Displacement of DNA-PKcs from DNA ends by the Werner syndrome protein.
The DNA-dependent protein kinase (DNA-PK) complex, which is composed of a DNA-dependent kinase subunit (DNA-PKcs) and the Ku70/80 heterodimer, is involved in DNA double-strand break repair by non-homologous end joining (NHEJ). Ku70/80 interacts with the Werner syndrome protein (WRN) and stimulates WRN exonuclease activity. To investigate a possible function of WRN in NHEJ, we have examined the relationship between DNA-PKcs, Ku and WRN. First, we showed that WRN forms a complex with DNA-PKcs and Ku in solution. Next, we determined whether this complex assembles on DNA ends. Interestingly, the addition of WRN to a Ku:DNA-PKcs:DNA complex results in the displacement of DNA-PKcs from the DNA, indicating that the triple complex WRN:Ku:DNA-PKcs cannot form on DNA ends. The displacement of DNA-PKcs from DNA requires the N- and C-terminal regions of WRN, both of which make direct contact with the Ku70/80 heterodimer. Moreover, exonuclease assays indicate that DNA-PKcs does not protect DNA from the nucleolytic action of WRN. These results suggest that WRN may influence the mechanism by which DNA ends are processed.
Li
Baomin
B
Department of Molecular Microbiology and Immunology, Keck School of Medicine, University of Southern California, 2011 Zonal Avenue, HMR 509, Los Angeles, CA 90089, USA.
Comai
Lucio
L
D034961:Antigens, Nuclear; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004265:DNA Helicases; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D024202:Electrophoretic Mobility Shift Assay; D005092:Exonucleases / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D000072200:Ku Autoantigen; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D004247:DNA; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; D005092:Exonucleases; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen