Fig. 7: SENP3 deSUMOylates MRE11 mainly after DNA end resection. | Nature Communications

Fig. 7: SENP3 deSUMOylates MRE11 mainly after DNA end resection.

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

Fig. 7

a HeLa cells were cotransfected with SFB-MRE11, His-SUMO2, and the indicated siRNA, followed by the treatment of 1 μM CPT for 8 h. MRE11 SUMOylation was analyzed by Ni-NTA pull-down and immunoblotting. b The knockdown efficiency of SENPs as in a was detected by quantitative real-time PCR (means ± SEM, n = 3 independent samples). c MRE11 SUMOylation in HEK293T cells expressing Myc-tagged SENP3 or SENP3-CI (the catalytically inactive mutant, C532S) was analyzed as in a. d, e The mutual interaction between MRE11 and SENP3 was examined by co-IP. f HeLa cells expressing Myc-tagged SENP3 were treated with 1 μM CPT for 1 h, and then subjected to immunofluorescence assay with or without pre-extraction. Scale bar, 10 μm. g SENP3-knockdown HeLa cells were transfected with SFB-MRE11 and His-SUMO2. After the treatment with 1 μM CPT for 6 h, soluble and chromatin fractions were isolated, and MRE11 SUMOylation was examined by Ni-NTA pull-down and immunoblotting. h SENP3-knockdown HeLa cells were treated with 1 μM CPT and released as indicated times. Then, MRE11 foci were analyzed. Scale bar, 10 μm. The data are presented as means ± SD, n (siCon+CPT 1 h; siSENP3+CPT 1 h; siCon+Release 9 h; siSENP3+Release 9 h) = 115; 104; 120; 107 cells. i SENP3-knockdown cells were treated with 1 μM CPT for 1 h and released for 9 h. DAPI staining was performed, and micronuclei were counted in over 500 cells. Data are presented as means ± SEM, n = 3 independent experiments. Scale bar, 10 μm.

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