Down-regulation of WNT/β-catenin signalling activity in <i>Dullard</i>^−/− embryos.

<p>(<b>A–B′′</b>) Curtailed expression of <i>Axin2</i> in the (<b>B–B′′</b>) E7.5 <i>Dullard</i>^−/− embryo compared with that in the (<b>A–A′′</b>) <i>Dullard</i>^+/− embryo. (<b>A, B</b>, bright-field images; <b>A′, B′</b>, Axin2 immunostaining; <b>A′′, B′′</b>, merged images). (<b>C–F′′′</b>) Expression of activated β-catenin (ABC) visualized by immunostaining and confocal microscopy of (<b>C–D′′′</b>) E7.5 no bud-stage <i>Dullard</i>^+/− embryos and (<b>E–F′′′</b>) E7.5 <i>Dullard</i>^−/− embryos. ABC expression in the posterior germ layer of the embryo, where Ifitm3-positive PGCs were localized, was weaker in the <i>Dullard^−/−</i> embryo that also lacked Ifitm3-postive PGCs. (<b>C, E</b>) Bright-field image; (<b>C′, E′</b>) ABC immunostaining; (<b>D–D′′′, F–F′′′</b>) magnified views of the boxed areas in panels <b>C′</b> and <b>E′</b>, respectively; (<b>D′′, F′′</b>) merged ABC and Iftm3 images; (<b>D′′′, F′′′</b>) merged images with DAPI nuclear staining; ABC immunostaining (green); Ifitm3 immunostaining (red), Scale bar = 100 µm (A–C′, E–E′) and 20 µm (<b>D–D′′′, F–F′′′</b>). (<b>G</b>) Western blot analysis of E7.75 wild-type (+/+) and <i>Dullard^+/−</i> (+/−) embryos showing reduced amounts of the active form of β-catenin in <i>Dullard^−/−</i> (−/−) embryos, but total β-catenin content was unchanged.