The stability system of the yeast 2 micron plasmid: analysis of plasmid and host encoded components
The work presented in this thesis aims to understand the molecular strategies used by an extrachromosomal selfish DNA element for its stable, high copy persistence. The model system studied here is the 2 micron plasmid found nearly ubiquitously in Saccharomyces yeast. By a combination of mutational and functional analysis of a plasmid coded protein Rep1p, an essential component of the stability system, we have provided support for the DNA-protein and proteinprotein interactions predicted to be important in plasmid maintenance. Using cell biological and molecular genetic methods, we have unveiled an apparent coupling of the pathways for plasmid and chromosome segregation. Mutations that affect equal partitioning of the chromosomes also affect the plasmid, and the two tend to missegregate in tandem. We have identified host factors that interact with components of the plasmid stability system, and may thus play a potential role in plasmid partitioning. In particular, we have found that the yeast cohesin complex, that bridges sister chromatids until they are ready to be unpaired and distributed to the daughter cells arising from a division event, may serve an analogous function in plasmid segregation. Our preliminary results suggest that the plasmid stability system follows the ‘recruitment model’, in which a functional complex is assembled by the sum of different sets of DNA-protein and protein-protein interactions. It is possible to reconstitute an active partitioning complex through an alternative set of interactions.