University of Tasmania
Browse

File(s) not publicly available

Molecular and biochemical characterisation of DNA-dependent protein kinase-defective rodent mutant irs-20

journal contribution
posted on 2023-05-17, 02:59 authored by Priestley, A, Beamish, HJ, David GellDavid Gell, Amatucci, AG, Muhlmann-Diaz, MC, Singleton, BK, Smith, GC, Blunt, T, Schalkwyk, LC, Bedford, JS, Jackson, SP, Jeggo, PA, Taccioli, GE
The catalytic subunit of the DNA-dependent protein kinase (DNA-PKcs) is a member of a sub-family of phosphatidylinositol (PI) 3-kinases termed PIK-related kinases. A distinguishing feature of this sub-family is the presence of a conserved C-terminal region downstream of a PI 3-kinase domain. Mutants defective in DNA-PKcs are sensitive to ionising radiation and are unable to carry out V(D)J recombination. Irs-20 is a DNA-PKcs-defective cell line with milder γ-ray sensitivity than two previously characterised mutants, V-3 and mouse scid cells. Here we show that the DNA-PKcs protein from irs-20 cells can bind to DNA but is unable to function as a protein kinase. To verify the defect in irs-20 cells and provide insight into the function and expression of DNA-PKcs in double-strand break repair and V(D)J recombination we introduced YACs encoding human and mouse DNA-PKcs into defective mutants and achieved complementation of the defective phenotypes. Furthermore, in irs-20 we identified a mutation in DNA-PKcs that causes substitution of a lysine for a glutamic acid in the fourth residue from the C-terminus. This represents a strong candidate for the inactivating mutation and provides supportive evidence that the extreme C-terminal motif is important for protein kinase activity.

History

Publication title

Nucleic Acids Research

Volume

26

Issue

8

Pagination

1965-1973

ISSN

0305-1048

Department/School

Menzies Institute for Medical Research

Publisher

Oxford Univ Press

Place of publication

Great Clarendon St, Oxford, England, Ox2 6Dp

Repository Status

  • Restricted

Socio-economic Objectives

Expanding knowledge in the biological sciences

Usage metrics

    University Of Tasmania

    Categories

    Exports

    RefWorks
    BibTeX
    Ref. manager
    Endnote
    DataCite
    NLM
    DC