Analysis of the genes involved in the maintenance of mini-chromosomes in yeast.

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Yeast plasmids containing the autonomously replicating sequence(ARS) and the centromere (CEN) can be maintained rather stably as mini- chromosomes in yeast cells. We selected the mutants in which mini- chromosomes were unstable at a higher temperature and examined those phenotypes. Our goal of this project is to define how the gene products function for either replication or partitioning.By using the system developed by Hartwell et al., we newly selected three ts mutants; two mutants formed pink colonies with white sectors (presumably defective in replication), one formed pink colonies with white and red sectors (non-disjunction ?). Now we are trying to clone the genes, by complementing activity of the ts phenotype.The GSTl gene which is essential for Gl-to-S transition, affects the stability of mini-chromosomes. The mini-chromosomes were unstable and easily lost at 34ﾟC. compared at 26ﾟC. The gene product (MW. 77K) has a homologous domain to polypeptide chain elongation factor lalpha. This gene was found to be the same as the frame-shift suppressor SUFl2, omnipotent suppressor sup2 (sup36) and allosuppressor sal3. We are in progress in studying the function of this protein.We also cloned the human homologue of the yeast GSTl gene from cDNA library of human fibroblast. The gene coded for a protein smaller than yeast GSTl but contained the complete domain homologous to EFlalpha and the consensus sequence for GTP-binding proteins. The mRNA level of the human GSTl gene was the maximum just before entering the S-phase.