Fig. 1

 Cloning and characterization of zebrafish Rab11-FIP4. (A) Schematic representation of protein structures of Rab11-FIP4 orthologues. The numbers indicate percentages of identical amino acids between any 2 proteins. Conserved motifs are indicated as boxes. EF, EF-hand; LZ, leucine-zipper-like motif; RBD, Rab11-binding domain. The predicted coiled-coil region is indicated at the top. A putative zRab11-FIP4B protein was predicted from its nucleotide sequence to be translated from Met138 of the A-form. (B) Expression of zRab11-FIP4 isoforms in various tissues of adult zebrafish was examined by semi-quantitative RT-PCR. β-actin was used as a control. (C) Expression of zRab11-FIP4 in developing zebrafish embryos. Whole-mount in situ hybridization was performed by using RNA prepared from full-length cDNA of zRab11-FIP4A as a probe. di, diencephalon; fb, forebrain; hy, hypothalamus; mb, midbrain; nr, neural retina; op, optic vesicle; ot, otic vesicle; sc, spinal cord. Scale bar indicates 200 μm. (D) Expression of A and B isoforms of zRab11-FIP4. Positions of antisense probes specific for each transcript (A and B) and common for both (common) along cDNAs are illustrated (left). The length of each probe was about 300 bases. In situ hybridization was performed by using 13-hpf embryos (right). Expression in the optic pit was pointed by arrowhead. (E) Expression of zRab11-FIP4 in the developing retina. In situ hybridization of transverse sections of embryonic heads was performed by using the same probe described above. The boxed area around the CMZ (104 hpf) is shown at higher magnification in the lower right most panel. Arrowhead indicates zRab11-FIP4 expressed in the CMZ. CMZ, ciliary marginal zone; gcl, retinal ganglion cell layer; inl, inner nuclear layer; onl, outer nuclear layer. Scale bars indicate 50 μm except in the lower right most panel, where the scale bar indicates 20 μm.