Fig. 1

The flh/noto/NOT1-expressing pineal progenitor region extends into the PPR. (A-H) Dual-colour in situ hybridisation for indicated genes on bud stage zebrafish embryos (anterior is leftwards). White asterisk in A marks pitx3-positive anterior PPR; black asterisks mark irx1b-positive posterior PPR. (D,F,H) Magnified views of the boxed areas in C,E,G, respectively; arrows indicate a putative pineal progenitor region; nc marks the neural crest in F,H. (I) In situ hybridisation for flh/noto (red) and immunohistochemical detection of DLX3 (green) in an early bud stage zebrafish embryo. (J) Magnification of boxed area in I showing partial overlap between flh/noto and DLX3 expression. (K) Transverse section of late bud stage embryo stained as in I and J at the level of the diencephalon. Arrow indicates the overlap between flh/noto and DLX3-expressing cells in the PPR (PrePlac.). All zebrafish wild-type double staining experiments were performed with at least 20 embryos, two independent experiments each. Images are representative of most or all embryos for each set of markers. (L) HH8.5 chick embryo stained using dual-colour in situ hybridisation for SOX2 (red) and NOT1 (blue; anterior is upwards). Arrowhead indicates level of section in M. (M) Transverse section through the neural tube of an embryo stained as in L at the level of the posterior forebrain (arrowhead in L). There is NOT1 staining in non-neural ectoderm overlying the SOX2-positive neural tube (n=3; two independent experiments). (N) Schematic colour-coded representation of gene expression patterns shown in A-K (Di, diencephalon; Hypoth, hypothalamus; Mes, mesencephalon; Tel, telencephalon). Scale bars: in A, 50 µm for A-C,E,G; in D,H, 15 µm for D,F,H,J; in I, 50 µm; 15 µm in K; 500 µm in L; 25 µm in M.