Rac1 is required for ciliogenesis in zebrafish and Xenopus. (A-B′′) Expression silencing of Rac1l using TB-MO rac1l (0.5 ng) results in pronephric cyst formation (stars in B′ and B′′) as compared with Co-MO (0.5ng) morphants (A′), as shown in a dorsal view with anterior to the left of a Tg(wt1b:EGFP) zebrafish embryo (A′,B′) and in a histological transverse section (B") of a 48 hpf embryo. (A,B) Embryos are shown in bright-field lateral view, with anterior to the left. (C) Quantification of pronephric cyst formation in 48 hpf zebrafish embryos injected with Co-MO (4.5 ng), TB-MO rac1 (4 ng), TB-MO rac1l (0.5 ng), TB-MO rac1 (4 ng)/rac1l (0.5 ng) and TB-MO rac1l (0.5 ng)+ rac1l mRNA (20pg). There was significant prevention of cyst formation by co-injection with rac1l mRNA (*Pd0.05). (D) Quantification of laterality defects by cmlc2 in situ hybridisation revealed impaired heart looping in 48 hpf zebrafish embryos injected with TB-MO rac1 (4ng), TB-MO rac1l (0.5 ng), and TB-MO rac1 (4 ng)/rac1l (0.5 ng) compared with Co-MO (4.5 ng). (C,D) The number of individual embryos analysed is indicated above each bar. (E-E′′) Analysis of electron micrographs revealed basal body docking defects (arrows in E′,E′′) on sections of 48 hpf zebrafish embryos injected with TB-MO rac1 (4 ng)/rac1l (0.5 ng). (F-F′′′) Xenopus embryos injected with Co-MO (40 ng) and centrin-GFP mRNA showed normal basal body docking and basal body distribution at stage 32; embryos were colabelled with phalloidin 568 (magenta). (G-H′′′) Xenopus embryos (stage 32) injected with TB-MO rac1 (40 ng) exhibited irregular basal body docking and distribution, ‘beads on a string’ pattern in H. Scale bars: 100 μm in A,B; 20 μm in A′,B′ ; 50 μm in B′′ ; 1 μm in E-E′′ ; 5 μm in H′′′.
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