DNA-PK is activated by nucleosomes and phosphorylates H2AX within the nucleosomes in an acetylation-dependent manner.
Eukaryotic DNA is organized into nucleosomes and higher order chromatin structure, which plays an important role in the regulation of many nuclear processes including DNA repair. Non-homologous end-joining, the major pathway for repairing DNA double-strand breaks (DSBs) in mammalian cells, is mediated by a set of proteins including DNA-dependent protein kinase (DNA-PK). DNA-PK is comprised of a large catalytic subunit, DNA-PKcs, and its regulatory subunit, Ku. Current models predict that Ku binds to the ends of broken DNA and DNA-PKcs is recruited to form the active kinase complex. Here we show that DNA-PK can be activated by nucleosomes through the ability of Ku to bind to the ends of nucleosomal DNA, and that the activated DNA-PK is capable of phosphorylating H2AX within the nucleosomes. Histone acetylation has little effect on the steps of Ku binding to nucleosomes and subsequent activation of DNA-PKcs. However, acetylation largely enhances the phosphorylation of H2AX by DNA-PK, and this acetylation effect is observed when H2AX exists in the context of nucleosomes but not in a free form. These results suggest that the phosphorylation of H2AX, known to be important for DSB repair, can be regulated by acetylation and may provide a mechanistic basis on which to understand the recent observations that histone acetylation critically functions in repairing DNA DSBs.
Park
Eun-Jung
EJ
Division of Molecular Life Sciences and Center for Cell Signalling Research, Ewha Womans University, Seoul 120-750, Korea.
Chan
Doug W
DW
Park
Ji-Hye
JH
Oettinger
Marjorie A
MA
Kwon
Jongbum
J
D000107:Acetylation; D034961:Antigens, Nuclear / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004789:Enzyme Activation; D006367:HeLa Cells; D006657:Histones / Q000378:metabolism*; D006801:Humans; D000072200:Ku Autoantigen; D009687:Nuclear Proteins; D009707:Nucleosomes / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*
D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; C468783:H2AX protein, human; D006657:Histones; D009687:Nuclear Proteins; D009707:Nucleosomes; D004247:DNA; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen