Abstract
Understanding the subcellular localization and regulatory mechanisms of DNA repair proteins is critical for elucidating the mechanisms of genotoxicity. Localization of DNA repair proteins is highly spatiotemporally regulated, and their dysregulation causes of various diseases, including cancer. Dysregulation also modulates cytotoxicity and genotoxicity caused by medical radiation and environmental mutagens. Non-homologous end joining (NHEJ) is the most important but error-prone pathway in DNA double-strand break (DSB) repair. NHEJ is initiated by binding of the Ku70/Ku80 heterodimer to the DSB. If a DSB is not repaired correctly, cells may die or undergo mutations. We have previously shown that EYFP-canine Ku70 localizes to the nuclei of canine cells. However, the mechanism through which canine Ku70 localizes to the nucleus remains unclear. In this study, we provide the first experimental evidence that canine Ku70 localizes to the nucleus and that its predicted nuclear localization signal is bound by the nuclear pore-targeting complex, importin-α/β. In addition, the transfected EYFP-canine Ku70 localized to the nucleus in human, hamster, and mouse cells, suggesting that canine Ku70 translocates and localizes to the nucleus via a common mechanism conserved among these four species. We also found that Ku80 is essential for the accumulation of canine Ku70 in DSBs. These findings provide fundamental information for understanding the regulatory mechanisms of Ku70 and the molecular mechanisms underlying the cytotoxicity and genotoxicity induced by DSBs in dogs.