Fig. 4

sf3b1 gain- and loss-of-function analyses. (A,B) The tst^b460 phenotype is partially rescued by sf3b1 misexpression. (A) Melanophores are absent in uninjected tst^b460 mutant embryos. (B) tst^b460 homozygous mutant embryos were injected with hs>sf3b1 and heat-shocked 2 or 3 times (6 hpf, 10 or 11 hpf, 22 or 23 hpf), 1h/each time. Several melanophores present in the dorsal trunk region and over the yolk region in injected tstb460 mutant embryos (arrows). (C-F) Phenocopy of tstb460 mutant embryos using morpholino knockdown analysis. (C) Lateral view of a live uninjected wildtype embryo at 30 hpf. (D) Lateral view of a tst^b460 homozygous mutant at 30 hpf. (E) Lateral view of a live sf3b1MO3rd morphant showing the absence of melanophores (arrowheads) and CNS (brain) cell death at 30 hpf. (F) Lateral view of a live sf3b1MOATG morphant showing phenocopy of the tst^b460 mutant at 30 hpf. (G) The phenotype was more severe in the sf3b1MOATG injected tst^b460 mutant as compared with uninjected tst^b460 mutant, suggesting that the tst^b460 allele is hypomorphic. (H) Variant transcripts in sf3b1MO3rd morphants. Lane1 and lane2: RT-PCR product from uninjected wildtype total RNA. Lane 3 and lane 4: RT-PCR product from sf3b1MO3rd morphant total RNA. (I) Western blot analysis of Sf3b1 protein levels in uninjected wildtype embryos and sf3b1MOATG morphants at 12 hpf. Lane 1: uninjected wildtype embryos; Lane 2: sf3b1MOATG morphants. (J) Schematic diagram of different sf3b1 transcripts in sf3b1MO3rd morphants. (K) Schematic diagram of different sf3b1 transcripts in hi3394a mutants.