Fig. 3

Hi904 mutation disrupts a conserved putative E3 ubiquitin ligase that is highly expressed in the CNS. (A) Alignment of amino acid sequence for human, zebrafish, and fly KIAA1323 orthologs shows highly conserved regions. (B) A schematic diagram of Hi904 mRNA indicates the location of the viral insert, as well as the location and orientation of PCR primers used for RT-PCR analysis (a). RT-PCR analysis of wild-type and mutant RNA at 36 hpf shows the absence of Hi904 mRNA, but not β-actin RNA in mutants (b). (C–K) Expression of Hi904 mRNA measured by in situ hybridization in wild-type embryos at four-cell stage (C), 6 hpf (D), 10 hpf (E), 14 hpf (G), 24 hpf (H and I), 36 hpf (J), and 48 hpf (K) and in Hi904 mutant embryo at 10 hpf (F). Expression at 48 hpf (K) is prominent in the inner ears (arrow), tectum (white arrowhead), and optic stalk (black arrowhead). (L, M) Expression of Hi904 mRNA in wild-type embryos (L) and dlA^hi781 mutants (M) at 12 hpf shows an increase of Hi904 in deltaA^hi781 mutants. (N) Expression of Hi904 mRNA in wild-type embryos after injection of synthetic RNA encoding zebrafish DeltaA (marked brown by immunostaining of co-injected β-gal) that suppresses Hi904 expression. Thin line indicates midline. ANK, ankyrin domain; RF, ringer finger domain; ZZ ZnF, ZZ zinc finger domain, retroviral insertion; P, primer; β-act, β-actin.