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PhD student position on the role played by chromatin dynamics in condensin-mediated 3D genome organization during mitosis

3 years PhD position funded by the ANR

Key words: 3D genome, mitotic chromosome, condensin, SMC, chromatin dynamics

Starting date: autumn 2022

Where: Team Chromatin Dynamics in Mitotic Chromosome Assembly

PhD supervisor: Pascal BERNARD (CNRS senior scientist and team leader).

Contact: pascal.bernard[at]ens-lyon.fr

 

A conserved, yet incompletely understood, principle in living organisms is the folding of the genome into loops by DNA-motors of the SMC family such as condensin and cohesin (1). In eukaryotes, large genomes are also packed into arrays of nucleosomes, which compacts DNA but also reduces its accessibility to non-histone factors. In essence, interplays between chromatin and eukaryotic SMCs are inevitable, but the nature and the consequences of such interplays remain largely unknown. The overall objective of this PhD project is to tackle that question by focusing on the condensin SMC complex.

Condensin (Fig. 1A) is best characterised as the driver of mitotic chromosome condensation, namely the folding of a chromatin fibre into a 3D mega-structure, the metaphase chromosome, shaped for accurate segregation in anaphase. When condensin is impaired, misshapen mitotic chromosomes fail to segregate in anaphase and form chromatin bridges, which triggers chromotrypsis and genetic instability. In vivo, condensin massively loads onto DNA upon mitotic entry and fold chromatin into arrays of loops (Fig. 1B), as shown by Hi-C. In vitro, condensin anchors DNA through direct electrostatic contacts with DNA and, through subsequent cycles of ATP hydrolysis, extrudes adjacent DNA into a loop of increasing size (Fig. 1C-D).  Such loop-extrusion reaction convincingly describes the structural properties of mitotic chromosomes. Yet, we still largely ignore how condensin achieves, in the crowed context of a chromatinized genome, the direct contacts with DNA that support its function. Conflicting results have been obtained thus far regarding the impact of nucleosomes on condensin (2, 3), leaving the question unanswered.

Our team studies the integrated functioning of condensin by using Fission yeast as a model system. Condensin and chromatin dynamics are highly conserved in this single-celled eukaryote. We use a combination of molecular genetics and state of art functional genomics (calibrated-ChIP-seq, Hi-C and Mnase-seq) and single molecule microscopy (SPT-PALM and SMLM) approaches to assess the impact of chromatin on condensin and mitotic chromosome assembly in vivo. We provided the first evidence that nucleosome eviction underlies condensin binding to chromatin in vivo (3). More recently, we identified nucleosome remodelling enzymes as binding partners of condensin and as candidates for coupling condensin-mediated loop extrusion to nucleosome dynamics. The available PhD project aims at precisely determining what is the role played by such remodelers in the loading of condensin, its loop formation activity, and in chromatin structure during mitosis. The overall objective being to determine whether and how chromatin must be remodelled to accommodate loop extrusion by condensin.

The project will be performed in close collaboration with the teams of Olivier Cuvier (CBI-Toulouse) for multi-omic analysis and of Daniel Jost (LBMC, ENS-Lyon) for biophysical modelling, and will consist into iterative cycles of experiments and data acquisition (P. Bernard lab); computational analysis (OC) and biophysical modelling (DJ) to reach explicit conclusions.

Technologies used during the PhD

Molecular genetics to control gene expression and protein levels ● Fluorescence microscopy ● Single molecule microscopy ● High resolution ChIP-seq, MNase-seq and Hi-C genomic approaches ● Bio-informatic analyses.

Candidate profile

The applicant must hold or be preparing a master degree in genomics, molecular genetics or a closely related field.

- Good background in chromatin structure and/or chromosome organization.

- Curiosity, autonomy and good organization and presentation skills.

- Basic knowledge of the UNIX and R programming languages for genomic data analysis is

not mandatory but would be a plus.

- Languages: French or English.

 

Bibliography

1.           I. F. Davidson, J.-M. Peters, Genome folding through loop extrusion by SMC complexes. Nat. Rev. Mol. Cell Biol. 22, 445–464 (2021).

2.           K. Tada, H. Susumu, T. Sakuno, Y. Watanabe, Condensin association with histone H2A shapes mitotic chromosomes. Nature. 474, 477–83 (2011).

3.           E. Toselli-Mollereau, X. Robellet, L. Fauque, S. Lemaire, C. Schiklenk, C. Klein, C. Hocquet, P. Legros, L. N’Guyen, L. Mouillard, E. Chautard, D. Auboeuf, C. H. Haering, P. Bernard, Nucleosome eviction in mitosis assists condensin loading and chromosome condensation. EMBO J. 35, 1565–1581 (2016).