Mechanosensitive nature of EPLIN and E-cadherin localization. (A) Schematic drawing of the points for laser irradiation. Peripheral actin bundles (a) or a part of the ZA (b) were cut with a laser. (B) Time-lapse images of EPLIN-EGFP after the laser irradiation at “a” (asterisk). Arrows indicate the original position of pAJ, which was inferred by the reduced EPLIN signals. EPLIN immediately accumulated at this position, with a concomitant up-regulation of EPLIN along the entire contiguous junctions (arrowhead). See also Video 5. A typical example of multiple experiments is shown. (C) Choosing two time points in B, cell–cell boundaries are traced with dotted lines. Relative fluorescence intensity across the cell boundaries was also measured at the sites marked as “x” and “y.” (D) Time-lapse images of EPLIN-EGFP after laser irradiation at “b.” Arrows in the first panel point to the original linear EPLIN signals: these signals were temporarily disordered or faded out, but reappeared at 5:00, as indicated by arrowheads in the last panel. Arrowheads in middle panels point to a transient up-regulation of EPLIN fluorescence signals, in which fluorescence increased ∼30% at 2:51. See also Video 7. (E) DLD1 cells double-transfected with E-cadherin–mKOR and EPLIN-EGFP were stretched to ∼150% of their original length. A typical colony is shown. Line profiles of fluorescence of E-cadherin–mKOR or EPLIN-EGFP signals across the junction, indicated by arrows, are shown at the right. Other junctions exhibiting similar changes are indicated with arrowheads. Quantification of fluorescence showed that the peak intensity of E-cadherin–mKOR and EPLIN-EGFP signals increased 18% (18 ± 08, n = 15, P < 0.05) and 17% (17 ± 07, n = 15, P < 0.05), respectively, in the junctions of stretched cells. Data represent mean ± SEM. Three independent experiments were performed. before, before stretch; after, after stretch. Time points are given in minutes and seconds. Bars, 10 µm.