Fig. 4

Metformin decreases RAD51 levels via ubiquitin/26S proteasome-mediated proteolysis. a Hs 578T and MDA-MB-231 cells were treated with 5 μM MG132, a proteasome inhibitor, for 0, 1, 2, 4, 8, or 16 h and harvested at the indicated time points for western blot analyses of RAD51 expression. Results represent mean ± SEM of five independent experiments. #, ##, ### vs. no treatment; ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 by one-way ANOVA followed by Bonferroni’s post hoc test. b Hs 578T (left panel) and MDA-MB-231 (right panel) cells were incubated with metformin (5 mM) for 24 h prior to MG132 (5 μM) treatment for 8 h. Cells were immunoblotted for RAD51 or β-actin (loading control). c Hs 578T (upper panel) and MDA-MB-231 (lower panel) cells were treated with metformin (5 mM) for 0, 6, 12, or 24 h. Protein complexes in the cell lysates were immunoprecipitated with anti-RAD51 antibodies, followed by immunoblotting with anti-ubiquitin antibodies. Whole cell lysates were immunoblotted to determine RAD51 and β-actin expression. Results represent the mean ± SEM of five independent experiments. #, ## vs. no treatment; ^#P < 0.05, ^##P < 0.01, **P < 0.01 by one-way ANOVA followed by Bonferroni’s post hoc test