Fig. 1

cep104 Knockdown in Zebrafish Embryos Leads to Ciliopathy Phenotypes

(A–C) 48 hpf morphant zebrafish display mild and severe pericardial edema and cardiac defects (^∗) following cep104 knockdown and additional phenotypes in severe morphants of mild tail curvature and microopthalmia, with a quantified reduction in area expressed as a ratio to control embryos of 0.45 (p < 0.0001, unpaired t test, n = 39 versus 28 control).

(D) Percentage of zebrafish displaying phenotypes following injection of cep104 splice MO and translation blocking morpholino cep104 ATG MO alone or in combination (control n = 98, cep104splice MO n = 166, cep104 ATG MO n = 95, cep104 splice MO + cep104ATG MO n = 77).

(E) Western blotting (WB) of cep104 at 48 hpf in zebrafish uninjected and injected with cep104 ATG MO and cep104 splice MO.

(F) IFM of cilia and cell junctions (a-acetylated tubulin, red) in Kupffer’s vesicle (KV; atypical protein kinase C [aPKC], green) at the 10-somite stage in control and cep104 knockdown embryos.

(G) Dot plots of the length of cilia in KV in control, cep104 splice MO knockdown, and cep104 splice MO and CEP104 mRNA co-injected zebrafish embryos (ANOVA with Tukey post hoc test, ^∗p < 0.05).

(H) 48 hpf cmlc2:GFP zebrafish treated with cep104 splice MO show changes to heart looping at 48 hpf, which is rescued by co-injection with CEP104 mRNA.

(I) Percentage of embryos displaying heart looping phenotypes following injection of cep104 splice MO and co-injection with CEP104 mRNA (^∗∗∗p < 0.0001, ^∗p = 0.0208, chi-square test of independence; control n = 186; cep104 splicing MO n = 132; cep104splicing MO + CEP104 mRNA n = 130).

(J and K) cep104 knockdown in 48 hpf islet-1:GFP transgenic fish leads to cranial nerve defects, rescued by co-injection with CEP104mRNA. Co-injection with CEP104 mRNA produces a partial rescue of phenotypes (^∗∗∗p < 0.0001, ^∗∗p = 0.0010, chi-square test of independence; control n = 200; cep104 splicing MO n = 80; cep104splicing MO + CEP104 mRNA n = 120).