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. 2021 Jul 21;148(14):dev199549. doi: 10.1242/dev.199549

Fig. 1.

Fig. 1.

MPDZ occupies the hair cell apical membrane and is required for normal stereocilia distribution and apical junction circumference in neonates. (A) Flat-mount views of the mouse auditory epithelium immunolabeled with MPDZ (red), ZO1 (green, apical junctions) and phalloidin (blue, F-actin) at E17.5, P0 and P4 (cochlear base). MPDZ signal is entirely lost in Mpdz mutants (MpdzDEL/DEL). (B) Projection (xz) view of the immunolabeled auditory epithelium at P0 as illustrated in the schematic on the left (high MPDZ enrichment at the bare zone shown in dark magenta; lower enrichment at the medial domain, light magenta). MPDZ is detected at the HC apical membrane, but not at junctions labeled with ZO1. (C) Phalloidin (F-actin) labeling of the hair bundle (cochlear base; maximal projection). In the absence of MPDZ, OHC and IHC hair bundles are misshapen from P0. (D) TRIOBP labeling of the stereocilia rootlets at P6 (maximal projection). Stereocilia are misaligned and form interrupted or supernumerary rows (arrowheads) in Mpdz mutants. (E) ZO1 labeling of the apical junctions at P4 (cochlear base). In the absence of MPDZ, the HC circumference is abnormally rounded, and the apical surface appears reduced (apical surface shaded in blue in MpdzDEL/+, red in MpdzDEL/DEL in a single HC). (F) Apical surface area at P0 and P4 based on ZO1 labeling. Here, and in the following figures, graphical representations show a 25-75% percentile box plot in which external bars show the minimum and maximum, the internal bar shows the median and the cross shows the mean. All individual data points are also shown. P0 base: MpdzDEL/+ N=3 animals, n=13 IHC, n=45 OHC; MpdzDEL/DEL N=3, n=13 IHC, n=40 OHC. P4 base: MpdzDEL/+ N=3, n=12 IHC, n=42 OHC; MpdzDEL/DEL N=3, n=14 IHC, n=38 OHC. P4 apex: MpdzDEL/+ N=3, n=13 IHC, n=36 OHC; MpdzDEL/DEL N=3, n=13 IHC, n=35 OHC (Mann–Whitney U-test; P0: IHC P=0.0523, OHC P=0.4417; P4 base: IHC **P=0.0054, OHC ****P<0.0001; P4 apex: IHC ****P<0.0001, OHC ****P<0.0001; ns, not significant). (G) Circularity of the HC apical circumference based on ZO1 labeling. Animal (N) and cell (n) numbers are the same as in F (Mann–Whitney U-test; P0: IHC **P=0.0025, OHC P=0.3513; P4 base: IHC P=0.1274, OHC ****P<0.0001; P4 apex: IHC P=0.3825, OHC **P=0.0023; ns, not significant). Scale bars: 10 µm (A,C-E); 5 µm (B).