Fig. 3.

The β-catenin γ-catenin DKO cells kept epithelial molecules. (A) Schematic representation and position of the gRNA target site in exon 3 of the γ-catenin gene (JUP). Targeted and PAM sequences are red and blue, respectively. (B) Genome sequence analysis. Two γ-catenin–negative clones were isolated, and extracted genomic DNA was sequenced. One clone showed a 1-nucleotide deletion in the proximity of the PAM site, and another clone showed a 1-nucleotide insertion. At least eight genome sequences of cell clones were analyzed, and all the clones gave the same results. (C) Cell morphology and immunofluorescent staining. Phase contrast microscopy shows that βγ-DKO cells acquired spheroidal morphology. Immunofluorescent staining using the anti–γ-catenin or anti–E-cadherin antibody. “Hygro” means that only the hygromycin resistance gene was transfected and denotes control cells. Scale bar, 50 µm. (D) Immunoblot analysis revealed that the βγ-DKO cells lost γ-catenin expression but retained the expression of E-cadherin and showed no changes in the p120-catenin splicing pattern. Vinculin served as a loading control.