Plk3 interacts with CtIP to promote resection in G1. (a) Interaction of GFP-CtIP and Plk3 after 5 Gy IR. HEK293T cells were treated with CtIP siRNA and transfected with a GFP-CtIP plasmid. GFP-CtIP or Plk3 were obtained by IP, and the precipitates were analyzed by immunoblotting. (b) Interaction of Plk3-ΔPBD with CtIP-S327A and CtIP-T847A at 30 min after 5 Gy. HEK293T cells were treated with CtIP and Plk3 siRNAs and transfected with GFP-CtIP and SFB-Plk3 plasmids. Plk3 was obtained by IP, and the precipitates were analyzed by immunoblotting. (c) CtIP phosphorylation in a Plk3-ΔPBD mutant in vivo. HEK293T cells were treated with CtIP and Plk3 siRNAs, transfected with GFP-CtIP and SFB-Plk3 plasmids, and irradiated with 5 Gy. GFP-CtIP was obtained by IP and analyzed for pS327 at 30 min after IR or pT847 at 2 h after IR by immunoblotting. (d) HEK293T cells were treated with CtIP siRNA, transfected with GFP-CtIP plasmids, and irradiated with 5 Gy. GFP-CtIP was obtained by IP and analyzed for pS327 or pT847 by immunoblotting. (e) pRPA foci in Flag-positive (Flag⁺), G1-phase HeLa cells after 2 Gy α-particle irradiation. Cells were treated with Plk3 siRNA and transfected with SFB-Plk3 plasmids. Transfection efficiencies were confirmed by Western blotting (±SEM from three experiments). siCtrl, control siRNA; siPlk3, Plk3 siRNA. ***, P < 0.001.