Fig. 3. Expression of CD9 redistributes TGF to the apical site in polarized epithelial cells. MDCK cells that express CD9, TGF or both proteins were grown in Transwell chambers to allow them to fully establish their polarized phenotype. (A) Cells were fixed and stained for CD9 or TGF, and immunofluorescence was visualized by confocal microscopy (xz section). (B) Expression of CD9 does not alter the distribution of transmembrane TGF that lacks the TGF core sequence in polarized epithelial cells. MDCK cells stably expressing CD9, TGFE or both were grown in Transwell chambers to establish a polarized phenotype. Cells were fixed, stained for CD9 or TGFE, and immunofluorescence was visualized by confocal microscopy (xz section). (C) Cells were subjected to biotinylation of cell-surface proteins in the apical or basolateral compartments, and cell-surface TGF and CD9 were visualized by immunoblotting. Arrow indicates the position of TGF form III. (D) Same experiment as in C, but visualizing the biotinylated E-cadherin or p58, two proteins that are largely expressed in the basolateral compartment, and the apical marker gp135.