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. Author manuscript; available in PMC: 2020 Jul 24.
Published in final edited form as: Oncogene. 2020 Jan 24;39(12):2467–2477. doi: 10.1038/s41388-020-1163-1

Fig. 2.

Fig. 2

MEK5 knockdown impairs DNA-PKcs phosphorylation in response to ionizing radiation. PC3 (a) or DU145 (b) cells were transiently transfected with Luciferase siRNA (siLUC) or siRNAs against MEK5 (#78). Four days later, cells were irradiated with 3 Gy γ-radiation, lysates were prepared at the indicated times and immunoblotted with the indicated antibodies. c PC3 cells stably expressing a control (shControl) or MEK5 (clone#12, upper; clone#22, lower) shRNA were exposed to 3 Gy of γ-rays and cells were lysed at the indicated times. Lysates were immunoblotted sequentially with the indicated antibodies. d DU145 cells stable expressing a scrambled (shControl) or MEK5 (clone#7) shRNA were exposed to 3 Gy of γ-rays and cells were lysed at different times and immunoblotted sequentially with anti-phospho-DNA-PKcs (S2056) and anti-total DNA-PKcs antibodies. e Ectopic expression of MEK5 restores activation of DNA-PKcs. PC3 cells stably expressing shControl, shMEK5 (clone#12), or shMEK5 transiently expressing MEK5-pcDNA3 vector were exposed to 3 Gy of γ-rays and lysed at the indicated times. Lysates were immunoblotted sequentially with phospho-DNA-PKcs (Ser2056), total DNA-PKcs, phospho-ATM (Ser1981) and total ATM antibodies. UI: unirradiated.