The mcm2 mutation of yeast affects replication, rather than segregation or amplification of the two micron plasmid

Abstract

We have studied the maintenance of the endogenous two micron (2μ) plasmid in a strain of yeast carrying the nuclear mutation mcm2. This mutation, earlier shown to affect the maintenance of yeast minichromosomes in an ARS-dependent manner, also affected the copy number of the 2μ plasmid. The effect was more pronounced at 35 °C leading to the elimination of the plasmid from the cells cultured at this temperature. The mutant cells could be efficiently cured of the circle by transformation with 2μ ORI-carrying hybrid vectors, an observation consistent with the low copy number of the endogenous plasmid. A chromosomal revertant of this mutant for another ARS(ARS1) was found also to confer stability on the 2μ ORI-carrying minichromosomes and had elevated levels of the endogenous plasmid. The mutation neither affected the segregation nor the amplification process mediated by site-specific recombination at FRT sites requiring the FLP gene-encoded protein action. ARS131C, an ARS that was unaffected in the mutant at 25 °C, could elevate the copy number of a 2μ hybrid vector in the mutant cells. In view of these results, some aspects of segregation and copy number control of the endogeneous plasmid have been discussed. We propose that the mutation impairs the 2μ ORI function, leading to its loss.