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Fig. 5 | Journal of Experimental & Clinical Cancer Research

Fig. 5

From: OTUD7B suppresses Smac mimetic-induced lung cancer cell invasion and migration via deubiquitinating TRAF3

Fig. 5

OTUD7B deubiquitinates TRAF3 to inhibit NIK. a Flag-TRAF3 and HA-OTUD7B plasmids were cotransfected into H1299 cells. TRAF3 or OTUD7B was immunoprecipitated with an anti-Flag or anti-HA antibody. b Myc-NIK and HA-OTUD7B plasmids were cotransfected into H1299 cells. NIK or OTUD7B was immunoprecipitated with an anti-Myc or anti-HA antibody. c H1299 cells were transfected with Flag-TRAF3 and HA-OTUD7B or the empty vector. The half-life of TRAF3 was assessed following treatment with cycloheximide (CHX) for 0–6 h. MG132 was applied to inhibit TRAF3 degradation as a positive control. Cell lysates were analysed by using anti-Flag and anti-HA antibodies. d H1299 cells were transfected with Flag-TRAF3, Ub and HA-OTUD7B or the empty vector. The ubiquitin assay was performed by Western blotting using an anti-ubiquitin antibody. e H1299 cells were transfected with Flag-TRAF3 or the empty vector for 48 h and then treated with 10 μM LCL161 for 24 h. Expression of TRAF3, NIK, p100 and p52 was examined by Western blotting. β-Actin expression served as a loading control. f NF-kB transcriptional activity was analysed by a luciferase assay, and the fold increase in luciferase activity is shown. g Flag-TRAF3 and Myc-NIK plasmids were cotransfected into H1299 cells. TRAF3 or NIK was immunoprecipitated with an anti-Flag or anti-Myc antibody. Representative results are shown. Plots are the mean ± SEM of data from three independent experiments.*P < 0.05

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