin-Mariamé, Janick Selves, Rosine Guimbaud, Edith Chipoulet, Caroline Delmas, Jean-Marc Rey, Pascal Pujol, Gilles Favre, Stefania Millevoi, Stéphan Vagner (2012 Jul 1)

Decreased efficiency of MSH6 mRNA polyadenylation linked to a 20-base-pair duplication in Lynch syndrome families.

Cell cycle (Georgetown, Tex.) : 2578-80 : DOI : 10.4161/cc.20625 En savoir plus
Résumé

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Stefania Millevoi, Hervé Moine, Stéphan Vagner (2012 Jul 1)

G-quadruplexes in RNA biology.

Wiley interdisciplinary reviews. RNA : 495-507 : DOI : 10.1002/wrna.1113 En savoir plus
Résumé

G-quadruplexes are noncanonical structures formed by G-rich DNA and RNA sequences that fold into a four-stranded conformation. Experimental studies and computational predictions show that RNA G-quadruplexes are present in transcripts associated with telomeres, in noncoding sequences of primary transcripts and within mature transcripts. RNA G-quadruplexes at these specific locations play important roles in key cellular functions, including telomere homeostasis and gene expression. Indeed, RNA G-quadruplexes appear as important regulators of pre-mRNA processing (splicing and polyadenylation), RNA turnover, mRNA targeting and translation. The regulatory mechanisms controlled by RNA G-quadruplexes involve the binding of protein factors that modulate G-quadruplex conformation and/or serve as a bridge to recruit additional protein regulators. In this review, we summarize the current knowledge on the role of G-quadruplexes in RNA biology with particular emphasis on the molecular mechanisms underlying their specific function in RNA metabolism occurring in physiological or pathological conditions.

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Sébastien Rouzeau, Fabrice P Cordelières, Géraldine Buhagiar-Labarchède, Ilse Hurbain, Rosine Onclercq-Delic, Simon Gemble, Laura Magnaghi-Jaulin, Christian Jaulin, Mounira Amor-Guéret (2012 Apr 26)

Bloom’s syndrome and PICH helicases cooperate with topoisomerase IIα in centromere disjunction before anaphase.

PloS one : e33905 : DOI : 10.1371/journal.pone.0033905 En savoir plus
Résumé

Centromeres are specialized chromosome domains that control chromosome segregation during mitosis, but little is known about the mechanisms underlying the maintenance of their integrity. Centromeric ultrafine anaphase bridges are physiological DNA structures thought to contain unresolved DNA catenations between the centromeres separating during anaphase. BLM and PICH helicases colocalize at these ultrafine anaphase bridges and promote their resolution. As PICH is detectable at centromeres from prometaphase onwards, we hypothesized that BLM might also be located at centromeres and that the two proteins might cooperate to resolve DNA catenations before the onset of anaphase. Using immunofluorescence analyses, we demonstrated the recruitment of BLM to centromeres from G2 phase to mitosis. With a combination of fluorescence in situ hybridization, electron microscopy, RNA interference, chromosome spreads and chromatin immunoprecipitation, we showed that both BLM-deficient and PICH-deficient prometaphase cells displayed changes in centromere structure. These cells also had a higher frequency of centromeric non disjunction in the absence of cohesin, suggesting the persistence of catenations. Both proteins were required for the correct recruitment to the centromere of active topoisomerase IIα, an enzyme specialized in the catenation/decatenation process. These observations reveal the existence of a functional relationship between BLM, PICH and topoisomerase IIα in the centromere decatenation process. They indicate that the higher frequency of centromeric ultrafine anaphase bridges in BLM-deficient cells and in cells treated with topoisomerase IIα inhibitors is probably due not only to unresolved physiological ultrafine anaphase bridges, but also to newly formed ultrafine anaphase bridges. We suggest that BLM and PICH cooperate in rendering centromeric catenates accessible to topoisomerase IIα, thereby facilitating correct centromere disjunction and preventing the formation of supernumerary centromeric ultrafine anaphase bridges.

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Jean Philippe Arnault, Christine Mateus, Bernard Escudier, Gorana Tomasic, Janine Wechsler, Emilie Hollville, Jean-Charles Soria, David Malka, Alain Sarasin, Magalie Larcher, Jocelyne André, Nyam Kamsu-Kom, Lise Boussemart, Ludovic Lacroix, Alain Spatz, Alexander M Eggermont, Sabine Druillennec, Stephan Vagner, Alain Eychène, Nicolas Dumaz, Caroline Robert (2012 Jan 1)

Skin tumors induced by sorafenib; paradoxic RAS-RAF pathway activation and oncogenic mutations of HRAS, TP53, and TGFBR1.

