The role of nephronophtisis proteins in motile cilia and renal tubules of Xenopus laevis
Final Report Abstract
The aim of the funded project was to elucidate the function of nephronophthisis related proteins in ciliogenesis and to determine the relationship of planar cell polarity signaling to tubulogenesis in Xenopus. Because little was known about the dynamics of tubule formation in embryogenesis of Xenopus, we performed extensive live cell imaging studies on tadpoles that expressed fluorescent labels in the developing renal system. We found extensive cell movement and intercalation as the pronephric tubules extended and elongated. The direction of tissue remodeling was correlated with the occurrence and positioning of multi-cellular rosettes. Rosette formation and tubule elongation were dependent on non-muscle myosin and planar cell polarity signaling. Interestingly, we detected that the collecting ducts of embryonic mice showed a similar shift towards lower order rosettes in a PCP-deficient background, which was is agreement with the observed phenotype in Xenopus. Therefore, rosette-based convergent extension movements guided by planar cell polarity signaling may constitute an evolutionarily conserved mechanism of renal tubule elongation. In a second project, we identified a novel nephronophthisis gene (ANKS6), the protein product of which was initially identified as a binding partner of NEK8 (NPHP9). ANKS6 was able to bridge the physical interaction of NEK8 and INVS (NPHP2) or NPHP3. All of these proteins localize to a distinct, proximal portion of the ciliary axoneme, previously described as Inv-compartment. Depletion experiments in Xenopus and zebrafish confirmed a ciliopathy typical phenotype of deficient pronephros development, disruption in left-right laterality and hydrocephalus. In collaboration with an international team of human geneticists, 6 families affected by nephronophthisis were identified that had a causal mutation in ANKS6. Thus, ANKS6 was classified by OMIM as NPHP16 and is now included in the genetic testing of unsolved cases of nephronophtisis. Further proteomic analysis found multiple protein interactors that were functionally follow up in subsequent projects. In conclusion, this data added to the growing body of knowledge of how multi-protein complexes at the primary cilium contribute to genetic kidney disease.
Publications
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(2012) Vertebrate kidney tubules elongate using a planar cell polaritydependent, rosette-based mechanism of convergent extension. Nature genetics 44, 1382-1387
Lienkamp SS, Liu K, Karner CM, Carroll TJ, Ronneberger O, Wallingford JB, and Walz G
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(2013) ANKS6 is a central component of a nephronophthisis module linking NEK8 to INVS and NPHP3. Nature genetics 45, 951-956
Hoff S, Halbritter J, Epting D, Frank V, Nguyen TM, van Reeuwijk J, Boehlke C, Schell C, Yasunaga T, Helmstadter M, Mergen M, Filhol E, Boldt K, Horn N, Ueffing M, Otto EA, Eisenberger T, Elting MW, van Wijk JA, Bockenhauer D, Sebire NJ, Rittig S, Vyberg M, Ring T, Pohl M, Pape L, Neuhaus TJ, Elshakhs NA, Koon SJ, Harris PC, Grahammer F, Huber TB, Kuehn EW, Kramer-Zucker A, Bolz HJ, Roepman R, Saunier S, Walz G, Hildebrandt F, Bergmann C, and Lienkamp SS
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(2014) Casein kinase 1 alpha phosphorylates the Wnt regulator Jade-1 and modulates its activity. The Journal of biological chemistry 289, 26344-26356
Borgal L, Rinschen MM, Dafinger C, Hoff S, Reinert MJ, Lamkemeyer T, Lienkamp SS, Benzing T, and Schermer B
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(2014) Interaction with the Bardet-Biedl gene product TRIM32/BBS11 modifies the half-life and localization of Glis2/NPHP7. The Journal of biological chemistry 289, 8390-8401
Ramachandran H, Schafer T, Kim Y, Herfurth K, Hoff S, Lienkamp SS, Kramer- Zucker A, and Walz G
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(2015) The Rac1 regulator ELMO controls basal body migration and docking in multiciliated cells through interaction with Ezrin. Development 142, 1553
Epting D, Slanchev K, Boehlke C, Hoff S, Loges NT, Yasunaga T, Indorf L, Nestel S, Lienkamp SS, Omran H, Kuehn EW, Ronneberger O, Walz G, and Kramer-Zucker A
