The tm4sf4 gene and regulation by nkx2.2a is conserved in zebrafish. (A) CLUSTAL W species alignment between human, mouse and zebrafish Tm4sf4. Identical (*), highly conserved (:) and weakly conserved (.) amino acids are indicated. Small/hydrophobic residues (red), acidic residues (blue), basic residues (magenta), hydroxyl/amine/basic residues (green) are shown. (B,C) Temporal qRT-PCR tm4sf4 mRNA analysis of early (B) and late (C) stage zebrafish embryos. (D-F) tm4sf4 in situ hybridization on wild-type (D,E) and nkx2.2a morphant (F) embryos. In, intestine; L, liver; P, pancreas. Images taken at 20× magnification. (G-I) qRT-PCR comparing uninjected (blue) and nkx2.2a morphant (red) embryo expression of tm4sf4 (G), insulin (H) and ghrelin (I). Error bars represent s.e.m.