Figure 3.
Figure 3. Loss of Sip1 prevents the completion of neural crest EMT. Whole-mount in situ hybridization of embryos electroporated with ContMO (A and C) or Sip1MO (B and D) at HH4 and collected at HH10. A and B, Sox10 expression; C and D, FoxD3 expression. While ContMO-electroporated embryos have normal expression of both genes, Sip1-electroporated embryos exhibit decreased expression of (B) Sox10 in the migratory crest (n = 11/13) and aberrantly maintain (D) FoxD3 in the dorsal neural tube (n = 7/10). (b′–d′′) Sections from levels indicated by dotted lines in B and D. (E) In situ hybridization for Sox10 on an HH13 embryo electroporated at HH4 with Sip1MO showing a maintenance of neural crest near the midline at the level of the midbrain (asterisk; n = 4/5). (F) Western blot analysis of Sip1 protein expression in HH9–10 embryos after injection of 1 mM ContMO, 1 mM Sip1MO, and 2 mM Sip1MO showing that Sip1 protein expression is greatly reduced after Sip1MO electroporation. The data shown here are representative of one experiment out of three repeats. In this experiment, protein was pooled from 8–10 chicken whole embryos. (G and H) Whole-mount immunohistochemistry for HNK1 (magnification of 10) comparing neural tube explants from embryos electroporated with either (G) Sip1MO (n = 8/8) or (H) ContMO (n = 6/6) showing lack of migration and clumping phenotype. Dashed lines indicate distance migrated by furthest cells from the explant. Explants are overlaid with bright-field images. (g′ and h′) A 40× image from dashed box in explants (G and H, respectively) demonstrating HNK1-positive cells that are either clumped outside of the neural tube in Sip1MO-injected explants (g′) or that have migrated away from the ContMO-injected explant (h′). (g′ and h′) High magnification image of explants showing HNK1 expression and Sip1MO or ContMO alone showing that HNK1-positive cells are clumped next to the explant (g′, white arrows) or have migrated away from the explant (h′, white arrowhead). Bars: (A–E) 200 µm; (b′–d′′) 150 µm; (G and H) 100 µm; and (g′ and h′) 30 µm. The data shown here are from a single representative experiment out of at least three repeats.

Loss of Sip1 prevents the completion of neural crest EMT. Whole-mount in situ hybridization of embryos electroporated with ContMO (A and C) or Sip1MO (B and D) at HH4 and collected at HH10. A and B, Sox10 expression; C and D, FoxD3 expression. While ContMO-electroporated embryos have normal expression of both genes, Sip1-electroporated embryos exhibit decreased expression of (B) Sox10 in the migratory crest (n = 11/13) and aberrantly maintain (D) FoxD3 in the dorsal neural tube (n = 7/10). (b′–d′′) Sections from levels indicated by dotted lines in B and D. (E) In situ hybridization for Sox10 on an HH13 embryo electroporated at HH4 with Sip1MO showing a maintenance of neural crest near the midline at the level of the midbrain (asterisk; n = 4/5). (F) Western blot analysis of Sip1 protein expression in HH9–10 embryos after injection of 1 mM ContMO, 1 mM Sip1MO, and 2 mM Sip1MO showing that Sip1 protein expression is greatly reduced after Sip1MO electroporation. The data shown here are representative of one experiment out of three repeats. In this experiment, protein was pooled from 8–10 chicken whole embryos. (G and H) Whole-mount immunohistochemistry for HNK1 (magnification of 10) comparing neural tube explants from embryos electroporated with either (G) Sip1MO (n = 8/8) or (H) ContMO (n = 6/6) showing lack of migration and clumping phenotype. Dashed lines indicate distance migrated by furthest cells from the explant. Explants are overlaid with bright-field images. (g′ and h′) A 40× image from dashed box in explants (G and H, respectively) demonstrating HNK1-positive cells that are either clumped outside of the neural tube in Sip1MO-injected explants (g′) or that have migrated away from the ContMO-injected explant (h′). (g′ and h′) High magnification image of explants showing HNK1 expression and Sip1MO or ContMO alone showing that HNK1-positive cells are clumped next to the explant (g′, white arrows) or have migrated away from the explant (h′, white arrowhead). Bars: (A–E) 200 µm; (b′–d′′) 150 µm; (G and H) 100 µm; and (g′ and h′) 30 µm. The data shown here are from a single representative experiment out of at least three repeats.

This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal