FOXP3 is an X-linked tumor suppressor gene and a master regulator in T regulatory cell function. This gene has been found to be mutated frequently in breast and prostate cancers and to inhibit tumor cell growth, but its functional significance in DNA repair has not been studied. We found that FOXP3 silencing stimulates homologous recombination-mediated DNA repair and also repair of g-irradiation-induced DNA damage. Expression profiling and chromatin- immunoprecipitation analyses revealed that FOXP3 regulated the BRCA1-mediated DNA repair program. Among 48 FOXP3-regulated DNA repair genes, BRCA1 and 12 others were direct targets of FOXP3 transcriptional control. Site-specific interaction of FOXP3 with the BRCA1 promoter repressed its transcription. Somatic FOXP3 mutants identified in breast cancer samples had reduced BRCA1 repressor activity, whereas FOXP3 silencing and knock-in of a prostate cancer-derived somatic FOXP3 mutant increased the radioresistance of cancer cells. Together our findings provide a missing link between FOXP3 function and DNA repair programs. © 2012 American Association for Cancer Research.