Break-induced replication: A review and an example in budding yeast

Break-induced replication (BIR) is a nonreciprocal recombination-dependent replication process that is an effective mechanism to repair a broken chrom osome. We review key roles played by BIR in maintaining genome integrity, i ncluding restarting DNA replication at broken replication forks and maintai ning telomeres in the absence of telomerase. Previous studies suggested tha t gene targeting does not occur by simple crossings-over between ends of th e linearized transforming fragment and the target chromosome, but involves extensive new DNA synthesis resembling BIR. We examined gene targeting in S accharomyces cerevisiae where only one end of the transformed DNA has homol ogy to chromosomal sequences. Linearized, centromere-containing plasmid DNA with the 5 ' end of the LEU2 gene at one end was transformed into a strain in which the 5 ' end of LEU2 was replaced by ADE1, preventing simple homol ogous gene replacement to become Leu2(+). Ade1(+) Leu2+ transformants were recovered in which the entire LEU2 gene and as much as 7 kb of additional s equences were found on the plasmid, joined by microhomologies characteristi c of nonhomologous end-joining (NHEJ). In other experiments, cells were tra nsformed with DNA fragments lacking an ARS and homologous to only 50 bp of ADEZ added to the ends of a URA3 gene. Autonomously replicating circles wer e recovered, containing URA3 and as much as 8 kb of ADEZ-adjacent sequences , including a nearby ARS, copied from chromosomal DNA, Thus, the end of a l inearized DNA fragment can initiate new DNA synthesis by BIR in which the n ewly synthesized DNA is displaced and subsequently forms circles by NHEJ.