Motile and Sensory Cilia Function in Zebrafish

Abstract

Advances in developmental genetics and human disease gene cloning have highlighted the essential roles played by cilia in developmental cell fate decisions, left–right asymmetry, and the pathology of human congenital disorders. Cilia-driven fluid flow in the embryonic node highlights the role of motile cilia in the generation of mechanical signals in development. Sensory cilia in photoreceptors and olfactory neurons highlight the role of cilia in phototransduction and G-protein coupled receptor signaling. Using a zebrafish knockdown screen of cilia candidate genes as an in vivo screening platform, we identified c21orf59, ccdc65, and c15orf26 as critical for cilia motility and generation of proper left-right asymmetry. c21orf59 and c15orf26 knockdown in zebrafish blocked outer dynein arm assembly, while ccdc65 knockdown altered cilia beat pattern. To study sensory cilia function we generated a new cilia assay based on calcium biosensor (GCaMP5) expression in zebrafish ciliated olfactory neurons (OSNs). Ciliated OSNs are activated by bile acid olfactory receptors on outward facing terminal sensory cilia. We found that ciliated OSNs responded with a robust fluorescence signal to specific odorants and that the cilia mutant fish oval, (ift88-/-) and ift172 knockdown larvae showed severely shortened OSN cilia and markedly reduced responses to bile acids while other odorant responses were unaffected. In summary, the zebrafish provides a robust platform for cilia gene discovery, in vivo functional assays of cilia mutant allele strength, and mechanisms of sensory cilia assembly and function. Supported by NIH grant DK053093