Fig. 5

GPX4/Gpx4b inhibits Wnt activity at the Tcf level. (A,B) Gpx4b inhibits the action of indicated Wnt activators in vivo. Representative images of embryos injected with 680 pg gfp mRNA, mRNA of each indicated Wnt activator (20 pg wnt3a, 50 pg β-catΔN and 80 pg vp16-Tcf3ΔN) and mRNA of each indicated Wnt activator plus 600 pg gpx4b mRNA at 12.5 hpf are shown in A. Quantitative results are shown in B. The frequency of embryos with the indicated phenotypes is shown in A. (C) Gpx4b inhibits Wnt activity induced by each indicated Wnt activator in vivo. Embryos were injected with TOPFlash reporter DNA with the indicated mRNAs (20 pg wnt3a, 50 pg β-catΔN, and 80 pg vp16-Tcf3ΔN). (D) Maternal loss of Gpx4b synergistically elevated β-catΔN- and vp16-tcf3ΔN-induced expression of boz, chd and sqt at 4.3 hpf. One-cell stage wild-type and maternal mutant embryos were injected with gfp (40 pg), β-catΔN (25 pg) or vp16-tcf3ΔN (40 pg) mRNA. The injected embryos were raised to 4.3 hpf and the expression levels of the indicated target genes were measured by qRT-PCR. (E,F) Knockdown of GPX4 synergistically enhances Wnt3a- and VP16-Tcf3ΔN-induced Wnt reporter activity. The indicated plasmid DNA (5 ng) was co-transfected with TOPFlash plasmid DNA into control or GPX4-knockdown HEK293T cells and the luciferase activity was measured. Results are from three independent experiments. Values are means±s.e.m. (n=3). *P<0.05; **P<0.01; ***P<0.001. Unpaired t-test, two-tailed. Scale bars: 200 µm.