**2.1.2 Phosphorylation on Tyrosine 315 by c-Abl**

Two years after Yuan and colleagues published their study, another group demonstrated that Rad51 can be phosphorylated. However Chen and colleagues did not observe the phosphorylation of Tyrosine 54 but detected the phosphorylation of another tyrosine residue, in position 315.

The authors used GST pull-down assays and immunoprecipitation to show that Rad51 forms a complex with c-Abl and ATM in cells. The association between the three proteins was independent of irradiation and DNA binding. The level of phosphorylation of Rad51 after irradiation of cells was investigated. The analyses of immunoprecipitated Rad51 with an anti-phosphoTyrosine antibody showed that the level of phosphorylation increases after irradiation. Rad51 was a direct substrate for c-Abl and the phosphorylation was dependent on both c-Abl and ATM. In order to determine which tyrosine residue was phosphorylated, the authors co-expressed c-Abl and wild type or mutated Rad51 in cells. Different tyrosine to phenylalanine Rad51 mutants were performed. Phenylalanine is an amino acid that cannot be phosphorylated. Thus, a signal would no longer be detected by the antiphosphoTyrosine antibody when the phosphorylated residue is mutated. The mutation of Y315 to phenylalanine abolished Rad51 phosphorylation, indicating that c-Abl phosphorylates Rad51 on this residue (Yuan et al., 1998).
