Fig. 7

Collagen detection in wild-type and uxs1 mutant skeletons. (A–D) Whole-mount (A, B) and horizontal section (C, D) in situ hybridization for col2a1a gene expression; (E, F) whole-mount immunostaining for Col2 protein; (G–L) whole-mount (G–J) and horizontal section (K, L) in situ hybridization for col10a1 gene expression. Expression of col2a1a increased in developing cartilage of uxs1^hi3357 mutants in lateral views of the head at 3 dpf (B) and ceratohyal sections at 5 dpf (D), compared to wild-type siblings (A, C). Longer substrate developing times demonstrated that col2a1a levels are high in cartilage of both wild type and mutant heads at 3 dpf (A′, B′). In contrast, although Col2a1 protein was easily detected in wild type cartilages (E) in ventral view at 5 dpf, it was not detected in mutant cartilages (F). Lateral (G, H) and ventral (I, J) whole-mount views showed that domains of col10a1 gene expression were greatly reduced in regions of endochondral and intramembranous skeletal elements in 5 dpf uxs1 mutants (H, J), compared to wild types (G, I). In situ hybridization on histological sections of the ceratohyal at 5 dpf illustrated reduced perichondral staining of col10a1 in mutants (L), compared with wild types (K). Also, chondrocyte expression of col10a1 was absent in mutants, although wild-type ceratohyal chondrocytes strongly expressed col10a1. Abbreviations: bsr, branchiostegal ray; cb1-5, ceratobranchials 1-5; ch, ceratohyal; cl, cleithrum; de, dentary; ent, entopterygoid; ep, ethmoid plate; f, fin; hm, hyomandibular; hs, hyosymplectic; m, Meckel′s cartilage; max, maxilla; op, opercle; pq, palatoquadrate; ps, parasphenoid.