Fig. 7. Localization of Ser827 phosphorylated ASIP/PAR-3 in mature and immature cell-cell contacts. ASIP/PAR-3 phosphorylated at Ser827 (A,D) and overexpressed T7-tagged ASIP/PAR-3 (B,E) were labeled with Cy3 and FITC, respectively. Four hours after Ca²⁺ switch, the overexpressed ASIP/PAR-3 (green; B,C) in mature cell-cell contacts overlaps the immunoreactivities of ASIP/PAR-3 phosphorylated at Ser827 (red; A,C, arrows). In contrast, 1 hour after switching, the immunoreactivities for ASIP/PAR-3 phosphorylated at Ser827 predominantly occupy the most apical tip of the immature cell-cell contacts (D,F; filled arrowhead), whereas overexpressed L-ASIP WT is detected also in the region slightly basal to the apical tip (E, F; unfilled arrowhead). The merged view of an X-Z optical section is shown in F with filled and unfilled arrowheads corresponding to the levels of the X-Y optical sections shown in G and H, respectively. The distance between the two X-Y optical sections is 1.75 µm. Merged X-Z views of other representatives are shown in I, J and K. Anti-S827-P antibody strongly stains cell-cell junctions of MDCK cells expressing T7-tagged ASIP/PAR-3 (L), whereas the antigen peptide abolishes the signals (M) in cell-cell junctions where T7-tagged ASIP/PAR-3 is concentrated (N). The distribution of overexpressed T7-tagged ASIP/PAR-3 in the same field as in M is shown in N. Bars, 10 µm.