Fig. 1.

Impaired vestibular and auditory function in B2-Sox9^cY440X/+ mutants. (A) Schematic diagram of the Sox9^cY440X/+ targeted allele comprising exon 3 flanked by LoxP sites followed by exon 3′ carrying the Y440X mutation and IRES-EGFP sequences. FRT, flippase recognition target. UTR, untranslated region. (B and C) Immunostaining of SOX9^Y440X in WT and Sox9^Y440X/+ (CD) ES (B) and cochlea (C) at E14.5 (n = 2). (D–F) B2-Sox9^cY440X/+ mutants displayed impaired vestibular function. (D) Reaching response: forelimbs stretched scored 1, curled ventrally scored 0, and curled slightly scored 0.5. (E) Contact righting: flipped back to upright position within 5 s scored 1, crawled upside down scored 0, and crawled sideways scored 0.5. (F) Negative geotaxis: turned and moved up the slope surface scored 1, failed to stay on the slope surface scored 0, and failed to turn and stayed scored 0.5 (WT n = 5; B2-Sox9^cY440X n = 6). (G) Thresholds for auditory brain stem response (sound pressure level [SPL]); 120-dB SPL was the highest level measured (n = 5). (H) Endocochlear potential in 4- to 7-mo-old WT and B2-Sox9^cY440X/+ mice (n = 4). (I and J) Cochlear ducts from WT (I) and B2-Sox9^cY440X/+ (J) adult animals at 6 mo. (K) Three-dimensional reconstruction of WT and B2-Sox9^cY440X/+ adult cochlea. Scala vestibule/scala media, scala media, and scala tympani are in blue, yellow, and pink, respectively. (L–S) Cochlear ducts from WT and B2-Sox9^cY440X/+ embryos at E14.5 (L and M), E15.0 (N and O), E15.5 (P and Q), and E16.5 (R and S; n = 2 to 4). (T and U) Transverse sections of WT and B2-Sox9^cY440X/+ ED (ed; arrowhead) at E16.5 (n = 2). Data are presented as means ± SEM. Results for the CD mutants are from the constitutively active Sox9^Y440X/+ samples unless labeled as conditional. ch, chondrocyte; co, cochlea; es, ES; hsd, horizontal semicircular duct; psd, posterior semicircular duct; sac, saccule; utr, utricle. (Scale bars: 40 µm in B and C; 100 µm in M, O, Q, and S; 400 µm in I, J, and U.) **P < 0.01; ***P < 0.001.