Although increasing evidence has suggested that the hMSH5 protein plays an important role in meiotic and mitotic DNA recombinational repair, its precise functions in recombination and DNA damage response are presently elusive. Here we show that the interaction between hMSH5 and c-Abl confers ionizing radiation (IR)-induced apoptotic response by promoting c-Abl activation and p73 accumulation, and these effects are greatly enhanced in cells expressing hMSH5(P29S) (i.e. the hMSH5 variant possessing a proline to serine change within the N-terminal (Px)(5) dipeptide repeat). Our current study provides the first evidence that the (Px)(5) dipeptide repeat plays an important role in modulating the interaction between hMSH5 and c-Abl and alteration of this dipeptide repeat in hMSH5(P29S) leads to increased IR sensitivity owing to enhanced caspase-3-mediated apoptosis. In addition, RNAi-mediated hMSH5 silencing leads to the reduction of apoptosis in IR-treated cells. In short, this study implicates a role for hMSH5 in DNA damage response involving c-Abl and p73, and suggests that mutations impairing this process could significantly affect normal cellular responses to anti-cancer treatments.