DEV015719 Supplementary Material

Files in this Data Supplement:

• Supplemental Table S1 -

  Adobe PDF

• Supplemental Figure S1 -

  Fig. S1. Onset of Smyhc gene expression in different muscles. ISH with smyhc1 (1 to 249 bp), smyhc2 (1 to 155 bp) or smyhc3 (1 to 129 bp) probes. Flat mounts of 12-somite stage embryos show adaxial cells express smyhc1 but not smyhc2 or smyhc3. Lateral views at 24 hpf, 48 hpf and 72 hpf show that smyhc2 is expressed from 48 hpf in the sca, iob and icp/scp, and smyhc3 is expressed in certain craniofacial muscles from 72 hpf. Transverse section of 24 hpf posterior trunk with smyhc1 (1 to 249 bp). ISH shows expression in the superficial slow fibres (green arrow) and muscle pioneers (pink arrow). Abbreviations: icp, infracarinalis posterior; iob, inferior obliquus; lap, levator arcus palatini; sca, supracarinalis anterior; scp, supracarinalis posterior; tv, transversus ventralis.

• Supplemental Figure S2 -

  Fig. S2. slow troponin C is expressed in all S58 positive slow fibres and in the pectoral fin muscle. (A) At 96 hpf slow troponin C (stnnC) is expressed in the slow fibres throughout the embryo and also in pectoral fin muscle. (B) At 96 hpf, slow troponin C ISH shows co-localisation with slow MyHC S58 antigen in the smyhc1:gfp-positive superficial slow fibres and in the sca muscle (dorsal view anterior trunk), in els and iob muscles (lateral view anterior trunk), and in scp and icp muscles (lateral view posterior tail). The pectoral fin muscle expresses slow troponin C but not slow MyHC S58 antigen. Scale bars: 25 µm. Abbreviations: els, embryonic lateralis superficialis; icp, infracarinalis posterior; iob, inferior obliquus; pf, pectoral fin; sca, supracarinalis anterior; scp, supracarinalis posterior.

• Supplemental Figure S3 -

  Fig. S3. smyhc1 expression in the craniofacial muscles is prdm1 independent. (A) Lateral views of wild-type sib and ubo (prdm1)^tp39 mutant head at 96 hpf with immunofluorescence to the general muscle MF20 antigen and cartilage Col2 II-II6B3 antigen. In the ubo (prdm1)^tp39 mutant, perturbations to the craniofacial muscles are associated with the major loss of skeletal elements in the posterior head mutant. The ceratobranchial cartilage is absent, and the sternohyoidus and inferior obliquus form abnormal attachments to the posterior basicranial commissure cartilage instead of the cleithrum. (B) Ventrolateral views of wild type and nrd (prdm1)^m805 mutants at 96 hpf with smyhc1 (1 to 249 bp), smyhc2 (1 to 155 bp) or smyhc3 (1 to 129 bp) ISH showing prdm1-independent expression. nrd (prdm1)^m805 mutants lack posterior transversus ventralis muscles. Scale bars: 25 µm. Abbreviations: bc, basicranial commissure; cb, ceratobranchial; iob, inferior obliquus; sh, sternohyoidus; tv, transversus ventralis.

• Movie 1 -

  Movie 1. After BrdU exposure at 18 hpf, smyhc1:gfp-expressing slow fibres are BrdU labelled only at the myotome margin. Animation scrolling from superficial to medial through a segment of a confocal stack showing a lateral view of the dorsal half of a somite at the level of the anus at 48 hpf following BrdU exposure at 18 hpf. BrdU staining is shown top left and in red and smyhc1:gfp is shown bottom left and in green. Co-localisation of smyhc1:gfp and BrdU is observed only in the nucleus of the smyhc1:gfp fibre closest to the dorsal margin.