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. 2021 Aug 27;7(35):eabb3799. doi: 10.1126/sciadv.abb3799

Fig. 5. 53BP1–lamin B1 interaction is dissociated after irradiation.

Fig. 5

(A) Fate of the lamin B1–53BP1 interaction after DNA damage. Quantification of PLA kinetics of the endogenous interaction after irradiation (2 Gy) from three independent experiments (left). Quantification of PLA kinetics upon lamin B1 overexpression after irradiation (2 Gy) from three independent experiments (right). (B) Impact of ATM activity inhibition on 53BP1–Lamin B1 dissociation after IR induced DNA damages. Proteins from cellular lysates treated with ATM inhibitor (ATMi, +) or vehicle (−) were subjected to immunoprecipitation before (NIR) or 1 hour after irradiation (2 Gy) with rabbit antibodies against lamin B1 or IgG as control with DNAse treatment. Coimmunoprecipitated proteins were revealed by Western blot with mouse anti-53BP1 and rabbit anti–lamin B1 antibodies (left) and quantified from four independent experiments (right). (C) Impact of 53BP1 N-terminal phosphorylation on the dissociation between lamin B1 and 53BP1 after IR. 53BP1 knockout cells transfected with WT 53BP1 vector (53BP1 WT) or with 28A mutant 53BP1 vector (53BP1 28A) were subjected to PLA using anti-53BP1 and lamin B1 antibodies, before (NIR) or 1 hour after irradiation (2 Gy). Example of in situ interaction between 53BP1 constructs and lamin B1 visualized as red fluorescent dots (left). Quantification of PLA dots per nucleus from three independent experiments is shown (right). For (B) and (C), percentage of lamin B1–53BP1 association after irradiation was normalized to corresponding nonirradiated conditions. Error bars, SEM. Scale bar, 10 μm. Unpaired t test P values **P < 0.005; ***P < 0.0001.