Clinical cancer research : an official journal of the American Association for Cancer Research : 263-72 : DOI : 10.1158/1078-0432.CCR-11-1344 En savoir plus
Résumé

The emergence of skin tumors in patients treated with sorafenib or with more recent BRAF inhibitors is an intriguing and potentially serious event. We carried out a clinical, pathologic, and molecular study of skin lesions occurring in patients receiving sorafenib.

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Année de publication : 2011

Carsten Janke, Jeannette Chloë Bulinski (2011 Nov 16)

Post-translational regulation of the microtubule cytoskeleton: mechanisms and functions.

Nature reviews. Molecular cell biology : 773-86 : DOI : 10.1038/nrm3227 En savoir plus
Résumé

Half a century of biochemical and biophysical experiments has provided attractive models that may explain the diverse functions of microtubules within cells and organisms. However, the notion of functionally distinct microtubule types has not been explored with similar intensity, mostly because mechanisms for generating divergent microtubule species were not yet known. Cells generate distinct microtubule subtypes through expression of different tubulin isotypes and through post-translational modifications, such as detyrosination and further cleavage to Δ2-tubulin, acetylation, polyglutamylation and polyglycylation. The recent discovery of enzymes responsible for many tubulin post-translational modifications has enabled functional studies demonstrating that these post-translational modifications may regulate microtubule functions through an amazing range of mechanisms.

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Ana Leticia Maragno, Martine Pironin, Hélène Alcalde, Xiuli Cong, Klaus-Peter Knobeloch, Frederic Tangy, Dong-Er Zhang, Jacques Ghysdael, Christine Tran Quang (2011 Oct 12)

ISG15 modulates development of the erythroid lineage.

PloS one : e26068 : DOI : 10.1371/journal.pone.0026068 En savoir plus
Résumé

Activation of erythropoietin receptor allows erythroblasts to generate erythrocytes. In a search for genes that are up-regulated during this differentiation process, we have identified ISG15 as being induced during late erythroid differentiation. ISG15 belongs to the ubiquitin-like protein family and is covalently linked to target proteins by the enzymes of the ISGylation machinery. Using both in vivo and in vitro differentiating erythroblasts, we show that expression of ISG15 as well as the ISGylation process related enzymes Ube1L, UbcM8 and Herc6 are induced during erythroid differentiation. Loss of ISG15 in mice results in decreased number of BFU-E/CFU-E in bone marrow, concomitant with an increased number of these cells in the spleen of these animals. ISG15(-/-) bone marrow and spleen-derived erythroblasts show a less differentiated phenotype both in vivo and in vitro, and over-expression of ISG15 in erythroblasts is found to facilitate erythroid differentiation. Furthermore, we have shown that important players of erythroid development, such as STAT5, Globin, PLC γ and ERK2 are ISGylated in erythroid cells. This establishes a new role for ISG15, besides its well-characterized anti-viral functions, during erythroid differentiation.

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Luc Friboulet, Daniel Barrios-Gonzales, Frédéric Commo, Ken André Olaussen, Stephan Vagner, Julien Adam, Aïcha Goubar, Nicolas Dorvault, Vladimir Lazar, Bastien Job, Benjamin Besse, Pierre Validire, Philippe Girard, Ludovic Lacroix, Johanna Hasmats, Fabienne Dufour, Fabrice André, Jean-Charles Soria (2011 Sep 1)

Molecular Characteristics of ERCC1-Negative versus ERCC1-Positive Tumors in Resected NSCLC.

Clinical cancer research : an official journal of the American Association for Cancer Research : 5562-72 : DOI : 10.1158/1078-0432.CCR-11-0790 En savoir plus
Résumé

Excision repair cross-complementation group 1 (ERCC1) is a protein involved in repair of DNA platinum adducts and stalled DNA replication forks. We and others have previously shown the influence of ERCC1 expression upon survival rates and benefit of cisplatin-based chemotherapy in patients with resected non-small-cell lung cancer (NSCLC). However, little is known about the molecular characteristics of ERCC1-positive and ERCC1-negative tumors.

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