Identification of a conserved minimal Xath5 distal enhancer that promotes retinal expression in vivo. (A) Pairwise mVISTA (http://genome.lbl.gov/vista/index.shtml) analysis of X. laevis and X. tropicalis Xath5 5′ distal noncoding sequences identifies a 1 kb highly conserved region. (B) A series of nested deletion constructs were generated to identify a minimal 152 bp enhancer that is sufficient to promote retinal specific expression. + strong expression, -/+ very weak expression, - no expression (C) A stage 33 pG1X5 distal 1 kb transgenic embryo shows retinal GFP expression. (D) The 152 bp distal enhancer also drives GFP in the retina. (E) A pG1X5 distal 73 bp transgenic embryo does not express GFP in the retina but only expresses in the pineal gland and olfactory placodes. (F–H) The zc38 transgenic zebrafish line exhibits specific expression in the retina as shown in confocal z-projections at 18 hpf (F), 24 hpf (G), 52 hpf (H) Anterior is to the right in all panels.

FGF signaling acts to regulate Xath5 expression through the 3.3 kb enhancer sequence but this effect is not mediated via the 152 bp distal enhancer. Overexpression of XFD mRNA in pG1 X5 3.3 kb transgenic embryos inhibits transgene expression on the injected side (A–B). Ath5 expression is blocked in zebrafish embryos treated with SU5402 vs. DMSO treated embryos (C–D). zc38 transgenic embryos treated with SU5402 express GFP throughout the retina, similar to DMSO treated controls (E–F).

Whole mount zc38 transgenic zebrafish showing transgene expression specifically in the retina. Anterior is to the right. ∗Yolk autofluorescence.