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Association of terminal deoxynucleotidyl transferase with Ku.

Terminal deoxynucleotidyl transferase (TdT) catalyzes the addition of nucleotides at the junctions of rearranging Ig and T cell receptor gene segments, thereby generating antigen receptor diversity. Ku is a heterodimeric protein composed of 70- and 86-kDa subunits that binds DNA ends and is required for V(D)J recombination and DNA double-strand break (DSB) repair. We provide evidence for a direct interaction between TdT and Ku proteins. Studies with a baculovirus expression system show that TdT can interact specifically with each of the Ku subunits and with the heterodimer. The interaction between Ku and TdT is also observed in pre-T cells with endogenously expressed proteins. The protein-protein interaction is DNA independent and occurs at physiological salt concentrations. Deletion mutagenesis experiments reveal that the N-terminal region of TdT (131 amino acids) is essential for interaction with the Ku heterodimer. This region, although not important for TdT polymerization activity, contains a BRCA1 C-terminal domain that has been shown to mediate interactions of proteins involved in DNA repair. The induction of DSBs in Cos-7 cells transfected with a human TdT expression construct resulted in the appearance of discrete nuclear foci in which TdT and Ku colocalize. The physical association of TdT with Ku suggests a possible mechanism by which TdT is recruited to the sites of DSBs such as V(D)J recombination intermediates.

Pubmed ID: 10570175

Authors

  • Mahajan KN
  • Gangi-Peterson L
  • Sorscher DH
  • Wang J
  • Gathy KN
  • Mahajan NP
  • Reeves WH
  • Mitchell BS

Journal

Proceedings of the National Academy of Sciences of the United States of America

Publication Data

November 23, 1999

Associated Grants

  • Agency: NCI NIH HHS, Id: CA34085
  • Agency: NCI NIH HHS, Id: CA67156
  • Agency: NIAMS NIH HHS, Id: R01AR40391

Mesh Terms

  • Animals
  • Antigens, Nuclear
  • Binding Sites
  • COS Cells
  • Cell Line
  • DNA
  • DNA Damage
  • DNA Helicases
  • DNA Nucleotidylexotransferase
  • DNA-Binding Proteins
  • Humans
  • Nuclear Proteins
  • Recombinant Fusion Proteins
  • Spodoptera
  • Tumor Cells, Cultured