Regulation of wnt-signaling by A20 through RIPK4.

<p>A) Wild-type (WT RKO) or A20 knockout RKO (KO RKO) were transfected with a TCF4-luciferase reporter and either a control or RIPK4 siRNA. Cells were stimulated with wnt3a for 8 hours. Luciferase activity was measured and normalized to Renilla luciferase. B) Wild-type (WT RKO) or A20 knockout RKO (KO RKO) were transfected with a TCF4-luciferase reporter and either a control or a RIPK4-K51R plasmid at different concentrations. Cells were stimulated with wnt3a for 8 hours. Luciferase activity was measured and normalized to Renilla luciferase. C) Wild-type (WT RKO) or A20 knockout RKO (KO RKO) were transfected with MYC-tagged RIPK4 and an HA-tagged K48-only ubiquitin construct. Cells were stimulated with wnt3a for the indicated time points. Lysates were subjected to immunoprecipitation with an antibody against MYC and then blotted with an antibody against HA. Inputs are shown below. Tubulin is shown as a loading control. ** = p < 0.01. Each panel is representative of at least three independent experiments.