Homologous recombination in eukaryotic organisms can occur between similar sequences at identical positions on homologous chromosomes (allelic recombination) or can involve dispersed repeated sequences (ectopic recombination). Recombination between dispersed repeats promotes sequence homogeneity between members of multigene families (concerted evolution), but also can generate novel sequences that increase genetic diversity. In addition, crossing-over between dispersed repeats can result in chromosomal rearrangements that not only affect an organism's reproductive fitness, but also can have direct impact on the organism. Genome rearrangements, for example, can have profound effects on the expression of genes near a breakpoint, and such changes in gene expression have been implicated in the development of certain cancers. In most cases, recombination between naturally-occurring repeated sequences is difficult, if not impossible, to detect selectively and to manipulate experimentally. Consequently, most of our current knowledge about ectopic recombination has come from studying interactions between artificially-constructed repeats. The genetically-amenable yeast Saccharomyces cerevisiae is particularly well-suited for studies of this sort since it is straightforward to construct artificial duplications of defined size and sequence at pre-selected genomic locations. The three major objectives of the proposed work center on examining aspects of ectopic recombination in S. cerevisiae. First, two parameters (substrate length and substrate homology) thought to be important in modulating the frequency of ectopic recombination will be examined systematically. The effect of substrate length on the frequency and resolution of ectopic interactions will be examined using heteroalleles of varying size. The effect of substrate homology will be addressed using a novel system involving recombination between partially homologous (homeologous) intronic sequences. The possible involvement of mismatch repair systems in limiting homeologous interactions also will be investigated. Second, the effect of crossing-over between nonhomologous chromosomes on meiotic chromosome segregation will be examined. Data obtained will indicate whether ectopic exchanges are functionally equivalent to exchanges involving homologous chromosomes in their ability to direct meiotic chromosome segregation. Third, an ectopic system will be used to identify novel types of recombination mutants in which gene conversion and exchange events are uncoupled. Such mutants are not readily identifiable using conventional allelic recombination systems and thus should provide additional information concerning the mechanism of recombination.
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