The Drosophila melanogaster DmRAD54 gene plays a crucial role in double-strand break repair after P-element excision and acts synergistically with Ku70 in the repair of X-ray damage

The RAD54 gene has an essential role in the repair of double-strand breaks (DSBs) via homologous recombination in yeast as well as in higher eukaryote s. A Drosophila melanogaster strain deficient in the RAD54 homolog DmRAD54 is characterized by increased X-ray and methyl methanesulfonate (MMS) sensi tivity. In addition, DmRAD54 is involved in the repair of DNA interstrand c ross-links, as is shown here. However, whereas X-ray-induced loss-of-hetero zygosity (LOH) events were completely absent in DmRAD54(-/-) flies, treatme nt with cross-linking agents or MMS resulted in only a slight reduction in LOH events in comparison with those in wild-type flies. To investigate the relative contributions of recombinational repair and nonhomologous end join ing in DSB repair, a DmRad54(-/-)/DmKu70(-/-) double mutant was generated. Compared with both single mutants, a strong synergistic increase in X-ray s ensitivity was observed in the double mutant. No similar increase in sensit ivity was seen after treatment with MMS. Apparently, the two DSB repair pat hways overlap much less in the repair of MMS-induced lesions than in that o f X-ray-induced lesions. Excision of P transposable elements in Drosophila involves the formation of site-specific DSBs. In the absence of the DmRAD54 gene product, no male flies could be recovered after the excision of a sin gle P element and the survival of females was reduced to 10% compared to th at of wild-type flies. P-element excision involves the formation of two DSB s which have identical 3' overhangs of 17 nucleotides. The crucial role of homologous recombination in the repair of these DSBs may be related to the very specific nature of the breaks.