Impaired binding of ZO-2 to the phospho-mimetic Occ-T400E/T404E/S408E construct. A) The indicated GST-Occ cytoplasmic tail fusion proteins were used to pull down HA-ZO-2 from transiently transfected MDCK C11 cells with GSH-agarose beads. Isolated protein complexes were analyzed by Western blotting with anti-HA antibody. Equal amounts of GST- fusion proteins were pulled down as detected with an anti-GST antibody in the lower panel. B) Quantification of 5 independent experiments as shown in (A). C) Purified GST-occludin C-terminal domain (GST-OccC) was prephosphorylated in vitro by purified CK2 and subsequently used to pull down FLAG-tagged ZO-2 from transiently transfected HEK-293 cells. Association of FLAG-ZO-2 was analyzed by Western blotting with the anti-FLAG M2 antibody. D) Densitometric quantification of 4 experiments as shown in (C).