Characterisierung der embryonalen Nierenentwicklung in Danio rerio und Eine Funktionelle Analyse von Arl13b/ Scorpion waehrend der Embryonalentwicklung in Danio rerio

Joubert syndrome and Joubert syndrome and related disorders are human diseases that are characterized by brain stem and cerebellar defects but can also be accompanied by polycystic kidney disease, retinitis pigmentosa, situs inversus among other defects. It is crucial to understand the normal kidney development in order to understand the cyst pathogenesis, however not much is known about embryonic kidney development in danio rerio. The present study aimed to elucidate the earliest events during embryonic kidney formation in Danio rerio. We show for the first time that cells of the intermediate mesoderm aquire apico-basolateral polarity, undergo cavitatation and exhibit oriented cilia concurrently around the 12 somite stage and are completed at the 14 somite stage. Furthermore the intermediate mesoderm undergoes extensive elongation and narrowing processes which are not due to proliferation and apoptosis but might be governed by the Planar cell polarity pathway. Arl13b/ Scorpion, a small GTPase, localizes to the cilium in various organs and plays a crucial role for ciliogenesis in the KV, retinal photoreceptor layer and the pronephric duct. As a conseqeunce, the Arl13b mutant or morphant embryos show laterality defects, photoreceptor degeneration due to apoptosis in the outer segment of the photoreceptor layer, duct dilation in the pronephros and kidney cyst formation. Performing careful histological analysis in scorpionhi459 mutant embryos, we could show that hyperproliferation and/ or defects in cell rearrangements occur as a consequence of dilation, and are not causative. In addition, scorpionhi459 mutant embryos extend cilia tufts from multicilliated cells which present a defective basal body alignment that is goverened by PCP but cilia on single ciliated cells can not be detected. Remaning cilia have a normal axonemal ultrastructure as shown by electron microscopy, but exhibit a defective beating pattern. The apicao-basolaterael polarity is not perturbed in mutant pronephric ducts. A deletion analysis of Arl13b/ Sco protein allowed us to study the roles of the truncation proteins in regards to cilia localization and the capability to rescue the mutant phenotype. The GTPase domain, coiled coil region and part of the C-terminus (C-terminal to amino acid 308) were necessary for proper ciliary targeting. We found a close correlation for cilia localization being necessary for the protein being able to rescue the mutant phenotype. In addition, despite the GTPase activity playing no role in trafficking the protein to the cilium, it was crucial for the rescue capability of the protein