The selfish yeast plasmid utilizes the condensin complex and condensed chromatin for faithful partitioning.

Equipartitioning by chromosome association and copy number correction by DNA amplification are at the heart of the evolutionary success of the selfish yeast 2-micron plasmid. The present analysis reveals frequent plasmid presence near telomeres (TELs) and centromeres (CENs) in mitotic cells, with a preference towards the former. Inactivation of Cdc14 causes plasmid missegregation, which is correlated to the non-disjunction of TELs (and of rDNA) under this condition. Induced missegregation of chromosome XII, one of the largest yeast chromosomes which harbors the rDNA array and is highly dependent on the condensin complex for proper disjunction, increases 2-micron plasmid missegregation. This is not the case when chromosome III, one of the smallest chromosomes, is forced to missegregate. Plasmid stability decreases when the condensin subunit Brn1 is inactivated. Brn1 is recruited to the plasmid partitioning locus (STB) with the assistance of the plasmid-coded partitioning proteins Rep1 and Rep2. Furthermore, in a dihybrid assay, Brn1 interacts with Rep1-Rep2. Taken together, these findings support a role for condensin and/or condensed chromatin in 2-micron plasmid propagation. They suggest that condensed chromosome loci are among favored sites utilized by the plasmid for its chromosome-associated segregation. By homing to condensed/quiescent chromosome locales, and not over-perturbing genome homeostasis, the plasmid may minimize fitness conflicts with its host. Analogous persistence strategies may be utilized by other extrachromosomal selfish genomes, for example, episomes of mammalian viruses that hitchhike on host chromosomes for their stable maintenance.

Meiosis-Specific Functions of Kinesin Motors in Cohesin Removal and Maintenance of Chromosome Integrity in Budding Yeast.

Kinesin motors provide the molecular forces at the kinetochore-microtubule interface and along the spindle to control chromosome segregation. During meiosis with two rounds of microtubule assembly-disassembly, the roles of motor proteins remain unexplored. We observed that in contrast to mitosis, Cin8 and Kip3 together are indispensable for meiosis. While examining meiosis in cin8Δ kip3Δ cells, we detected chromosome breakage in the meiosis II cells. The double mutant exhibits a delay in cohesin removal during anaphase I. Consequently, some cells fail to undergo meiosis II and form dyads, while some, as they progress through meiosis II, cause a defect in chromosome integrity. We believe that in the latter cells, an imbalance of spindle-mediated force and the simultaneous persistence of cohesin on chromosomes cause their breakage. We provide evidence that tension generated by Cin8 and Kip3 through microtubule cross-linking is essential for signaling efficient cohesin removal and the maintenance of chromosome integrity during meiosis.

Evidence of Zip1 Promoting Sister Kinetochore Mono-orientation During Meiosis in Budding Yeast.

Halving of the genome during meiosis I is achieved as the homologous chromosomes move to the opposite spindle poles whereas the sister chromatids stay together and move to the same pole. This requires that the sister kinetochores should take a side-by-side orientation in order to connect to the microtubules emanating from the same pole. Factors that constrain sister kinetochores to adopt such orientation are therefore crucial to achieve reductional chromosome segregation in meiosis I. In budding yeast, a protein complex, known as monopolin, is involved in conjoining of the sister kinetochores and thus facilitates their binding to the microtubules from the same pole. In this study, we report Zip1, a synaptonemal complex component, as another factor that might help the sister kinetochores to take the side-by-side orientation and promote their mono-orientation on the meiosis I spindle. From our results, we propose that the localization of Zip1 at the centromere may provide an additional constraining factor that promotes monopolin to cross-link the sister kinetochores enabling them to mono-orient.

The 2 micron plasmid: a selfish genetic element with an optimized survival strategy within Saccharomyces cerevisiae.

Since its discovery in the early 70s, the 2 micron plasmid of Saccharomyces cerevisiae continues to intrigue researchers with its high protein-coding capacity and a selfish nature yet high stability, earning it the title of a 'miniaturized selfish genetic element'. It codes for four proteins (Rep1, Rep2, Raf1, and Flp) vital for its own survival and recruits several host factors (RSC2, Cohesin, Cse4, Kip1, Bik1, Bim1, and microtubules) for its faithful segregation during cell division. The plasmid maintains a high-copy number with the help of Flp-mediated recombination. The plasmids organize in the form of clusters that hitch-hike the host chromosomes presumably with the help of the plasmid-encoded Rep proteins and host factors such as microtubules, Kip1 motor, and microtubule-associated proteins Bik1 and Bim1. Although there is no known yeast cell phenotype associated with the 2 micron plasmid, excessive copies of the plasmid are lethal for the cells, warranting a tight control over the plasmid copy number. This control is achieved through a combination of feedback loops involving the 2 micron encoded proteins. Thus, faithful segregation and a concomitant tightly controlled plasmid copy number ensure an optimized benign parasitism of the 2 micron plasmid within budding yeast.

Microtubule-associated proteins, Bik1 and Bim1, are required for faithful partitioning of the endogenous 2 micron plasmids in budding yeast.

The 2 µ plasmid of budding yeast shows high mitotic stability similar to that of chromosomes by using its self-encoded systems, namely partitioning and amplification. The partitioning system consists of the plasmid-borne proteins Rep1, Rep2 and a cis-acting locus STB that, along with several host factors, ensures efficient segregation of the plasmid. The plasmids show high stability as they presumably co-segregate with chromosomes through utilization of various host factors. To acquire these host factors, the plasmids are thought to localize to a certain sub-nuclear locale probably assisted by the motor protein, Kip1 and microtubules. Here, we show that the microtubule-associated proteins Bik1 and Bim1 are also important host factors in this process, perhaps by acting as an adapter between the plasmid and the motor and thus helping to anchor the plasmid to microtubules. Abrogation of Kip1 recruitment at STB in the absence of Bik1 argues for its function at STB upstream of Kip1. Consistent with this, both Bik1 and Bim1 associate with plasmids without any assistance from the Rep proteins. As observed earlier with other host factors, lack of Bik1 or Bim1 also causes a cohesion defect between sister plasmids leading to plasmid missegregation.