Reciprocal inhibition of Wnt signaling pathways pattern the interconnection of epithelial tubules in the regenerating zebrafish kidney

In the adult zebrafish kidney, nephrogenesis occurs as a regenerative response to injury and provides a model to explore cell signaling pathways required for nephron formation and engraftment. Differentiating kidney tubules interconnect with collecting system epithelia to generate a pathway for fluid excretion. We show that canonical Wnt signaling induces a mesenchymal, invasive cell phenotype and is required, along with Src kinase and Rac1, to generate basal cell protrusions on new nephrons. The Wnt ligands wnt9b and wnt4 are both required for new nephron formation after injury. