Fig. 3

Generation of zebrafish ahr1a and ahr1b mutants.

(A, B) Depiction of genomic locus for ahr1a (A) and ahr1b (B). TALEN pairs were designed targeting the bHLH domain in exon 2 of both genes. Genomic lesions in the TALEN spacer indicated in alignment to WT sequence, and cartoons show predicted protein products. For ahr1b, alleles mu133-mu135 were generated in a WT genetic background, while mu136-mu145 were made in an ahr2^hu3335 homozygous mutant background. (C) Brightfield images of WT, ahr1a -/-, ahr1b,2 -/- and triple AHR mutant embryos at 5 dpf. No gross morphological defects in AHR mutants are apparent. Boxes indicate region of imaging for different vascular beds in D-I. Numbers correspond to “number of embryos with inflated swim bladders/total number of embryos of that genotype”. Scale bar is 1000 um. (D-I) Maximum intensity projections of confocal z-stacks comparing the trunk (D, E), brain (F, G) and liver (H, I) vascular beds in ahr1a -/- and triple AHR mutant zebrafish embryos at 5 dpf. Vascular patterning appears normal in both mutants compared to WT. Yellow arrowheads point to forming intercostal vessels in the trunk. Orange brackets mark midbrain vessels, yellow brackets mark hindbrain vessels. Red arrows show the liver. Numbers indicate the “number of embryos of a particular genotype with vascular characteristics as in image/total numbers of that genotype analyzed.” Scale bar is 100 um. Abbreviations—AHR: aryl hydrocarbon receptor, bHLH: basic helix-loop-helix, hpf: hours post fertilization, TALEN: transcription activator-like effector nuclease, WT: wildtype.