vps33b expression is regulated by hnf6/vhnf1. (A,B) vps33b expression in 3 dpf wild-type (cont) hnf6 morpholino-injected (A) and vhnf1mutant larvae (B) as determined by quantitative PCR. vps33b expression is reduced by 75% and 60%, respectively, in the hnf6 morpholino-injected larvae and vhnf1 mutants. Amplification levels normalized to wild type. (C-F) Whole-mount RNA in situ hybridization of 3 dpf wild-type (C,E) and hnf6 (D,F) morpholino-injected larvae using a ceruloplasmin (C,D) or a vps33b (E-F) riboprobe. Liver (black arrows) ceruloplasmin expression is unchanged in the hnf6 morpholino-injected larva, whereas decreased liver vps33b expression is evident. (G) Comparison of vps33b expression in 3 dpf wild-type larvae (cont) and in 3 dpf larvae injected with hnf6 morpholino (hnf6 MO), hnf6 morpholino and vhnf1mRNA, vhnf1 mRNA, or hnf6 mRNA. These experiments are normalized to control expression and show decreased vps33b expression (by 75%, as above) in hnf6-deficient larvae that is restored with co-injection of vhnf1 RNA. Microinjection of vhnf1 on its own increases vps33b expression (2.2×), as does hnf6 RNA to a lesser degree (1.3×). Error bars represent s.e.m. from six separate experiments.
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