Fig. 7. Shroom2 recruitment at the forming TJs after calcium switch in MDCK cells. The lines were used to calculate the fluorescence intensity in line scan. In micrographs, dots with numbers indicate fluorescence intensity at cell edges. Blue asterisks indicate nontransfected cells that do not express GFP-mShrm2. In the fluorescence intensity profiles, vertical blue lines delineate the emplacement of these cells that were used as an internal control for fluorescence quantification. After 20 hours of incubation in low-Ca²⁺ medium (t=0), cells were fixed and analyzed 1 hour (t=1 hour), 2 hours (t=2 hours) and 7 hours (t=7 hours) after Ca²⁺ repletion. At t=0, the fluorescence intensity profiles of transfected cells indicate that both ZO-1 (red) and GFP-mShrm2 (green) are distributed throughout the cytoplasm. At t=1 hour, ZO-1 is recruited at cell-cell junctions in a discontinuous pattern, whereas GFP-mShrm2 is not yet present at the junctions. At t=2 hours, GFP-mShrm2 is mainly detected at cell-cell junctions and the intensity of the labeling at the junctions correlates with that of ZO-1. At t=7 hours, ZO-1 and GFP-mShrm2 display continuous fluorescence patterns along the junctions, and both protein labelings are more intense at tricellular junctions. Bars, 20 µm.