A NEW-TYPE OF ESCHERICHIA-COLI RECOMBINATIONAL HOTSPOT WHICH REQUIRES FOR ACTIVITY BOTH DNA-REPLICATION TERMINATION EVENTS AND THE CHI-SEQUENCE

Homologous recombination on the chromosome is in many cases uniform, but in both procaryotes and eucaryotes there are specific regions Or sites, named ''hotspots'', where homologous recombination occurs at a higher rate. DNA replication origin in procaryotes (phage) is one example (1). Another example is the ''HOT1'' site in yeast, which has activity to stimulate recombination, homologously, in adjacent regions (2). The molecular mechanisms involved in enhancing homologous recombinations are not well understood. Microscopically, there is a site, the homologous recombination of the surrounding region of which is stimulated. The ''Chi'' site is such a recombinational hotspot and was first identified in lambda phage (3-5). The Chi site enhances recombination not just in its vicinity but even as far away as 10 kb (6, 7). The Chi consists of an 8 bp specific sequence, 5'-GCTGGTGG-3', distributed in Escherichia coli chromosomal DNA (one per 5 to 15 kb on the average) (8, 9). RecBCD, which is a Chi responsive enzyme, enters into duplex DNA, probably through a double-stranded (ds)-break (''cos'' site in the case of lambda phage) and moves in it with concomitant DNA degradation. The exonuclease activity of the enzyme seems to be modulated by Chi only when the enzyme approaches Chi from the correct side, the result being an enhancement of homologous recombination at the surrounding region (6, 10-18). For various analyses, the lambda phage system has been most extensively used, because in the E. coli system there are numerous Chi sites and the entrance for the enzyme on the circular chromosome has yet to be identified (6, 19). While a recombinational hotspot site on the E. coli chromosome has been identified, mechanisms related to enhancement remain unknown (20). Recently, we identified a new type of recombination hotspot, termed Hot, and a detailed analysis was made (21, 22). We obtained evidence that the homologous recombination is closely linked with DNA replication fork blocking events.