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Fig. 7. Inhibition of XNF-ATc3 blocks CE in neural plate explants. (A) Neural plate explants from stage 12-12.5 were cultured and photographed hourly. Upper panels show a control neural plate explant. The middle panels show a neural plate explant of an embryo injected with 2 ng DN XNF-ATc3 and 200 pg GFP. The lower panels show GFP fluorescence. The upper graph illustrates the change in length (δL) and the change in width (δW) over time. The lower graph shows the change in neural tube length (δNT) between 2 and 4 hours. Stars indicate where the DN XNF-ATc3 injected explants are significantly different than the controls in an unpaired Student's t-test. (B) Neural plate explants incubated in 400 nM CsA (lower panels). As a vehicle control, neural plate explants were incubated in 0.33% ethanol (upper panels). The graph describes the change in length (δL) and change in width (δW) over time. Stars indicate where the CsA-treated explants were significantly different from the control and ethanol treated explants using an unpaired Student's t-test. (C) CsA treatment does not lead to cell fate changes in neural plate explants. In situ hybridization using antisense probes for XAG-1, Krox20 and HoxB9. Left panel shows two control embryos used to stage the explants (left, posterior view; right, anterior view). Middle panel presents control neural plate explants, while the right panel shows explants incubated in 400 nM CsA. |
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Fig. 3. Activation of XNF-ATc3 generates neural CE defects and anterior truncations. (A) Xenopus embryos were injected at the one-cell stage with 100 pg (middle panels) or 500 pg (right panels) of CA XNF-ATc3. Embryos injected with 100 pg CA XNF-ATc3 resembled control embryos during gastrula stages (first row, vegetal view of stage 11; second row, vegetal view of stage 12.5), but developed neural tube CE defects in 31% of embryos at neurula stages (third row: dorsal view of stage 17). Embryos injected with 500 pg CA XNF-ATc3 exhibited slight defects at stage 12.5 and neural CE defects in 81% of injected embryos. Control embryos developed no neural CE defects. (B) Xenopus embryos were injected at the one-cell stage with 50 pg CA XNF-ATc3 and analyzed for Krox20, Eng, Pax3, Sox2 and HoxB9 expression at the neurula stage. For XAG-1 in situ hybridization embryos were injected with 250 pg CA XNF-ATc3 and 100 pg lacZ (red) for lineage tracing. |
