GENE-CONVERSION TRACT DIRECTIONALITY IS INFLUENCED BY THE CHROMOSOME ENVIRONMENT

Spontaneous and double-strand break (DSB)induced gene conversion in Sa ccharomyces cerevisiae was assayed using non-tandem chromosomal direct repeat crosses and plasmid x chromosome crosses. Each cross involved identical ura3 alleles marked with phenotypically silent restriction f ragment length polymorphic (RFLP) mutations at approximately 100-bp in tervals. DSBs introduced in vivo at HO sites in one allele stimulated recombination to Ura(+) by more than two orders of magnitude. Spontane ous gene-conversion products were isolated from a related strain lacki ng a functional HO nuclease gene. The multiple markers did not appear to influence the frequency of direct repeat deletions for spontaneous or DSB-induced events. DSB-induced conversion reflected efficient mism atch repair of heteroduplex DNA. Conversion frequencies of equidistant markers on opposites sides of the DSB were similar in the direct repe at cross. In contrast, markers 5' of the DSB (promoter-proximal) conve rted more often than 3' markers in plasmid x chromosome crosses, a pos sible consequence of crossing-over associated with long conversion tra cts. With direct repeats, bidirectional tracts (extending 5' and 3' of the DSB) occurred twice as often as in a plasmid x chromosome cross i n which DSBs were introduced into the plasmid-borne allele. A key diff erence between the direct-repeat and plasmid x chromosome crosses is t hat the ends of a broken plasmid are linked, whereas the ends of a bro ken chromosome are unlinked. We tested whether linkage of ends influen ced tract directionality using a second plasmid x chromosome cross in which DSBs were introduced into the chromosomal allele and found few b idirectional tracts. Thus, chromosome environment, but not linkage of ends, influences tract directionality. The similar tract spectra of th e two plasmid x chromosome crosses suggest that similar mechanisms are involved whether recombination is initiated by DSBs in plasmid or chr omosomal alleles.