Figure 8.

E-cadherin–α-catenin chimera cannot rescue tTJ formation in α-catenin KO cells. (A) Schematic of the E-cadherin–α-catenin chimeric construct. Star indicates FLAG-tag. (B) Whole-cell lysates of EpH4 WT cells, α-catenin KO cells, α-catenin KO cells expressing WT α-catenin (α-catenin res), and α-catenin KO cells expressing E-cadherin–α-catenin chimera (chimera res) were immunoblotted with the indicated antibodies. Molecular weight measurements are in kD. (C) Phase-contrast (top panels) and immunofluorescence images of WT, α-catenin KO, α-catenin rescued, and chimera rescued EpH4 cells. Cells were stained with using anti-afadin pAb as the AJ marker and anti-occludin mAb as the bTJ marker (middle and lower panels). Scale bar: 20 µm. (D) Immunofluorescence images showing anti–claudin-3 pAb (green) and an anti-FLAG mAb (magenta) staining of a co-culture of α-catenin rescued cells and chimera rescued cells. Dotted line overlays the border between α-catenin rescued cells and chimera rescued cells, which are indicated by the asterisk. Insets are enlarged below. Blue and orange insets correspond to α-catenin rescued cells and chimera rescued cells, respectively. Scale bar: 20 μm. (E) The rate of tTJ disruption was quantified as in Fig. 1 D. Student’s t test; **, P < 0.01. Source data are available for this figure: SourceData F8.