Adult (four months) craniofacial phenotypes in the established cyp1b1-KO zebrafish line (F3 and F4). (A–L) Phenotypes of homozygous (-/-) cyp1b1-KO F3 zebrafish. (A) Lateral craniofacial shape was assessed by the L1/L2 ratio, where L1 (yellow line) is the dorsoventral head length centered in the pupil and L2 (red line) is the anteroposterior head length from the center of the pupil to the end of the lower jaw (white dashed line). (B) Jaw asymmetry was evaluated measuring the dorsal angle (da) formed between the longitudinal central body axis (yellow dashed line) and an imaginary line linking the supraoccipital axis and the joining point of the two Meckel’s cartilages (green line). (C,D) Ventral morphology of wild type-like head. (E–H) Phenotype 1 (Ph1) was characterized by jaw asymmetry, i.e., da > 5°, which is clearly visible in (F). (I–L) Phenotype 2 (Ph2) was identified by altered craniofacial shape defined as the L1/L2 ratio > 1.87, which is 15% higher than the mean value (1.63) in wildtype zebrafish. White arrowhead: altered quadrate bone. Red arrowhead: altered palatoquadrate bone. (M) Percentage of F3 craniofacial phenotypes. (N) Percentage of F4 craniofacial phenotypes. F3 cyp1b1-KO siblings with mutant or wild type-like phenotypes were inbred to obtain the F4 KO progeny (-/-). Wildtype (+/+) F3 progenitors were crossed in parallel as a control. The images in (A–L) are representative of the results observed in 33 homozygous (-/-) cyp1b1-KO F3 zebrafish. Scale bar = 2.5 mm.
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