The GeBi team is co-directed by Emmanuelle Fabre and Pascale Lesage, two leaders in the field of 3D genome organization and mobile DNA. Our lab is investigating the processes that govern genome organization and integrity using eukaryotic models including yeast and mouse embryonic stem cells (mESC).
Eukaryotic chromosomes are non-randomly organized in the 3D space of the nucleus and their spatial organization is involved in fundamental genome functions, such as repair of DNA damage and transcription. Further, genome integrity and gene expression are constantly challenged by transposable elements (TEs) representing respectively 3% to 50% of yeast and human genomes.
Given the very high degree of conservation of DNA-repair machineries, TE mobilization and impact on genomes and mechanisms of chromatin organization between yeast and humans, our long-term goal is to take advantage from our understanding in yeast to determine if similar mechanisms operate in human cells. In this respect, opening up to translational research with the teams at St Louis Hospital is a constant concern.
Molecular bases of TE integration site selection (ISS)
Transposable elements (TEs) are dynamic genomic units able to move within the genome and are present in all domains of life. They have been shown to be a powerful force of evolution and act positively in long-term adaptation to different environments. However, TEs can also be deleterious in the short-term. Their integration into the host genome can inactivate/deregulate gene expression or induce large chromosomal rearrangements. In humans, more than hundred heritable diseases or cancers have been assigned to de novo TE insertions. To understand how TEs contribute to genome stability, expression and evolution, we are deciphering the molecular bases of their integration site selection. To this purpose, we are combining molecular genetic tools and genome-wide approaches to characterize the multiple factors that contribute to the integration site preferences of the yeast Ty1 LTR-retrotransposon for Pol III-transcribed genes.
The different factors that can influence integration site selection, from nuclear entry to chromosomal DNA. An interaction between the retroelement integrase and DNA or chromatin associated proteins is determinant to tether the complex of integration at the site of integration. After Sultana, Zamborlini, Cristofari & Lesage, Nature Reviews in Genetics, 2017.
Understanding the mechanisms that control chromosome mobility and repair upon DNA damage in the context of 3D chromosome organization
Genomes are constantly under endogenous or exogenous genotoxic threats. Double strand breaks of chromatin are the most deleterious damages. Left unrepaired, cell will die; erroneously repaired, cell might engage to cancer. Cells have therefore evolved sophisticated and conserved mechanisms to ensure genome integrity. Among those, chromosome mobility, at the site of damage and elsewhere, in undamaged chromosomal domains, are conserved features of threaten genomes. On the other hand, genomes are not randomly organized and chromosome organization is functionally important for faithful repair to occur. Given the importance for genome integrity and cell maintenance, we aim at understanding how 3D chromosomes are organized and what controls chromosome mobility and repair.
A scheme of yeast chromosomes (grey lines) organization as defined with the help of gene localization by in vivo high-through put microscopy, chromosome capture configuration (HiC) and polymer modeling. Genome is mobile (concentric circles). We interrogate the interplay between chromosome organization and genome dynamics with DNA damage (Double Strand Breaks). After Fabre & Zimmer, Nucleus, 2018.
Our group has developed tools allowing for induction of double strand breaks (DSBs) at desired positions in the genome. By combining yeast genetics and genomics, cell biology in living cells together with live-video microscopy and image analyses tools, we study chromosome mobility and architecture upon DNA damage. We are interested in developing interdisciplinary aspects of our research and we regularly collaborate with physicists to feed chromosome models based on polymer physics and super resolution imaging (PALM microscopy).
The recent discoveries of our team are focused on the essential role of histone H2A phosphorylation in chromosome mobility in the context of 3D chromosome organization. The current objectives of the group are to understand how H2A phosphorylation sensitizes cells for repair, to define the contributions of other epigenetic marks and to comprehend the role of chromosome architecture in this process.
Heterochromatin organization and genome integrity in pluripotent cells
The establishment of heterochromatin is essential to repress transcription of DNA repeats and contributes to maintain genome integrity. During early embryonic development, the active DNA demethylation occurring before embryo implantation challenges heterochromatin formation. In this context, alternative mechanisms are established to form the heterochromatin compartment.
