Fig. 1: MRE11 SUMOylation is increased in response to DNA damage. | Nature Communications

Fig. 1: MRE11 SUMOylation is increased in response to DNA damage.

From: Crosstalk between SUMOylation and ubiquitylation controls DNA end resection by maintaining MRE11 homeostasis on chromatin

Fig. 1

a, b HeLa cells were treated with or without 20 μM ML792 for 1 h and the indicated times of 1 μM CPT (a), or the indicated doses of ML792 and 1 μM CPT for 1 h (b) and the whole cell extracts were subjected to immunoblotting. c, d ML792 repressed pRPA2 (S4/S8) foci formation in S/G2 phase HeLa cells after 1 μM CPT treatment for 45 min. Scale bar, 10 μm. The data are presented as means ± SD, n (DMSO) = 117 cells, n (ML792) = 119 cells. e MRE11 was modified by SUMO1/2/3. HEK293T cells were cotransfected with SFB-MRE11 and 10×His-SUMO1/2/3, followed by Ni-NTA pull-down with guanidine denaturing buffer. f NBS1 SUMOylation was tested as in e. g MRE11 SUMOylation was enhanced by treatment with DSB-inducing agents (1 μM CPT for 2 h, 100 μM etoposide for 3 h, and 1 μM cisplatin for 3 h) but not HU (2 mM, 3 h). h MRE11 SUMOylation increased after 1 μM CPT treatment in a time-dependent manner. i NBS1 SUMOylation was downregulated upon treatment with DNA-damaging agents as in g. j NBS1 SUMOylation gradually decreased along CPT treatment. k The indicated purified proteins were analyzed by Coomassie blue staining, and then used in reconstituted reactions to test MRE11 SUMOylation in vitro. l SUMOylation of endogenous MRE11 was examined in HEK293T cells treated with or without CPT (1 μM, 6 h). m HEK293T cells expressing SFB-MRE11 and His-SUMO2 were fractionated into soluble and chromatin fractions, followed by the analysis of MRE11 SUMOylation.

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