Figure 4.

USP7 Depletion in APC-Truncated CRC Suppresses Wnt Activation by Restoring β-Catenin Ubiquitination

(A–C) Relative TOP/FOP activities of the APC4 (A), SW480 (B), and HEK293T (C) cells with control or USP7 CRISPR targeting.

(D) AXIN1 complexes were immunoprecipitated in APC4 cells with or without USP7 targeting followed by western blotting using the indicated antibodies.

(E) Cells were treated with cycloheximide (Chx) (50 μg/μL), and lysates were collected at different time points as indicated for immunoblotting of ABC (active β-catenin) and control GAPDH.

(F) Colony formation assay in parental HEK293T, SW480, Caco2, and APC4 cells and the corresponding USP7 CRISPR-deleted cells.

(G) Quantitation of number of colonies in (F). Experiments were performed in triplicates.

(H) Colony formation assay of APC4 and Caco2 cells upon transient transfection of USP7 CRISPR and/or β-catenin S33Y mutant plasmids. The corresponding quantitation is indicated on the right.

(I–K) Relative TOP/FOP activities of the HCT116 p53 WT (I and J) and HCT116 p53^−/− (K) cells with indicated CRISPR targeting.

Error bars represent SE from at least three independent experiments (^∗∗p < 0.01; ^∗∗∗p < 0.001; ns, not significant). See also Figure S4.