Fig. S3

ret-expressing cells in the opercular muscles are not derived from neural crest (NC) and connective cell markers are present in Ret signalling mutants. (A,B) Injection of morpholinos to tfap2a and tfap2c results in a loss of NC cells (data not shown), but ret is still expressed in the arches (1,2) of tfap2+c morphants (B) compared with wild type (WT; A). (C,C′) A comparison of ret expression in the arches (1,2) to GFP⁺ NC cells in fli1:GFP animals (C), shows no expression of ret in NC cells. In the second arch the ret⁺ cells are surrounded by GFP⁺ cells, which suggests that they are mesodermal (C′). (D,E) Connective tissues and opercular bone (op, arrowhead) labelled by zns-5 appear normal in ret^hu2846 mutants (D) compared with WT (E). (F,G) ten-w expression in presumptive connective tissues (arrowhead) associated with the forming opercular bone (op) is unchanged in stm^hy024 mutants (F) at 65 hpf compared with WT (G). (H,I) At 4 dpf, ten-w-expressing cells on the opercular bone are associated with the ao and lo muscles (arrowheads) and are likely to be tendon precursors. Expression of ten-w is still present in these tendon cells in ret^hu2846 mutants (I) relative to WT (H), but is reduced, reflecting the reduced number of muscle fibres connecting to the bone. (J-M) At 3 dpf and onwards, ten-c expression is in presumptive connective cells (arrowheads) associated with the opercular bone. ten-c expression in these cells is not obviously reduced in either stm^hy024 mutants (K) or ret^hu2846 mutants (M) compared with WT (J,L). ah, adductor hyoideous; allg, anterior lateral line ganglia; ao, adductor operculi; e, eye; lo, levator operculi. Scale bars: 100 μm in A-E; 20 μm in C′,F-M.