Fig. S1. Phenotypes of arl13b morphants. (A) Injected at modest dosage, arl13 morphants show similar phenotypes to sco^hi459 mutants. The morphants can be rescued by co-injection of arl13b mRNA. c-mo, embryos injected with a standard control morpholino; sco-mo, embryos injected with the arl13b morpholino; sco-mo+mRNA, embryos co-injected with arl13b morpholino and mRNA. (B) Injected at higher dosage, arl13 morphants show more severe phenotypes at 1 dpf. mph, morphant; wt, wild type. (C) A shield-stage embryo injected with FM1-43 in dorsal view. (D,E) Side views of injected embryos at the tailbud stage. Arrows point to the tip of the head and the tail. mo+sco, embryos co-injected with sco morpholino and mRNA (D); mo+gfp, embryos co-injected with sco morpholino and eGFP mRNA (E).

Fig. S2. Point mutations in the small GTPase domain, and coiled-coil deletion disrupt cilia rescue activity. (A,B) Cilia in the pronephric duct (arrow) at 50 hpf as shown by anti-acetylated tubulin immunostaining in whole-mount embryos; side views at low magnification. Mut, sco^hi459 mutant sibling; wt, wild type. (C) 265.5 pg mRNA encoding alr13b alleles translationally fused to gfp were injected into hi459^+/−×hi459^+/− embryos at the one- to four-cell stage. Embryos were then scored at 2 dpf for body curvature, and stained with antibody against acetylated tubulin to evaluate the pronephric cilia phenotype. P-value is calculated from a chi-squared test with 1 degree of freedom. Results from a representative experiment are shown.

Movie 1. Ciliary motility in a wild-type sibling at 5 dpf. Images were taken at 200 frames per second and are played at 6 frames per second. Arrow indicates beating cilia. Scale bar: 15 m.

Movie 2. Ciliary motility in a sco^hi459 mutant embryo at 5 dpf. Images were taken at 200 frames per second and are played at 6 frames per second. Arrow indicates beating cilia. Scale bar: 15 m.