Fig. 5: MCL1 negatively impacts p73 DNA binding in a p73TD-dependent manner.
From: MCL1 binds and negatively regulates the transcriptional function of tumor suppressor p73
a Direct FPA between p73 (either p73DBD alone or p73DBD-TD) and FITC-labeled DNA response elements (i.e. FITC-GADD45 or FITC-NOXA). All FPAs were performed in biological and technical triplicate. Each plot shows the three data points from one representative assay. b Quantification of the EMSA using p73DBD or p73DBD-TD with an increasing concentration of recombinant MCL1. The amount of unbound DNA was quantified through intensity values calculated by the free probe. An unpaired t-test was used for each concentration of MCL1 (DBD vs. DBD-TD), N = 3. Error is the SEM. p-value for 50 μM = 0.043, 100 μM = 0.0153, 200 μM = 0.0005. c and d Representative EMSAs on native TGX gels visualizing FITC-GADD45 DNA (F-DNA). c is the p73DBD alone whereas d is the p73DBD-TD. Coomassie staining is used to visualize the protein with increasing amounts of MCL1 added into the reaction. Lanes for both gels are as follows: (1) 200 μM MCL1 alone (No F-DNA), (2) F-DNA alone (no protein), (3) 200 μM MCL1 + F-DNA, (4) 10 μM p73 protein with 1 μM F-DNA, (5)–(9) 10 μM p73 with F-DNA+ increasing concentration of MCL1 (10–200 μM).
