Fig. 6

Fig. 6

a The Wnt reporter gene expressing cell line, HEK293STF, was transfected with control (CTRL) siRNA or one of two distinct CRBN siRNA and treated with PBS or Wnt3a (100 ng/mL) for 48 h. Luciferase activity was subsequently determined and normalized to total protein concentration. A quantification of Wnt reporter activity (mean ± SEM, n = 3 independent experiments) is shown. b HEK293STF cells were transfected with a control plasmid or a plasmid encoding wild-type (WT) Flag-tagged CRBN, or the indicated Flag-tagged CRBN mutants for 48 h. Luciferase activity was determined and normalized to total protein concentration. A quantification of Wnt reporter activity (mean ± SEM, n = 3 independent experiments) is shown. c A schematic of a mouse intestinal organoid showing the presumptive crypt (C) and villus (V) regions. d Mouse intestinal organoids were infected with control (Ctrl) or one of two distinct Crbn shRNA (marked by GFP expression) and then cultured in (i) basal media or (ii–iv) 25% exogenous Wnt conditioned media (L-WRN) for 5 days. Representative images are shown (n = 3 independent experiments). Scale bar = 100 μm. e. Quantification of the percent of organoids that exhibit a more basal-like phenotype (mean ± SD, n = 3 technical replicates) in a representative experiment (n = 3 independent experiments) is shown. Asterisks in a, b, and e indicate statistical significance (two-tailed Student’s t-test, **p value < 0.01, ****p value < 0.0001).