Conventional organization of a mammalian nucleus. Different landmarks contribute to genome segregation into functional compartments. The nuclear envelope defines the edge of the nucleus. The nuclear lamina is a meshwork of proteins that interact with heterochromatic genomic regions called lamina-associated domains (LADs), only interrupted by nuclear pore complexes (NPC). The two main other landmarks: the chromocenter, composed of clustered pericentromeric heterochromatin (PCH), and the nucleolus. (right) region of the nucleus with the histone marks associated with the different compartments. LADs at the periphery are enriched in H3K9me2/3 modifications and H3K27me3 at LAD borders. Heterochromatin regions show different levels of HP1 with higher concentration at PCH. The NPC interacts with euchromatin domains. Ac, acetylation; me2/3, di or tri-methylation; DNA 5mC, DNA methylation; HP1, Heterochromatin Protein 1; LBR, Lamin B Receptor; NET, Nuclear Envelope Transmembrane protein. After Canat , Veillet , Bonnet &Therizols. Brief Funct Genomics, 2020.
Using mouse embryonic stem cells (mESC) as a model, our team focuses on the role of the histone variant H3.3 and its chaperone Daxx in the formation of heterochromatin at tandem and interspaced repeats.
Molecular bases of TE integration site selection (ISS)
The Ty1 LTR-retrotransposon of S. cerevisiae preferentially integrates into a 1-kb window upstream of RNA polymerase III (Pol III)-transcribed genes. Because tRNA genes are in multicopy and thus individually are non-essential, this integration pattern allows Ty1 to replicate and yet minimizes genetic damage to its host.
► We have identified an interaction between Ty1 integrase (IN) and the Pol I and Pol III common subunit AC40.
► We have demonstrated that this interaction is a predominant determinant of Ty1 integration upstream of Pol III-transcribed genes. Loss of IN1/AC40 interaction leads to a redistribution of Ty1 insertions, mainly to subtelomeres (Bridier-Nahmias, Tchalikian-Cosson et al, Science 2015<).
► We have also identified the targeting domain of Ty1 that interacts with AC40 in the bipartite nuclear localization signal of IN1 located at its C-terminus (Asif-Laidin et al. EMBO J 2020).
► Using genome-wide approaches, we have shown that this sequence is required for the recruitment of IN1 to Pol III and Pol I-transcribed genes and the integration of Ty1 at Pol III-transcribed genes.
► Replacement of the targeting domain of Ty5 retrotransposon by that of Ty1 induces Ty5 integration into Pol III-transcribed genes.
Determinants of chromosome organization in budding yeast
► Genes occupy a defined territory of limited volume (in collaboration with O.Gadal (LBME, Toulouse, France) and C. Zimmer (Institut Pasteur, Paris, France); (Berger et al., 2008).
► Subtelomeres position close to nuclear periphery depends on chromosome arm length, nucleolar volume and centromere attachment to the spindle pole body (yeast microtubule organizing center) and subtelomeres of similar sized chromosome arms have a higher probability to come into contact; (Therizols et al., 2010).
► Strong gene activation causes their relocation in specific and distinct areas of the nuclear periphery (in collaboration with J. Brickner, Northwestern University, USA); (Brickner et al., 2012).
► Polymer physics recapitulates coarse grain organization of yeast chromosomes arms (in collaboration with C. Zimmer (Institut Pasteur, Paris, France); (Wong et al., 2012) (Arbona et al., 2017).
► Actin is important for subtelomere positionning and dynamics (in collaboration with R. Koszul, Institut Pasteur, Paris, France) (Spichal et al., 2016).
Functional role of chromosomes dynamic organization in DNA repair and stress
To understand the interplay between chromosome dynamic organization and DSB repair, we have shown :
► Subtelomeric transcriptional repression and DSB repair depend on the integrity of the nuclear pore complexes (NPC); (Therizols et al., JCB, 2006).
► Subtelomere territories are sufficient to predict DSB repair efficiency by homologous recombination (in collaboration with M. Kupiec, Tel Aviv University, Israel); (Agmon et al., NCB, 2013).
► Chromosome organization is important for osmotic stress response memory as seen by microfluidic lineage (in collaboration with P.Hersen, Institut Curie, Paris, France). (Meriem et al., Cells, 2019).
► The constitutively phosphorylated H2A-S129E mutant shows enhanced chromatin dynamics due to chromatin structural modification, leading to efficient checkpoint activation and NHEJ-repair, even in the absence of Rad9 (Garcia Fernandez et al., J.Cell Sciences, 2021)
Determinants of chromosome organization in mES cells
► Daxx is essential for heterochromatin maintenance of ground-state embryonic stem cells, preventing the cells from DNA damage accumulation. (Canat et al., 2021, BioRXiv).
