Fig 4

(A) capza1a^ex5 mutants harbour an insertion of 20 bp into exon 5 of capza1a, resulting in a frame shift. (B) The mutant allele capza1b^ex5 was generated by deletion of 23 bp and insertion of 1 bp into exon 5 of capza1b, causing a frame shift. (C) At 3 dpf, in situ hybridization showed that the trunk expression of capza1a found in siblings was strikingly reduced in capza1a^ex5 homozygotes. (D) At 3 dpf, whole-mount in situ hybridization showed that the expression of capza1b was severely reduced in capza1b^ex5 homozygotes compared to siblings. (E) At 3 dpf, the birefringence of single capza1a^ex5/+ and capza1b^ex5/+ heterozygotes as well as single capza1a^ex5/ex5 homozygotes was comparable to WT siblings (capza1a^+/+;capza1b^+/+, capza1a^ex5/+;capza1b^+/+, capza1a^+/+;capza1b^ex5/+ and capza1a^ex5/ex5;capza1b^+/+ all were 100 ± 1%). However, single capza1b^ex5/ex5 homozygotes were significantly reduced in their birefringence and larvae that were additionally heterozygous for capza1a^ex5/ex5 were significantly reduced further (capza1a^+/+;capza1b^ex5/ex5 were 81 ± 1% and capza1a^ex5/+;capza1b^ex5/ex5 were 75 ± 1%). In addition, compound heterozygotes capza1a^ex5/+;capza1b^ex5/ex5 and capza1a^ex5/ex5 homozygotes that were capza1b^ex5/+ heterozygous were also significantly reduced in their birefringence (capza1a^ex5/+;capza1b^ex5/+ were 92 ± 2% and capza1a^ex5/ex5;capza1b^ex5/+ were 86 ± 1%). Data are mean ± SEM; * P < 0.05, ** P < 0.01 and *** P < 0.001 calculated by one-way ANOVA with post hoc Tukey’s test; n = 5 clutches. Scale bar sizes are 300 μm.