Fig. 9

hnf-6 regulates expression of vhnf1 and other liver genes. (A–D) Relative expression of vhnf1, hnf-4, hnf-3β, and ceruloplasmin (cer) genes in 3 dpf WT and hnf-6 morphant larvae. Data derived from real-time quantitative PCR using tbp gene expression as a standard. Error bars represent SEM of five separate experiments on pools of five to eight larvae. These data show that hnf-6 positively regulates vhnf1, hnf-4, and hnf-3β. Specificity of these results is supported by the normal expression of the hepatocyte marker cer in the hnf-6 morphants (D). (E–F) Left dorsolateral views of 3 dpf WT (E) and hnf-6 morphant (F) larvae processed for vhnf1 RNA in situ hybridization. Strong expression in the paired pronephric ducts (arrows) is seen in both larvae. However, there is minimal vhnf1 expression in the liver of the hnf-6 morphant (arrowhead). The larva shown is representative of 40% of hnf-6 morphants, with a range of expression patterns evident in the remainder. (G–I) Confocal projections through the liver of 5 dpf WT (G), hnf-6 morphant (H) larvae, and a larva coinjected with the hnf-6 morpholino and vhnf1 RNA (I). There is partial rescue of the hnf-6 morphant biliary phenotype in the vhnf1 RNA-injected larva (I). Ducts in this larva are narrowed and longer than in the uninjected morphant (H). Short, connecting ducts are also present in the vhnf1 RNA-injected larva. (J–L) Schematized versions of (G–I). Long ducts were traced in blue, interconnecting ducts in green, and terminal ductules in red. Compared with the wild-type larvae (J), there are shorter long ducts (blue), and fewer interconnecting ducts (green) and terminal ductules (red) in the hnf-6 morphant larva (K). Partial phenotype rescue occurs with coinjection of vhnf1 RNA and the hnf-6 morpholino (L).