XRCC4 (X-ray cross-complementation group 4) is a protein associated with DNA ligase IV, which is thought to join two DNA ends at the final step of DNA double-strand break repair through non-homologous end-joining. It has been shown that, in response to irradiation or treatment with DNA damaging agents, XRCC4 undergoes phosphorylation, requiring DNA-PK. Here we explored possible role of ATM, which is structurally related to DNA-PK, in the regulation of XRCC4. The radiosensitizing effects of DNA-PK inhibitor and/or ATM inhibitor were dependent on XRCC4. DNA-PK inhibitor and ATM inhibitor did not affect the ionizing radiation-induced chromatin recruitment of XRCC4. Ionizing radiation-induced phosphorylation of XRCC4 in the chromatin-bound fraction was largely inhibited by DNA-PK inhibitor but further diminished by the combination with ATM inhibitor. The present results indicated that XRCC4 phosphorylation is mediated through ATM as well as DNA-PK, although DNA-PK plays the major role. We would propose a possible model that DNA-PK and ATM acts in parallel upstream of XRCC4, regulating through phosphorylation.