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Fig. 2. Nanos1-depleted PGCs enter apoptosis. (A) PGCs deficient in Nanos1 are not apoptotic throughout early development (stage 18). WISH Xpat (germ plasm marker, green) and TUNEL staining (red) of WT, Nanos1-Ctrl-MO- and Nanos1-MO-injected embryos. Arrows indicate TUNEL-positive cells in neural ectoderm. (B) TUNEL-positive PGCs are detected during tailbud stage (stage 28) in Nanos1-depleted embryos, but not in control embryos. Arrows indicate TUNEL-positive PGCs. Confocal images were taken from the region indicated in the diagram (red box). Scale bars: 100 μm in A; 150 μm in B. |
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Fig. 7. VegT RNA is stabilized in PGCs after Nanos1 knockdown. (A) VegT PBE mediates the degradation of the reporter message DsRED. PGCs were detected by either WISH for Xpat (top panels) or for DsRED antisense probe (bottom panels) in tailbud stage 31 embryos. Note that the DsRED reporter with a PBE was specifically degraded. Arrows indicate PGCs. (B) Real-time PCR analysis of VegT expression at stages 8 and 10. Expression of VegT in Nanos1 knockdown samples was normalized to that of wild-type samples. Real-time PCR was performed in duplicate, and experiments were repeated three times and showed a similar pattern. Error bars indicate s.e. P-value by unpaired Student’s t-test. |
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Fig. 1. Nanos1-depleted Xenopus embryos are deficient in PGCs. (A) Nanos1-MO blocks the translation of endogenous nanos1. Confocal analysis showing the expression of Nanos1 (red) and Xiwi (germ plasm marker, green) at the eight-cell stage of embryos previously injected with Nanos1-Ctrl-MO or Nanos1-MO at the one-cell stage. Uninjected embryos served as controls. Arrows indicate Nanos1 signal. (B) Knockdown of Nanos1 results in loss of PGCs. WISH Xpat analysis of stage 37/38 wild-type (WT), Nanos1-Ctrl-MO- and Nanos1-MO-injected embryos. For the rescue, Nanos1-MO embryos were injected with nanos1-mut RNA. (C) Summary of PGC loss at different stages after Nanos1 knockdown. P-values by one-way ANOVA. Error bars indicate s.e. (D) Hematoxylin and Eosin-stained sections of gonads from WT (stage 54), Nanos1-Ctrl-MO (stage 55), Nanos1-MO (stage 54) and rescued (stage 54) embryos, Arrows indicate germ cells. Note that nanos1 morphants were completely rescued by nanos1-mut RNA injection. Scale bars: 130 μm in A; 100 μm in B; 20 μm in D. |
