crip2 is expressed in the endothelium of the dorsal aorta. (A-F) In situ hybridization for crip2 in wild-type (n=35) (A), (n=15) (B), (n=20) (C) and in cripDM (n=35) (D), (n=15) (E), (n=20) (F) embryos at 26 hpf (A,B,D,E) and 36 hpf (C,F). Lateral views, anterior to the left (A,C,D,F). Transverse views, dorsal to the top (B,E). Scale bars: 100 µm. (G) Schematic depicts the zebrafish crip2 gene in black and the wild-type and predicted crip2 mutant protein products in blue. The five base pair deletion in exon 2 (red bar), generated through CRISPR mutagenesis, results in an early stop codon in exon 3, indicated by the red asterisk. The predicted protein lacks the second half of the first LIM binding domain. (H) Schematic depicts the zebrafish crip3 gene in black and the wild-type and predicted crip3 mutant protein products in blue. The 11 base pair deletion in exon 2 (red bar), generated through CRISPR mutagenesis, results in an early stop codon in exon 3, indicated by the red asterisk. The predicted protein lacks the second half of the first LIM binding domain. (I,J) Representative live images of wild-type (n=10) (I) and cripDM (n=10) (J) embryos at 52 hpf. Lateral views, anterior to the left. Scale bar: 250 µm. (K,L) Quantitative PCR analyses of crip2 (K) and crip3 (L) expression in wild-type and cripDM embryos at 26 hpf shows significant reduction in both genes, employing unpaired, two-tailed nonparametric t-tests (**P=0.0067 in K and ****P<0.0001 in L). Mean and standard error of each dataset are shown.
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