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Fig. 6.
ZNF703/tlp-1 is a novel regulator functioning as an Msx/vab-15 modulator both in worm lateral neuroblasts and vertebrate NPB. (A) Embryonic expression of worm ZNF703/tlp-1. A comma-stage embryo carrying the Ptlp-1::H1::mCherry reporter trangene is shown. Arrowheads indicate P neuroblasts. Arrow indicates the mother cell of V5 and Q neuroblasts. (Scale bar, 10 μm.) (B–E) Phenotypes of ZNF703/tlp-1 and Msx/vab-15 mutant worms of migration and differentiation of lateral neuroblasts. The y axis represents the number of VD neurons recognized by the Punc-25::GFP marker (B) or the fraction of worms showing the specified defect (C–E). The number of scored worms is shown. (F) The expression of ZNF703 in Xenopus embryos from stage (st)12 to st19. (Scale bar, 0.5 mm.) The red dot lines represent the location of cross section. (G) Whole-mount in situ hybridization assay in Xenopus embryos (st16 to st17) to examine the effect of MO knockdown of Msx/vab-15 and/or ZNF703/tlp-1 on the expression of the neural plate marker Sox2, neural crest specifier FoxD3, and Rohon–Beard sensory neuron progenitor marker Runx1 and DRG progenitor marker Islet1. Asterisks indicate the injected side of an embryo. The red lines and green lines indicate the length of left and right parts of neural tubes, respectively. # indicates mRNA resistant to the ZNF703 MO. Fractions of scored embryos with phenotype are shown. (Scale bar, 0.5 mm.) Error bars represent standard errors. |
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Fig. S3.
Assessment of effects of ZNF703 and Msx1 depletion in Xenopus. (A) Immunoblots (IBs) showing that ZNF703 MO efficiently blocks translation of an MO-complementary Znf703-HA mRNA but not an MO-resistant Znf703-HA# mRNA. (B) Immunoblots showing that Msx1 MO efficiently blocks translation of an MO-complementary Msx1-HA mRNA but not an MO-resistant Msx1-HA# mRNA. β-Actin serves as a loading control in A and B. (C) Depletion of ZNF703 alone through injecting a mix of MOs up to 40 ng has no discernable effects on the expression of foxd3, sox2, runx1, and islet. However, depletion of Msx1 using 10 to 15 ng MO discernably decreased the expression of foxd3, runx1, and islet1 and expanded the expression domain of sox2. Asterisks indicate the injected side of an embryo. (Scale bar, 0.5 mm.) |
