Homologous recombination systems can be divided into two broad classes depending on how double-chain breaks are treated: those that do and those that do not focus recombination at double-chain ends. Those that do include yeast meiotic and mitotic cells, several prokaryotic pathways, mammalian cells, and oocytes of Xenopus laevis. The wild-type recombinat!on pathway of E. coli (Rec) is the only known member of the second class, in which double-chain ends are not recombination hotspots. An octameric sequence, Chi, is a hotspot in the Rec pathway, and a double-chain break must be made elsewhere on a Chi-containing molecule for Chi (and Rec) to be used. Molecular mechanisms of homologous recombination in a representative of each class will be studied, using a common methodology. We are interested in Xenopus oocytes as a possible model system for meiotic recombination that can be probed using added DNAs. In the Rec system, we wish to elucidate the mechanism by which the prototypic recombinator, Chi, works. The problems in each system are different and complementary. The common methodology is genetical and physical analysis of phage lambdal and bacterial plasmid DNAs recovered as products of Xenopus-and Rec- mediated recombination. Experiments with Rec will constitute tests of two recombination models involving replication or transcription. Our work on chain bias of heteroduplex patches will be extended. Our preliminary finding that RNase H inhibits Rec-mediated resolution will be pursued. Single-chain gapped molecules will be tested as potential initiators of recombination. Tests will be made for Chi-stimulated recombination of Holliday junctions in vivo. In Xenopus oocytes, whether added DNAs recombine following the rules that meiotic chromosomes must (overall reciprocality; conservation of molecules) will be tested. Roles of double-chain breaks and of chain-specific exonuclease from oocytes will be investigated. We will ask whether substrates other than double-chain breaks are recombinagenic. Models for meiotic recombination will be tested. Whether the oocyte recombination machinery can recognize Chi will be examined.
