REPLICATION SLIPPAGE BETWEEN DISTANT SHORT REPEATS IN SACCHAROMYCES-CEREVISIAE DEPENDS ON THE DIRECTION OF REPLICATION AND THE RAD50 AND RAD52 GENES

Small direct repeats, which are frequent in all genomes, are a potenti al source of genome instability, To study the occurrence and genetic c ontrol of repeat-associated deletions, we developed a system in the ye ast Saccharomyces cerevisiae that was based on small direct repeats se parated by either random sequences or inverted repeats. Deletions were examined in the LYS2 gene, using a set of 31- to 156-bp inserts that included inserts with no apparent potential for secondary structure as well as two quasipalindromes. All inserts were flanked by 6- to 9-bp direct repeats of LYS2 sequence, providing an opportunity for Lys(+) r eversion via precise excision. Reversions could arise by extended dele tions involving either direct repeats or random sequences and by -1- o r +2-bp frameshift mutations, The deletion breakpoints were always ass ociated with short (3- to 9-bp) perfect or imperfect direct repeats, C ompared with the POL(+) strain, deletions between small direct repeats were increased as much as 100-fold, and the spectrum was changed in a temperature-sensitive DNA polymerase delta pol3-t mutant, suggesting a role for replication. The type of deletion depended on orientation r elative to the origin of replication, On the basis of these results, w e propose (i) that extended deletions between small repeats arise by r eplication slippage and (ii) that the deletions occur primarily in eit her the leading or lagging strand. The RAD50 and RAD52 genes, which ar e required for the recombinational repair of many kinds of DNA double- strand breaks, appeared to be required also for the production of up t o 90% of the deletions arising between separated repeats in the pol3-t mutant, suggesting a newly identified role for these genes in genome stability and possibly replication.