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. 2015 Jan 14;4(2):197–205. doi: 10.1242/bio.201410629

Fig. 2. Y520 and Y667 contained within the YXXΦ motif are important for basolateral targeting of prestin in MDCK cells.

Fig. 2.

The figure shows MDCK cells transiently transfected with wild type prestin and mutations of prestin: Y520Q, Y526Q, Y616Q and Y667Q. The cells were fixed after 36 hours. Prestin was tagged with YFP at its C-terminus (green) and the cells were stained with an antibody to β-catenin. Shown are serial X-Y confocal sections along the z axis (A). The corresponding X-Z sections are shown at the bottom. Mutations at Y520 and Y667 result in a failure to target the basolateral surface of the cell along with intracellular retention of the protein and apical trafficking. (B,C) Pearson's correlation of prestin with β-catenin and a ratio of prestin fluorescence on the surface of the cell compared to the total in the cell. The mean Pearson's correlation values were: wt 0.23 (+/−0.028SE, n = 7); Y520Q −0.045 (+/−0.016 SE, n = 7); Y526Q 0.17 (+/−0.025 SE, n = 14); Y616Q 0.175 (+/−0.033 SE, n = 14); Y667Q 0.01386A (+/−.03 SE, n = 10). The differences between wt and Y520Q and Y667Q were considered significant. A one way ANOVA (parametric) with Bonferroni post test comparison yielded a p value of <0.001 for wt vs Y520Q and wt vs Y667Q; wt vs Y616Q and Y667Q were not significant. The mean surface to total ratios were: wt 0.67 (+/−0.016 SE, n = 22); Y520Q −0.39 (+/−0.0129 SE, n = 20); Y526Q 0.78 (+/−0.013 SE, n = 13); Y616Q 0.73 (+/−0.02 SE, n = 13); Y667Q 0.40 (+/−.044 SE, n = 13). Here too the differences between wt and Y520Q and Y667Q were significant. A one-way ANOVA (parametric) with Bonferroni post-test comparison yielded a p value of <0.001 for wt vs Y520Q and wt vs Y667Q. The scale bar is 10 microns.