Gene conversions and crossing over during homologous and homeologous ectopic recombination in Saccharomyces cerevisiae

The pma1-105 mutation reduces the activity of the yeast plasma membrane H(+)-ATPase and causes cells to be both low pH and ammonium ion sensitive and resistant to the antibiotic hygromycin B. Revertants that can grow at pH 3.0 and on ammonium-containing plates frequently arise by ectopic recombination between pma1-105 and PMA2, a diverged gene that shares 85% DNA sequence identity with PMA1. The gene conversion tracts of revertants of pma1-105 were determined by DNA sequencing the hybrid PMA1::PMA2 genes. Gene conversion tracts ranged from 18-774 bp. The boundaries of these replacements were short (3-26 bp) regions of sequences that were identical between PMA1 and PMA2. These boundaries were not located at the regions of greatest shared identity between the two PMA genes. Similar results were obtained among low pH-resistant revertants of another mutation, pma1-147. One gene conversion was obtained in which the resulting PMA1::PMA2 hybrid was low pH-resistant but still hygromycin B-resistant. This partially active gene differs from a wild-type revertant only by the presence of two PMA2-encoded amino acid substitutions. Thus, some regions of PMA2 are not fully interchangeable with PMA1. We have also compared the efficiency of recombination between pma1-105 and either homeologous PMA2 sequence or homologous PMA1 donor sequences inserted at the same location. PMA2 x pma1-105 recombination occurred at a rate approximately 75-fold less than PMA1 x pma1-105 events. The difference in homology between the interacting sequences did not affect the proportion of gene conversion events associated with a cross-over, as in both cases approximately 5% of the Pma+ recombinants had undergone reciprocal translocations between the two chromosomes carrying pma1-105 and the donor PMA sequences. Reciprocal translocations were identified by a simple and generally useful nutritional test.     

The fluctuating phenotype of the lymphohematopoietic stem cell with cell cycle transit

When damaged by hydrogen peroxide, peripheral blood lymphocytes undergo cell death by apoptosis in the absence of internucleosomal DNA cleavage, while, in the same cells, other apoptosis-inducing treatments bring DNA cleavage to completion. However, the formation of internucleosomal DNA fragments is readily obtained if cells are pretreated with a divalent metal chelator, TPEN, at micromolar concentrations. Since the coadministration of equimolar zinc concentrations abrogates the formation of the ladder, a zinc-inhibitable endonucleolytic activity is accounted for the effect. Most notably, subtraction of zinc ions does not increase the percentage of cells undergoing apoptosis, but rather results in a rescue from death.