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Fig. 6 Fsta antagonizes the activity of Spaw and Activin by blocking binding to their receptors. (A) Schematic diagram of experimental setup for (B) WISH of lefty1 is used for a readout of Nodal/Activin activity. Immunostaining of EGFP shows grafted cells. Donor cells from embryos injected with EGFP mRNA or + fsta or truncated fsta mRNA (?5/?4/?TB) were grafted into host embryos that were overexpressed with spaw or inhbb. WT embryos were used as control. Numbers in the bottom right of panels indicate the number of embryos with the phenotype shown out of the total number of embryos examined. At least 20 embryos were examined in each experiment. (C) Interaction of Fsta?Flag, EGFP?Spaw, and HA?Tdgf1 in zebrafish embryos as assessed using competitive protein binding assay. (D) Interaction of EGFP?Spaw with its candidate receptors in zebrafish embryos as assessed using Co-IP. (E) Interaction of Fsta?Flag, EGFP?Spaw, and Acvr2ab?HA in zebrafish embryos as assessed using competitive protein binding assays. (F) Interaction of Fsta?Flag, EGFP?Spaw, and Acvr1bb?HA in zebrafish embryos as assessed using competitive protein binding assays. (Scale bar, 100 ?m.)