| Summary: | Sister chromatid cohesion (cohesion) is a process that ensures accurate segregation of genetic material into daughter cells during mitosis in eukaryotic cells. Cohesion is mediated by a highly conserved ring-shaped protein complex called cohesin. Cohesion is thought to arise from the topological entrapment of replicated sister DNAs inside the cohesin ring. Although cohesin can load onto, and topologically entrap individual DNAs prior to DNA replication, cohesion is established during S-phase in concert with DNA replication. Analysis of small circular DNAs in yeast has revealed that cohesion establishment can occur by two pathways operating in parallel. Firstly, cohesin can load onto replicated sister DNAs de novo in a manner that is dependent on the cohesin loading protein Scc2. Secondly, there exists a ‘conversion’ pathway in which cohesin binds to unreplicated DNA during G1, and remains associated with DNA throughout S-phase during which it is converted into cohesive cohesin. This process is independent of Scc2 but requires a group of replisome-associated factors which include Ctf4. Whether this cohesin conversion pathway generates cohesion of yeast chromosomes, and the molecular mechanisms underlying conversion, remain largely unknown. It is thought that opening of cohesin’s Smc1-Smc3 hinge is crucial for entrapment of individual DNAs, but whether this is the case for entrapment of sister DNAs is currently unclear. I have developed a unique assay to measure cohesion generated solely by cohesin rings associated with DNA from G1. Using this assay, we show for the first time that conversion of non-cohesive cohesin to cohesive cohesin occurs on S. cerevisiae chromosomes during DNA replication, and that this process does not require opening of the Smc1-Smc3 hinge. Furthermore, the conversion observed was significantly reduced by deletion of Ctf4 but not significantly impacted by inactivation of the cohesin loader Scc2. These data provide crucial mechanistic insight into the process of cohesin conversion, and unequivocally demonstrate that while hinge opening is required for loading of cohesin rings onto DNA, it is not required for conversion of these rings into cohesive forms.
|
|---|