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Abstract

Eukaryotic chromosomes undergo massive three-dimensional rearrangements over the course of the cell cycle. Prior to cell division, chromosomes are condensed into rod-shaped structures which ensures the faithful transmission of the genetic material into daughter cells. Emerging evidence from biochemical and chromosome conformation capture exper- iments suggests that condensin, an evolutionary conserved multi-subunit protein com- plex, catalyzes chromosome condensation through the extrusion of DNA loops using en- ergy derived from ATP hydrolysis. However, the underlying molecular mechanism re- mains largely elusive. The objective of my PhD thesis was to gain insights into the structural basis of DNA loop extrusion by condensin. In this thesis, I report the structure of Saccharomyces cerevisiae (Sc) condensin bound to ATP and to DNA solved by cryo-electron microscopy (cryo-EM) at resolutions of 3.5 to 3.9 Å. The structure reveals that DNA binds at two distinct condensin modules, which I named “core” and “periphery”. At both modules, DNA is entrapped into topological compartments formed by the condensin subunits. In addition, I present a low-resolution structure (~10 Å) of Sc condensin core bound to DNA, without ATP. The integration of structural information obtained during my PhD with previously available condensin structures, biochemical experiments and single-molecule data results in a detailed model for condensin-mediated DNA loop extrusion. My work provides a molecular explanation for the motor and anchor functions that are required for condensin to reel DNA into a loop while stably holding onto the DNA. The molecular rearrangements of condensin core upon DNA and ATP binding suggest a “power-stroke” motor mechanism of con- densin. In addition, I present first experiments that investigate the DNA sequence specificity of condensin binding, its association with nucleosomes, and its interaction with the general transcription factor TFIIIC. These new experimental data pave the way to understand how condensin is loaded onto chromatin.