Fig. 1.

Fig. 1.

Tenocytes at cranial muscle attachments express scxa, sox9a and col1a. (A-C) Single-plane ventral views, anterior to the left, of 72 hpf zebrafish embryos showing in situ hybridization chain reaction (isHCR) staining for expression of sox9a (green), scxa (magenta) and col1a1a (cyan) mRNA at cranial muscle-cartilage attachments (entheses). (D) Similar view of cell nuclei labeled with DAPI. (E-G) Z-stack ventral views of 72 hpf Tg(sox10:gfp;BAC scx:mCherry) embryos stained by immunofluorescence for GFP (E), mCherry (F) and myosin heavy chain (MHC; G). (H) Cartoon showing a ventral view of a 72 hpf embryo illustrating chondrocytes expressing sox9a (green) and tenocytes expressing scxa (red), with dual expression of both genes in tenocytes of entheses. (I-K) Z-stack ventral views, anterior to the left, of 96 hpf embryos stained by immunofluorescence for Col2a (I), Tsp4b (J) and Col2a/Tsp4b colocalization (K). (L-N) Z-stack ventral views of 5 dpf embryos stained by immunofluorescence for Col1a (L), Tsp4b (M) and Col1a/Tsp4b colocalization (N). Arrowheads indicate tendons (white) and ligaments (orange) attaching with mc, ch, pq and bh cartilages. Yellow asterisks show the developing entheseal ECM of the ch-SH tendon. (O) Cartoon showing ECM organization of a tendon attaching a muscle to a mammalian long bone, with fibrocartilage and Col2a⁺ matrix near the bony junction (green), Col1a-rich matrix in the tendon midbody (blue) and Tsp4 (red) in the myotendinous junction (MTJ). (P) Cartoon showing ECM organization of the elongated tendon in zebrafish attaching the SH muscle to the ch cartilage, with similar Col2a⁺ (green), Col1a⁺ (blue) and Tsp4⁺ (red) defining similar regions. AM – adductor mandibularis; bh-IMP-IH, basihyal-intermandibularis posterior-interhyal; ch, ceratohyal; ch-HH, ceratohyal-hyohyal tendon; ch-IH, ceratohyal-interhyal tendon; ch-SH, ceratohyal-sternohyoideus tendon; HH, hyohyal; IMA; intermandibularis anterior; IMP, intermandibularis posterior; Mc, Meckels cartilage; mc-AM, Meckels adductor mandibularis tendon; mc-IMA, Meckels-intermandibularis anterior tendon; mc-pq, Meckels-palatoquadrate ligament; pq, palatoquadrate; pq-AM, palatoquadrate-adductor mandibularis tendon. Scale bars: 20 μm.