Fig. 6

Sec is dispensable for Wnt inhibition. (A) Gpx4b^WT and Gpx4b^U67C inhibit endogenous Wnt activity. The indicated plasmid DNA (200 ng) was co-transfected with TOPFlash plasmid DNA into HEK293T cells. (B) Classification of ventralized phenotypes at 24 hpf caused by forced expression of 600 pg gpx4b^U67C or GPX4^U73C mRNA. (C) Percentages of embryos in each category as shown in B. Results are from three independent experiments and the total embryo numbers are given at the top. (D) Gpx4b-C inhibits endogenous Wnt activity. The indicated plasmid DNA (600 ng) was co-transfected with TOPFlash plasmid DNA into HEK293T cells and the luciferase activity was measured. (E) Gpx4b-C inhibits Wnt3a and β-CatΔN activity in vitro. The indicated plasmid DNA (20 ng Wnt3a and 50 ng β-CatΔN) was co-transfected with TOPFlash plasmid DNA into HEK293T cells and the luciferase activity was measured. (F) Gpx4b-C inhibits Wnt3a and β-CatΔN action in vivo. Quantitative results are shown as in Fig. 5B. Embryos injected with 650 pg gfp mRNA, mRNA of each indicated Wnt activator (20 pg wnt3a and 50 pg β-catΔN) and mRNA of each indicated Wnt activator plus 600 pg gpx4b-c mRNA at 12.5 hpf. The total embryo numbers from three independent experiments are shown at the top of each bar. Values are mean±s.e.m. (n=3). *P<0.05; ***P<0.001. Unpaired t-test, two-tailed. Scale bar: 200 µm.