The Role of the Wnt/PCP Formin Daam1 in Renal Ciliogenesis

Abstract

16Kidneys are composed of numerous ciliated epithelial tubules called nephrons. Each nephron functions to reabsorb 17 nutrients and concentrate waste products into urine. Defects in primary cilia are associated with abnormal formation of 18 nephrons and cyst formation in a wide range of kidney disorders. Previous work in Xenopus laevis and zebrafish embryos 19 established that loss of components that make up the Wnt/PCP pathway, Daam1 and ArhGEF19 (wGEF) perturb kidney 20 tubulogenesis. Dishevelled, which activates both the canonical and non-canonical Wnt/PCP pathway, affect cilia 21 formation in multiciliated cells. In this study, we investigated the role of the noncanoncial Wnt/PCP components Daam1 22and ArhGEF19 (wGEF) in renal ciliogenesis utilizing polarized mammalian kidney epithelia cells (MDCKII and IMCD3) and 23Xenopus laevis embryonic kidney. We demonstrate that knockdown of Daam1 and ArhGEF19 in MDCKII and IMCD3 cells 24 leads to loss of cilia, and Daam1's effect on ciliogenesis is mediated by the formin-activity of Daam1. Moreover, Daam1 25 co-localizes with the ciliary transport protein IFT88. Interestingly, knocking down Daam1 in Xenopus kidney does not 26 lead to loss of cilia. This data suggests a new role for Daam1 in the formation of primary cilia. 27 catenin, the Wnt co-transcription factor, results in a reduction of cilia (Zhu et al., 2015). In mouse and Xenopus laevis a 39 protein called Chibby1, which functions to shuttle β-catenin out of the nucleus, localizes to the cilia and is required for 40 proper ciliogenesis and kidney development (Lee et al., 2014; Shi et al., 2014). Additionally, mutations in genes that 41 regulate cilia formation typically results in increased sensitivity to Wnt ligands, although the mechanism for this 42 phenomenon is heavily debated (Corbit et al., 2008). Moreover, Dishevelled, a component of both the canonical and 43 non-canonical Wnt pathway, is required for actin assembly and positioning of the basal bodies on the surface of the cell 44 during ciliogenesis in X. laevis multiciliated skin cells (Park et al., 2008). 45Actin filaments are important for proper ciliogenesis. In motile multiciliated cells, the cilia are connected by an actin 46 network (Werner et al., 2011). Although this actin network is not clearly visible around primary cilia, application of drugs 47