Examination of the physical properties and genetic requirements of mating type interconversion

Peter Houston, Eugenia Xu, James Broach. Dept. of Molecular Biology, Princeton University, LTL 301, Princeton, NJ, 08540, United States of America.

Haploid wildtype yeast can change mating type as much as once every generation. The HO endonuclease initiates the process by cutting at MAT. This DNA break is then repaired by homologous recombination with one of two donor sequences, HML or HMR. The different mating types a and α are biased in the selection of donor: a chooses HMLα, and α chooses HMRa. This nonrandom cell type dependent selection ensures that cells switch their mating type instead of futile replacement of the same mating type allele. A physical examination of the pairing of MAT with HML and HMR should provide insight into the process. Preliminary results show that large scale preorganization of the chromosome does not occur. This result suggests that sampling of HML and HMR may be occurring after the DNA break and the choice of donor occurs at the commitment to recombination. Perhaps the rate limiting step that controls selection occurs during formation of a stable strand invasion intermediate or at the recruitment of a competent replication holoenzyme. The observation of pairing in single cells will allow us to dissect these events through the gratuitous use of mutants and drugs. Previous genetic analysis of donor preference has been elusive since the assays of the process were too cumbersome. We have developed a facile assay of donor selection that allows the global genetic analysis of mutants that affect the process. Results of these studies and their implications will be discussed.

Program Nr. 279C from 2004 Yeast meeting

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