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. 2001 Jan 23;98(3):1036–1041. doi: 10.1073/pnas.021282898

Figure 5.

Figure 5

HU neither requires functional ATM, nor interferes with the γ IR-dependent activation of ATM kinase. (A) HCT116 cells were pretreated with wortmannin in DMSO (W; lanes 4 and 7, 20 μM; lanes 2, 5 and 8, 50 μM) or DMSO (lanes 1, 3, and 6) for 2 h, then either exposed to γ IR (10 Gy) or HU (1.5 mM) for 6 h, and cell extracts were analyzed by Western blotting using either PAb 1801 (p53) or phospho-serine 15-specific antibodies (pS15). Actin was used as a loading control. NT, not treated. (B) Normal and AT−/− lymphoblasts were untreated (−) or treated with γ IR (3.5 Gy). Cells were harvested at the indicated hours after treatment, and lysates from each sample were analyzed as described in A. (C) Normal and AT−/− lymphoblasts were either untreated (−) or treated with HU (1.5 mM). Cells were harvested at the indicated hours and analyzed as in A. (D) SAOS-2 cells were grown without treatment (−) or exposed to HU treatment for 13 h (HU and HU + γ) or subjected to γ IR for 1 h (HU+ γ and γ). All samples were treated with MG132 proteosome inhibitor for 2 h before lysis to allow HDM-2 accumulation. Total cellular extracts were immunoblotted with the two indicated mAbs to HDM-2. Actin was used as a normalization control.