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. 2001 Jul 15;15(14):1833–1844.

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Copyright © 2001, Cold Spring Harbor Laboratory Press

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The ATM phosphorylation site in E2F1 is required for DNA damaged induction. (A) 293T cells were transfected with HA-E2F1 or HA-E2F3 and then treated with neocarzinostatin (NCS) for the indicated time. HA-tagged E2F proteins were detected by immunoblotting with an HA-specific antibody. (B) Degradation of E2F1 by the proteasome. 293T cells were transfected with HA-E2F1, HA-E2F1^S31A, or HA-E2F1^ΔN85 and then treated with MG-132 at 20 μM for 5 h. Proteins were detected by immunoblotting with an HA specific antibody. (C) 293T cells were transfected with HA-E2F1, HA-E2F1^S31A, or HA-E2F1^ΔN85 and then treated with NCS for the indicated time. Proteins were detected by immunoblotting with an HA specific antibody. (D) Induction of E2F1 in response to DNA damage requires ATM. 293T cells were transfected with HA-E2F1 along with HA-ATM^Ki or vector. The cells were then treated with NCS for the indicated time. Transfected E2F1 was detected with an HA-specific antibody (top). Expression of transfected ATM^ki protein was confirmed by immunoprecipitation with HA beads followed by immunoblotting with ATM antibody (Ab-3) (bottom).