A Novel Small Peptide Inhibitor of NFκB, RH10, Blocks Oxidative Stress-Dependent Phenotypes in Cancer

Figure 1

Cloning and testing of RH truncated mutants. (a) To test the ability of truncated mutants to bind IκB, we performed a coimmunoprecipitation assay, using Myc tag for immunoprecipitation. The first two lines are negative controls: (1) agarose beads + Ab anti-Myc; (2) agarose beads + secondary Ab + cell lysate; lines 3–6 include agarose beads + Ab anti-Myc + lysate of cells transfected as described in the legend. Line 7 is a positive control (cell lysate). Images show the ability of the mutants to precipitate IκB. Since the mutants are too small to be identified by electrophoresis, immunoglobulins (Ig) were used as a control of immunoprecipitation. The images are the representative of the results from the three independent experiments. (b) IκB and caspase 3 levels were assessed by Western blot in response to mutants expression. Actin was used as loading control. All the mutants are able to increase both IκB and caspase 3 levels. The images are the representative of the results from the three independent experiments. Densitometric analysis (bar graph) shows fold of increase of IκB and cleaved caspase 3 levels vs. control ( vs. control). Data are reported as mean ± SD.

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