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 ?-Cat?N activity in vitro. The indicated plasmid DNA (20?ng Wnt3a and 50?ng ?-Cat?N) was co-transfected with TOPFlash plasmid DNA into HEK293T cells and the luciferase activity was measured. (F) Gpx4b-C inhibits Wnt3a and ?-Cat?N 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 ?-cat?N) 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.