GENOMIC STABILITY AND RECOMBINATIONAL INTERACTIONS

基因组中相关的重复元素之间的重组可能

Recombination between related, repeated elements within the genome may lead to alterations in adjacent sequences or translocations, both potentially damaging to normal metabolism. We are investigating which genes in Saccharomyces contribute to or impinge upon recombination between diverged DNAs in yeast (homeologous). We have examined repair following transformation of plasmids containing a gap in homologous or homeologous DNAs (15% diverged), using chromosomal homology to provide the missing information; our observations suggest that repair of the two types of DNAs may be affected by different mechanisms. We have also examined spontaneous mitotic recombination between chromosomes and diverged DNAs located on centromere containing plasmids and established that mismatch previously identified repair systems have little influence on rates of homeologous recombination. Surprisingly, two genes required for many types of recombination, RAD51 and RAD52, had no influence on homeologous recombination in this system, again suggesting a separate mechanism for recombination between diverged DNAs. Our studies have now been extended to the examination of UV-induced recombination of diverged DNAs in the plasmid-chromosome assay described above. Recombination is induced at approximately the same rate as for homologous DNAs, in contrast to results previously obtained in our laboratory for ionizing radiation. In E. coli, at least some types of recombination events between diverged DNAs are inhibited or prohibited by the mismatch repair system. We are testing the hypothesis that this inhibition is largely executed during initial formation of heteroduplexes, and that it might be bypassed by pre-forming heteroduplexes. We previously obtained data suggesting that, in E. coli, the survival of a mixture of two forms of pre-formed, highly heteroduplex molecules depended on the polarity of DNA ends in each form. We have developed a technique for purifying each of the two forms, so that the individual dependences can be examined. After modifications of the procedure to remove minor contaminants, we will be in a position to answer the question posited above.