Fig. 6: The absence of LOXHD1 leads to BAIAP2L2 mislocalization.

a Anti-BAIAP2L2 immunofluorescence on P21 apical HCs shows a strong signal at transducing stereocilia tips which is absent in the mechanotransduction deficient IHC mutants Tmie^KO/KO and Tmc1^KO/KO. In Loxhd1^Delta/Delta, a strong BAIAP2L2 signal is detected in row 2 stereocilia but with a different distribution than control Loxhd1^Delta/+ littermates. Boxes correspond to magnified panels. Scale bars = 2 µm. Representative results from experiments replicated once (Tmie^KO) or twice (Tmc1^KO and Loxhd1^Delta). b Stereocilia classification based on the BAIAP2L2 fluorescence signal distribution pattern: only at the tip, at both the tip and along the shaft, at the shaft, or absent from stereocilia. Data are represented as mean ± SEM. The experiment was replicated twice. Per group: n_cells ≥ 8; Mann–Whitney test, two-tailed. c SUB-immunogold-SEM anti-BAIAP2L2 shows the enrichment of gold beads at the tip of rows 2 and 3 P21 IHC stereocilia in Loxhd1^+/+, while gold beads were more distant from the tip in Loxhd1^Delta/Delta. Arrowheads indicate the gold beads closest to the tips. Scale bars = 500 nm. Distance between BAIAP2L2-gold beads to tip of rows 2 (d) and 3 (e) stereocilia measured from SUB-immunogold-SEM micrographs in Loxhd1^+/+ and Loxhd1^Delta/Delta P21. IHCs corresponded to ~70% apical, ~12% medial, and ~18% basal. Per group: P21 row 2 (d) n_cells ≥ 34; n_stereocilia ≥ 168; row 3 (e) n_cells ≥ 34; n_stereocilia = 155 for Loxhd1^+/+ and 4 for Loxhd1^Delta/Delta; The experiment was replicated twice. Stereocilia classification based on the BAIAP2L2-gold distribution pattern: with at least one gold bead present regardless of its position, with at least one gold bead in the first 100 nm, with at least one gold bead below 100 nm, and without any gold beads. Represented as mean % ± SEM. Row 2 (f): per group: n_cells ≥ 33; row 3 (g): per group: n_cells = 32. Mann-Whitney test, two-tailed. For all experiments. ns (non significative) = p > 0.05; *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001; ****p ≤ 0.0001. The exact p values are presented in Supplementary Table 1. Source data are provided as a Source Data file.