Regulation of Chromosome Condensation by Intrinsically Disordered Regions in Condensin

Abstract

Faithful transmission of the genome during mitosis is an irreversible event that determines the survival and propagation of all living organisms. A prerequisite for this process is the large-scale restructuring of amorphous chromatin into distinct, rod-shaped chromosomes at the onset of cell division. Failure to perform effective mitotic chromosome assembly can lead to chromosomal instability, a fundamental driver of tumourigenesis. In all eukaryotic organisms, chromatin compaction is promoted by a highly conserved, pentameric complex called condensin. While fungal species contain a sole condensin complex that is responsible for chromosome assembly, higher eukaryotes including humans possess two complexes (condensins I and II) that exhibit divergent spatio-temporal regulation and make differential contributions to chromatin compaction. It has been proposed that condensin utilizes its ring-shaped architecture together with its ATPase motor domains to generate large DNA loops that contribute to higher-order chromosome organization. However, the fundamental mechanism underpinning this process and its regulation in time and space remain largely unresolved. In this thesis, I provide compelling evidence for the existence of an intrinsically disordered, autonomous DNA-recruitment module located at the amino-terminal end of the Smc4 subunit of condensin. First, I demonstrated that this region critically modulates condensin dynamics and its association with chromatin in the budding yeast Saccharomyces cerevisiae. I showed that it is capable of phase separation with DNA and that its removal from condensin causes lethality. Next, I characterized a critical DNA-binding motif at the N-terminus of this module that when deleted, leads to dramatic chromosome condensation defects and segregation errors. Finally, I explored whether an analogous DNA-recruitment module exists within human condensins I and II. I identified an intrinsically disordered region at the C-terminus of at least one regulatory subunit of each human condensin complex that has a strong affinity for DNA. Importantly, mutations in condensin have been associated with cancer and neurodevelopmental disorders in humans. I demonstrated that a pathogenic mutation in the DNA-binding module of condensin I associated with autosomal recessive microcephaly causes erroneous DNA-binding activity. Taken together, this thesis reveals a critical function for the often over-looked intrinsically disordered regions (IDRs) of the condensin complex in mitotic chromosome assembly. The notion that condensin contains at least one DNA-binding IDR is a significant proposal, as this module presumably increases the search radius of condensin for its genomic substrates and could recruit the complex to distal DNA regions that lie beyond the core structure.