Additional file 1 of Development of the hyolaryngeal architecture in horseshoe bats: insights into the evolution of the pulse generation for laryngeal echolocation

Additional file 1. The coding of the craniocervical ossification or calcification in all specimens of Rhinolophus sp. and Mus musculus, examined this study

Expression of <i>Lrp4</i> and <i>Wise</i> during palatal rugae development.

(A-H) Sagittal sections of developing palate between midline and molar region showing in situ hybridization of Lrp4 (A, C, E, G) and Wise (B, D, F, H) in wild-type at E12.5 (A, B), E13.5 (C, D), E14.5 (E, F) and E16.5 (G, H). (C-H) Box showing large magnification images of developing rugae region. (I, J) Whole mount in situ hybridization of Lrp4 (I) and Wise (J) in wild-type palatal rugae at E14.5. Scale bar; 100μm (box in C-H), 200μm (A-D), 300μm (E, F), 400μm (G-J).</p

Wnt signaling in <i>Lrp4</i> and <i>Wise</i> mutant mice.

(A-C) Sagittal sections of developing palate at region between midline and molar showing in situ hybridization of Axin2 in wild-type (A), Wise (B) and Lrp4 (C) mutants at E14.5. Scale bar; 200μm. (D, E) Whole mount explants showing Lef1transcription in palatal rugae of wild-type (D) and Wise (F) mutant mice.</p

Shh and Fgf signaling in <i>Lrp4</i> and <i>Wise</i> mutants during palatal rugae development.

(A, B, Q, R) Sagittal sections of developing palate between midline and molar region showing in situ hybridization of Gli1 (A, B) and Erm (Q, R) in wild-type (A, Q), Shh-K14 (B) mice and Lrp4 mutant mice (R) at E13.5. Arrowheads indicating Erm expression in developing palatal rugae. (A) Box showing large magnification images of developing rugae region. (C, D) SEM image of palatal rugae of wild-type (C) and Shh-K14 (D) mice at birth. Arrowheads indicating palatal rugae presumptive region. (E-P) Whole mount explants showing Shh (E-N) and Erm (O, P) transcription in palatal rugae of wild-type (E, G, I, K, M, O), Lrp4 (N) and Wise (F, H, J, L, P) mutant mice at E12.5 (G, H), 13.5 (I, J, O, P), E14.0 (K, L) and E14.5 (E, F, M, N). Scale bar; 200μm.</p

<i>Wise</i> expression during palatal rugae development.

Diagrammatic representation of Wise expression during the developmen of palatal rugae. Light blue representing Wise expression. A, B; E11.5-E12.0, C; E12.0-E12.5, D; E12.5-E13.0, E; 13.0-E13.5, F; E13.5-E14.0, G; E14.5–15.0, H;16.0-E16.5. P2; Wise expression domain posterior to ruga 2, A8; Wise expression domain anterior to ruga 8, P4-1; Wise expression domain posterior to ruga 4, P4-2; Wise expression domain between P4-1 and A8 Wise expression domain, P6; Wise expression domain posterior to ruga 6.</p

Palatal rugae phenotypes in <i>Lrp4</i> and <i>Wise</i> mutant mice.

Oral views of palatal rugae in adult wild-type (A), Lrp4 (B) and Wise mutants (C). (D, E) Pseudo SEM image of palatal rugae of wild-type (D) and Wise mutant (E) mice at E13.5-E14.0. Arrowheads indicating rugae.</p

Murine palatal rugae development.

(A) SEM image of adult murine palatal rugae. (B) Diagrammatic representation of the developmental stages of palatal rugae formation (sagittal view). Mes; Mesenchyme, Epi; Epithelium.</p

<i>Wise</i> and <i>Lrp4</i> expression in <i>Lrp4</i> and <i>Wise</i> mutant mice.

Whole mount explants showing Lrp4 (A-D) and Wise (E, F) transcription in palatal rugae of wild-type (A, C, E), Wise (B, D) and Lrp4 (F) mutant mice at E13.5-E14.0 (A, B, E, F) and E14.0-E14.5 (C, D). Scale bar; 200μm (A-D), 50μm (E, F).</p

<i>Wise</i> expression during sequence of rugae appearance.

(A-L) Oral views of palatal rugae showing expression of Shh (left palatal shelf) and Wise (right palatal shelf) in wild-type. Green and red arrowheads indicating region corresponding ruga 2 and 8, respectively. Left and right panel showing developing palatal shelf obtained from same embryo. Blue arrowheads indicating region corresponding newly formed ruga. Scale bar; 200μm (A-H), 400μm (I-L). (M) Oral views of palatal rugae showing expression of Shh (red) and Wise (blue) in wild-type. A-D; E11.5-E12.0, E, F; E12.0-E12.5, G, H; E12.5-E13.0, I, J; 13.0-E13.5, K, L, M; E13.5-E14.0. Images of Wise expression were horizontally flipped.</p

Bmp signaling in palatal rugae development.

(A-L) Sagittal sections of developing palate between midline and molar region showing in situ hybridization of Bmp2 (A, B), Bmp4 (C, D), Bmp5 (E, F), Bmp7 (G, H) and Noggin (I, J) at E14.5 (A, C, E, G, I) and E16.5 (B, D, F, H, J) and immunohistochemistry of p-Smad1/5/9 (K, L) in wild-type (A-K) and Wise mutants (L) at E13.5. Box showing large magnification images of developing rugae area. (M) Schematic of Bmp signaling activity and Noggin expression in palatal rugae in sagittal view. Oral views of palatal rugae in adult Noggin-K14 (N) and Bmp4-K14 mice (O). Red arrowheads indicate disorganized rugae. Scale bar; 200μm.</p