PTPL1 is required for apical membrane clustering. (A) Western blot of W4:shCTRL or W4:shPTPL1 lysates probed for PTPL1 and β-catenin. (B) Localization of Lifeact-Ruby in W4:shCTRL or PTPL1-depleted W4 cells. Scale bars, 5 μm. (C) Quantification of apical membrane size in W4:shCTRL and W4:shPTPL1 cells. Red bars represent the average. Red error bars represent the SEM (n > 17). *, P < 0.05 using independent sample t tests. (D) Relative PTPL1 mRNA expression levels as determined by qPCR. Values in the graph represent the average. Error bars represent the SEM (n = 6; 2 biological and 3 technical replicates). *, P < 0.05 using independent sample t tests. (E, F, and G) Localization of the brush border marker GFP-EBP50 (E), apical membrane marker CD66 (F), and basolateral domain marker CD71 (G) in polarized W4:shCTRL and W4:shPTPL1 cells in combination with the actin marker Lifeact-Ruby or phalloidin-Alexa Fluor 568 and DAPI. Scale bars, 5 μm.