Molecular bases of TE integration site selection (ISS)
► A nuclear pore sub-complex restricts the propagation of Ty retrotransposons by limiting their transcription, Amandine Bonnet, Carole Chaput, Benoit Palancade, Pascale Lesage, bioRxiv 2021.01.05.425438; doi: https://doi.org/10.1101/2021.01.05.425438
► Light and shadow on the mechanisms of integration site selection in yeast Ty retrotransposon families. Bonnet A, Lesage P.Curr Genet. 2021 Feb 15. doi: 10.1007/s00294-021-01154-7. Online ahead of print.PMID: 33590295 Review.
► A small targeting domain in Ty1 integrase is sufficient to direct retrotransposon integration upstream of tRNA genes. Asif-Laidin A, Conesa C, Bonnet A, Grison C, Adhya I, Menouni R, Fayol H, Palmic N, Acker J, Lesage P.EMBO J. 2020 Sep 1;39(17):e104337. doi: 10.15252/embj.2019104337. Epub 2020 Jul 17.PMID: 32677087
► Barkova, A., A. Asif-Laidin, and P. Lesage, Genome-Wide Mapping of Yeast Retrotransposon Integration Target Sites. Methods Enzymol, (2018). 612: p. 197-223.
► Sultana, T., A. Zamborlini, G. Cristofari, and P. Lesage, Integration site selection by retroviruses and transposable elements in eukaryotes. Nat Rev Genet, (2017). 18(5): p. 292-308.
► Curcio, M.J., S. Lutz, and P. Lesage, The Ty1 LTR-retrotransposon of budding yeast, Saccharomyces cerevisiae. Microbiol Spectr, (2015). 3(2): p. 1-35.
► Bridier-Nahmias, A., A. Tchalikian-Cosson, J.A. Baller, R. Menouni, H. Fayol, A. Flores, A. Saib, M. Werner, D.F. Voytas, and P. Lesage, Retrotransposons. An RNA polymerase III subunit determines sites of retrotransposon integration. Science, (2015). 348(6234): p. 585-8.
► Bridier-Nahmias, A. and P. Lesage, Two large-scale analyses of Ty1 LTR-retrotransposon de novo insertion events indicate that Ty1 targets nucleosomal DNA near the H2A/H2B interface. Mob DNA, (2012). 3(1): p. 22.
► Servant, G., C. Pennetier, and P. Lesage, Remodeling yeast gene transcription by activating the Ty1 long terminal repeat retrotransposon under severe adenine deficiency. Mol Cell Biol, (2008). 28(17): p. 5543-54.
► Lesage, P. and A.L. Todeschini, Happy together: the life and times of Ty retrotransposons and their hosts. Cytogenet Genome Res, (2005). 110(1-4): p. 70-90.
Determinants of chromosome organization in budding yeast & Functional role of chromosome dynamic organization in DNA repair and stress
► Fabiola Garcia Fernandez†, Etienne Almayrac†, Ànnia Carré Simon, Renaud Batrin, Yasmine Khalil, Michel Boissac and Emmanuelle Fabre*. (2022). Global chromatin mobility induced by a DSB is dictated by chromosomal conformation and defines the outcome of Homologous Recombination. eLife. https://doi.org/10.7554/eLife.78015
► Fabiola Garcia Fernandez, Brenda Lemos, Yasmine Khalil, Renaud Batrin, James E. Haber and Fabre, E*. (2021) Modified chromosome structure caused by phosphomimetic H2A modulates the DNA damage response by increasing chromatin mobility in yeast, Journal of Cell Science, 134(6). PMID : 33622771.
► Bruhn, C., Ajazi, A., Ferrari, E., Lan, M.C., Batrin, R., Choudhary, R., Walvekar, A., Laxman, S., Longhese, M. P., Fabre, E., Bustamente , M., and Foiani., M. (2020). The Rad53CHK1/CHK2-Spt21NPAT and Tel1ATM axes couple glucose 1 tolerance to histone dosage and subtelomeric silencing. Nature Comm. 11(1): p 4154-4168. PMID: 32814778.
► Zimmer, C. and E. Fabre, Chromatin mobility upon DNA damage: state of the art and remaining questions. Curr Genet, (2019). 65(1): p. 1-9.
► Meriem, Z.B., Y. Khalil, P. Hersen*, and E. Fabre*, Hyperosmotic Stress Response Memory is Modulated by Gene Positioning in Yeast. Cells, (2019). 8(6).
