Fig. 3.

Par3 expression is important for both Ca²⁺ switch and AMPK-mediated zonula occludens-1 (ZO-1) localization to the plasma membrane. Madin-Darby canine kidney (MDCK) cells grown to confluency were incubated in media containing 5 μM Ca²⁺ [low-Ca²⁺ media (LCM)] for 16 h before being introduced to fresh LCM, LCM containing 2 mM AICAR, or media containing 1.8 mM Ca²⁺ (Ca²⁺ switch) for 1 or 2 h as indicated. A and C: cells were fixed and immunostained for ZO-1. Scale bar, 30 μm. Arrowheads indicate sites of junction assembly. B and D: quantification of ZO-1 localization at sites of junction assembly. Data were analyzed for each experimental condition based on four randomly obtained views from three independent coverslips. Error bars represent the SE of the length of Par3 per cell within each of the four selected fields of view. ****Significant difference due to the presence of AICAR (P < 0.0001) or the Ca²⁺ switch (P < 0.0001) compared with LCM conditions based on Student’s t-test. KD, knockdown; WT, wild type.