Figure 7.

The Overexpression of the SF-Dlg3^YA1+2 Nedd4(-2)-Binding Mutants Dominantly Interferes with AB Cell Polarity In Vivo

(A) Immunoblot (anti-Flag) showing the equal expression of SF-Dlg3 (lane 2) or SF-Dlg3^YA1+2 (lane 3) proteins in stable ESC clones. Untransfected ESCs are shown as negative control, and Actin served as loading control.

(B) Lateral views of control, SF-Dlg3, and SF-Dlg3^YA1+2 ESC-derived embryos. Note the axis elongation defects in the SF-Dlg3^YA1+2 embryos at E7.75 (scale bars, 200 μm) and E8.5 (scale bars, 300 μm).

(C and D) Colocalization of the TJ marker ZO-1 with SF-Dlg3 (C) or SF-Dlg3^YA1+2 (D) as revealed by quantitative statistical colocalization on two-color confocal images. Raw images used for colocalization measurements are presented in (E) and (F) (overlays).

(E-H) Analysis of TJ formation by whole-mount immunofluorescence in the endoderm (E and F), ventral node (G), and ectoderm (H). Note the epithelial holes (F, white arrowheads) and rupturing (G) in SF-Dlg3^YA1+2 mutants as compared to the control (E). Scale bars represent 25 μm.

See also the graphical abstract.