Fig. 1

Fig. 1

PCP signaling is required within FBMNs and in their r4 environment.

(A) Schematic showing a dorsal view of a 48 hours post fertilization (hpf) zebrafish hindbrain with anterior to the top. Facial Branchiomotor neurons (FBMNs) (green) migrate posteriorly from rhombomere (r) 4 to r6, leaving a trailing axon that exits from r4. The enhancer element islet-1 (isl1) CREST drives expression in branchiomotor neurons (green); the hoxb1a element drives expression in r4 (light blue); egr2b drives expression in r3 and r5 (yellow) and shh drives expression in the floorplate (purple). (B,C, G-L) Live or (H) fixed confocal images showing dorsal views of the hindbrain of 48 hpf zebrafish embryos with anterior to the top. Brackets mark rhombomere (r) position. Scale Bar: 50μm (B) Tg(isl1:GFP) expression in a wild type embryo at 48 hpf. (C) Tg(isl1:Dvl-DEP-GFP) embryo with unmigrated Dvl-DEP-GFP-expressing FBMNs in r4. (D,E) Dorsal view of E12.5 mouse hindbrains with FBMNs (magenta) labeled with anti-Isl1 antibody. Dotted lines indicate the length of facial motor nucleus. Scale Bar: 100μm (D) Migrating FBMNs in Vangl2^LoxP/LoxP control embryos. N = 6 embryos. (E) Blocked FBMNs in Vangl2^LoxP/LoxP;Isl1^Cre embryos. N = 9 embryos. (F) Quantitation of FBMN migration stream length in Vangl2^LoxP/LoxP control embryos and Vangl2^LoxP/LoxP;Isl1^Cre embryos. ***p = 0.0003. Significance was determined using an unpaired, two-tail t-test. (G-L) FBMNs (magenta) are either expressing Tg(isl1:mRFP)(G,I-L) or are stained with anti-Isl1 (H). (G,H) Tg(egr2b:KalTA4)-driven expression of Tg(UAS:Xdd1-GFP) (G) and Tg(UAS:Fzd3aΔC-GFP) (H), throughout r3 and r5 does not block FBMN migration. (I,J) Tg(hoxb1a:Gal4)-driven expression of Tg(UAS:Xdd1-GFP) (I) and Tg(UAS:Fzd3aΔC-GFP) (J), throughout r4 blocks FBMN migration out of r4. (K,L) Chimeric embryos with transplant conditions indicated as donor→ host. Cascade blue-dextran marks all donor-derived cells (blue) and Tg(isl-1:mRFP) marks all donor-derived FBMNs (magenta). (K) Wild type donor-derived FBMNs migrate normally in a non-transgenic control host. N = 37 embryos, 378 FBMNs. (L) Wild type donor-derived FBMNs fail to migrate out of r4 that is expressing Tg(UAS:Xdd1-GFP). N = 26 embryos, 190 FBMNs. Inset: same image without the magenta channel showing that donor-derived FBMNs (blue, circled) are not themselves expressing Xdd1-GFP (green). (M) Histograms indicate the percent of donor-derived FBMNs at 48 hpf that failed to migrate (r4), migrated partially (r5) or migrated fully (r6). Each histogram corresponds to the chimeric condition in the image to its left and numbers indicate the number of FBMNs represented in each bar.