|
Fig. 5 Sdc2 controls DFC/KV-specific expression of Tbx16, upstream of both Fgf2-dependent KV morphogenesis and Fgf2-independent cilia length regulation. (A-F) KV morphogenesis and cilia length defects in spadetail (spt; tbx16) mutants. (A,C) Normal KV morphology (A) and cilia length (C) in wild type or spt heterozygotes. (B,D) spt mutants had defective KV morphogenesis (B) and cilia length (D), similar to sdc2 morphants. (E,F) Quantification of (E) cilia length and (F) cilia number (spt+/?: eight embryos, 361 cilia; spt-/-: nine embryos, 242 cilia) indicated that both are decreased in spt mutants (*P<0.05, **P<0.001). Error bars indicate s.e.m. (G-L) Sdc2 regulates Tbx16 expression in DFCs/KV. (G,H) Green arrowheads indicate (G) tbx16 RNA and (H) Tbx16 protein expression in DFCs in wild-type embryos. (I,J) Red arrowheads indicate the position of DFCs lacking (I) tbx16 mRNA and (J) Tbx16 protein expression in sdc2 morphants. By contrast, tbx16 RNA and Tbx16 protein expression in adjacent non-DFC domains was normal, revealing a domain-specific regulation of Tbx16 by Sdc2. (K) fgf2 was downregulated in spt mutants (compare with wild type in Fig. 4B). (L) Synergy between Sdc2, Tbx16 and Fgf2 was tested by injection of subeffective amounts of each MO individually (sdc2 SB2 MO, n=431; tbx16 MO, n=396; fgf2 MO, n=475) compared with co-injections of pairs of subthreshold MOs. Co-injection of sdc2 SB2 and tbx16 MOs (n=251) and co-injection of tbx16 and fgf2 MOs (n=111) resulted in significantly affected spaw expression (P<0.001) compared with single injections of subthreshold MOs. P-values by Student’s t-test.