Human Ku70 interacts with heterochromatin protein 1alpha.
Ku is involved in the metabolism of DNA ends, DNA repair, and the maintenance of telomeres. It consists of a heterodimer of 70- and 80-kDa subunits. Recently we have demonstrated that Ku70 interacted with TRF2, a mammalian telomere-binding protein. Using the same yeast two-hybrid screening system, we now show that Ku70 also interacts with heterochromatin protein 1alpha (HP1alpha), a protein known to be associated with telomeres as well as heterochromatin. HP1 is a suppressor of the position effect variegation in Drosophila and acts as a transcriptional suppressor in mammalian cells. The interaction with Ku70 in the two-hybrid system was confirmed by a glutathione S-transferase pull-down study using bacterial recombinant proteins in vitro. The interaction was also reproduced in vivo in HeLa cells, where endogenous Ku70 coimmunoprecipitated with HP1alpha. This interaction was more effective in acidic pH and weakened considerably as the pH of the reaction buffer was elevated up to 7.5. Ku80 did not interact with HP1alpha directly. The interaction domains of Ku70 and HP1alpha included the Leu-Ser repeat (amino acids 200-385) and the chromo shadow domain, respectively. Ku70 was largely colocalized with transfected HP1alpha but not with a C-terminal deletion mutant, HP1alpha(Delta)C. In contrast to HP1alpha, Ku70 did not repress transcriptional activity of the reporter gene when tethered to DNA after transfection to mammalian cells. The implication of this interaction is discussed.
Song
K
K
Department of Biochemistry, University of Ulsan College of Medicine, Seoul 138-736, Korea.
Jung
Y
Y
Jung
D
D
Lee
I
I
D034961:Antigens, Nuclear; D001665:Binding Sites; D000090266:Chromobox Protein Homolog 5; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism*; D004265:DNA Helicases; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D000072200:Ku Autoantigen; D009687:Nuclear Proteins / Q000378:metabolism*; D012097:Repressor Proteins / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins; D014162:Transfection
D034961:Antigens, Nuclear; C000715594:CBX5 protein, human; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; C081647:high affinity DNA-binding factor, S cerevisiae; D000090266:Chromobox Protein Homolog 5; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen