Fig. 2. Loss of PATJ expression leads to Pals1 disorganization. (A) Cell extracts (80 µg) from control (CT Cl 8) and PATJ KD clones (Cl 4 and 12) were analyzed by SDS-PAGE and western blotting with affinity-purified antibodies against Pals1. Tubulin was used as an internal control for protein loading. (B,C) Confocal microscopic X-Y (B) or Z (C) sections at the level of tight junctions of control (CT Cl 8) and PATJ KD clone (Cl 12) of Caco2 cells. Cells grown on filters were labeled with affinity-purified antibodies against human PATJ (in red) and Pals1 (in blue). Arrowheads indicate the colocalization of the two proteins. Note that in the absence of PATJ labeling there is no accumulation of Pals1 at the tight junctions. Bars, 10 µm.