(a) Embryos expressing ACVR1 HA-tagged constructs were immunostained for HA at the shield stage (early gastrula); ACVR1 domain schematics of the constructs are above each set of corresponding images. Nuclei (yellow), βcatenin (cyan), ACVR1-HA (magenta). Scale bars = 40 μm. ΔAcvr1 constructs lack residues 35–100, which contain the cysteine-rich ligand binding motifs. Uninjected (N = 6) and embryos injected with: Acvr1 (N = 6), ΔAcvr1 (N = 5), Acvr1-R206H (N = 9), ΔAcvr1-R206H (N = 4), Acvr1-G328R (N = 5), and ΔAcvr1-G328R (N = 5). (b) BMP signaling dose-dependent 12–30 hpf zebrafish embryo phenotypes: severe dorsalization (C5-C4, dark blue), mild dorsalization (C3-C1, light blue), wild-type development (WT, violet), mild ventralization (V1-V3, light orange), and severe ventralization (V4-V5, dark orange). (c) Bubble plot circle sizes correspond to the percent of total embryos within a condition that are in a particular phenotypic category. Examples are shown for: 100%, 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20% and 10%. (d) Representative phenotypes of wild-type embryos (evaluated at 12 and 30 hpf) and embryos injected with acvr1l MO (evaluated at 12 hpf; none survive to 30 hpf). (e–g) DV phenotypes of 12–30 hpf embryos injected with Acvr1 or ΔAcvr1 mRNA alone or together with acvr1l MO. Data are from four pooled experiments. (e) Acvr1 or ΔAcvr1 injected embryos. Columns: 1, N = 240; 2, N = 121; 3, N = 140; 4, N = 153; 5, N = 96; 6, N = 118. (e’) Representative 12 or 30 hpf phenotypes. (f) Acvr1-R206H or ΔAcvr1-R206H injected embryos. Columns: 1, N = 240; 2, N = 70; 3, N = 84; 4, N = 153; 5, N = 73; 6, N = 91. (f’) Representative 30 hpf phenotypes. (g) Acvr1-G328R or ΔAcvr1-G328R injected embryos. Columns: 1, N = 240; 2, N = 88; 3, N = 79; 4, N = 153; 5, N = 78; 6, N = 71. (g’) Representative 30 hpf phenotypes.
|
