PARENT SESSION
1:30 PM to 3:30 PM
Monday, April 22, 2002
Poster Session 12 DNA Damage and Repair I
Room: Nevada Exhibition Center
(P17-160) Persistent ATM activation in Ku80-/- fibroblasts.
Burma, Sandeep*,1, Kurmasa, Akihiro2, Cary, Robert3, Iliakis, George4, Li, Gloria5, Jackson, Stephen6, Chen, David1, 1 Lawrence Berkeley National Laboratory, Berkeley, CA2 Tottori University, Tottori, JAPAN3 Los Alamos Laboratory, Los Alamos, NM4 Thomas Jefferson University, Philadelphia, PA5 Memorial Sloan-Kettering Cancer Center, New York, NY6 Wellcome/CRC Institute, University of Cambridge, Cambridge, UK
ABSTRACT-
DNA double-strand breaks (DSBs) generated by ionizing radiation (IR) result in the activation of Ataxia Telangiectasia Mutated (ATM) kinase which, in turn, phosphorylates and activates a number of downstream targets including the tumor suppressor p53 leading ultimately to cell cycle arrest. These breaks also constitute a high affinity target for Ku, a highly abundant heterodimer of 70kDa and 80kDa subunits (Ku70/Ku80). Ku is the DNA-binding component of the DNA-dependent protein kinase (DNA-PK) and is believed to recruit and activate the catalytic subunit DNA-PKcs at the sites of DNA damage. Here we demonstrate that IR-induced ATM activation, p53 phosphorylation, and p21 induction are persistent in Ku80-/- mouse fibroblasts, but not in DNA-PKcs-/- fibroblasts. p53 phosphorylation and cell cycle arrest are persistent in Ku80-/- cells and transient in DNA-PKcs-/- cells even when these cells are irradiated with "equal survival" doses at which they are expected to harbor approximately equal levels of unrepaired DSBs. Moreover, both DNA-PKcs-/- and Ku80-/- cells are grossly impaired in DSB rejoining as measured by pulsed-field gel electrophoresis. Therefore, persistent ATM activation in Ku80-/- cells appears not to be due to the persistence of unrejoined DSBs per se and may be indicative of a novel function of Ku in regulating ATM kinase activation. In vitro, Ku can inhibit the binding of ATM to DNA ends suggesting that Ku may act to directly or indirectly prevent persistent ATM activation in response to endogenous or exogenous DNA damage. This work was funded by US Dept. of Energy and NIH grant CA50519 to D.J.C.
KEYWORDS: ATM, Ku, DNA double-strand breaks, DNA-PK