Dimerization and Nuclear Localization of Ku Proteins

Abstract

Ku, a heterodimer of Ku70 and Ku80, plays a key role in multiple nuclear processes, e.g. DNA repair, chromosome maintenance, and transcription regulation. Heterodimerization is essential for Ku-dependent DNA repair in vivo, although its role is poorly understood. Some lines of evidence suggest that heterodimerization is required for the stabilization of Ku70 and Ku80. Here we show that the heterodimerization of these Ku subunits is important for their nuclear entry. When transfected into Ku-deficient xrs-6 cells, exogenous Ku70 and Ku80 tagged with green fluorescent protein accumulated into the nucleus, whereas each nuclear localization signal (NLS)-dysfunctional mutant was undetectable in the nucleus, supporting the idea that each Ku can translocate to the nucleus through its own NLS. On the other hand, the nuclear accumulation of each NLS-dysfunctional mutant was markedly enhanced by the presence of an exogenous wild-type counterpart. In Ku-expressing HeLa cells, each NLS-dysfunctional mutant, as well as wild-type Ku70 and Ku80, was still detectable in the nucleus, whereas the double mutant of each Ku subunit with decreased functions of both nuclear targeting and dimerization was undetectable in the nucleus. Our results indicate that each Ku subunit can translocate to the nucleus not only through its own NLS but also through heterodimerization with each other.

Establishment of Ku70-Deficient Lung Epithelial Cell Lines and Their Hypersensitivity to Low-Dose X-Irradiation

“…The nuclear translocation of Ku70 is, at least in part, controlled at a nuclear localization signal (NLS)-recognition step by NLS receptors and regulated by the heterodimerization with Ku80 [12][13][14][15]17]. In hamster cells, Ku70/Ku80 heterodimer accumulation at DSBs produced using a 405-nm laser starts immediately after irradiation [18], strongly supporting the idea that the heterodimer of Ku70 and Ku80 is a sensor of DSBs in the nuclei.…”

“…It was also reported that Ku70 is translocated from the cytoplasm to the nucleus in rat and mouse fibroblasts after X-irradiation [2,29]. On the other hand, we have demonstrated that the nuclear translocation of Ku70 is, at least in part, controlled at a nuclear localization signal (NLS)-recognition step by NLS receptors and regulated by heterodimerization with Ku80 [12][13][14][15]17].…”

Transient Association of Ku with Nuclear Substrates Characterized Using Fluorescence Photobleaching

“…However, a separate study has shown that equivalent amounts of Ku86 are immunoprecipitated with antiserum specific to either Ku70 or the Ku heterodimer, and dimerization is necessary for efficient nuclear translocation of each subunit (42). These results therefore indicate that nuclear Ku70 occurs as a heterodimer and that Ku associated with matrix does so as a heterodimer of Ku70 and Ku86.…”

Interaction between Ku80 protein and a widely used antibody to adenomatous polyposis coli

“…This protein complex, which is located primarily in the nucleus (Bertinato et al, 2001;Koike et al, 2001), has key roles in numerous cellular processes, including DNA repair, chromosome maintenance and regulation of transcription (Ramsden and Gellert, 1998). In most instances, the Ku heterodimer is in a complex with the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs).…”