► Fabre, E. and C. Zimmer, From dynamic chromatin architecture to DNA damage repair and back. Nucleus, (2018). 9(1): p. 161-170.
► Spichal, M. and E. Fabre, The emerging role of the cytoskeleton in chromosome dynamics. Frontiers in Genetics, (2017). in press: p. 681-92.
► Herbert, S., A. Brion, J.M. Arbona, M. Lelek, A. Veillet, B. Lelandais, J. Parmar, F.G. Fernandez, E. Almayrac, Y. Khalil, E. Birgy, E. Fabre*, and C. Zimmer*, Chromatin stiffening underlies enhanced locus mobility after DNA damage in budding yeast. EMBO J, (2017). 36(17): p. 2595-2608.
► Arbona, J.M., S. Herbert, E. Fabre, and C. Zimmer, Inferring the physical properties of yeast chromatin through Bayesian analysis of whole nucleus simulations. Genome Biol, (2017). 18(1): p. 81.
► Almayrac, E. and E. Fabre, Yeast nucleus: a model for chromosome folding principles. Nuclear Architecture and Dynamics hosted by Christophe Lavelle & Jean-Marc Victor in Translational Epigenetics series. Elsevier. (2017). Book chapter.
► Spichal, M., A. Brion, S. Herbert, A. Cournac, M. Marbouty, C. Zimmer, R. Koszul*, and E. Fabre*, Evidence for a dual role of actin in regulating chromosome organization and dynamics in yeast. J Cell Sci, (2016). 129(4): p. 681-92.
► Spichal, M. and E. Fabre, Subnuclear architecture of telomeres and subtelomeres in yeast. Springer-Verlag GmbH Berlin Heidelberg, (2014). Book chapter( XIV, 271 p. ): p. 13-39.
► Agmon, N., B. Liefshitz, C. Zimmer*, E. Fabre*, and M. Kupiec*, Effect of nuclear architecture on the efficiency of double-strand break repair. Nat Cell Biol, (2013). 15(6): p. 694-9.
► Wong, H., H. Marie-Nelly, S. Herbert, P. Carrivain, H. Blanc, R. Koszul, E. Fabre, and C. Zimmer, A predictive computational model of the dynamic 3D interphase yeast nucleus. Curr Biol, (2012). 22(20): p. 1881-90.
► Brickner, D.G., S. Ahmed, L. Meldi, A. Thompson, W. Light, M. Young, T.L. Hickman, F. Chu, E. Fabre, and J.H. Brickner, Transcription factor binding to a DNA zip code controls interchromosomal clustering at the nuclear periphery. Dev Cell, (2012). 22(6): p. 1234-46.
► Zimmer, C. and E. Fabre, Principles of chromosomal organization: lessons from yeast. J Cell Biol, (2011). 192(5): p. 723-33.
► Therizols, P., T. Duong, B. Dujon, C. Zimmer, and E. Fabre, Chromosome arm length and nuclear constraints determine the dynamic relationship of yeast subtelomeres. Proc Natl Acad Sci U S A, (2010). 107(5): p. 2025-30.
Determinants of chromosome organization in mES cells
► A. Canat, A. Veillet, R. Illingworth, E. Fabre, P. Therizols. (2021), DAXX safeguards pericentromeric heterochromatin formation in embryonic stem cells. bioRXiv, 2021.04.28.441827.
► Canat, A., A. Veillet, A. Bonnet, and P. Therizols, Genome anchoring to nuclear landmarks drives functional compartmentalization of the nuclear space. Brief Funct Genomics, (2020). 19(2): p. 101-110.
► Benabdallah, N.S., I. Williamson, R.S. Illingworth, L. Kane, S. Boyle, D. Sengupta, G.R. Grimes, P. Therizols, and W.A. Bickmore, Decreased Enhancer-Promoter Proximity Accompanying Enhancer Activation. Mol Cell, (2019). 76(3): p. 473-484 e7.
► Florescu, A.M., P. Therizols, and A. Rosa, Large Scale Chromosome Folding Is Stable against Local Changes in Chromatin Structure. PLoS Comput Biol, (2016). 12(6): p. e1004987.
► Therizols, P., R.S. Illingworth, C. Courilleau, S. Boyle, A.J. Wood, and W.A. Bickmore, Chromatin decondensation is sufficient to alter nuclear organization in embryonic stem cells. Science, (2014). 346(6214): p. 1238-42.
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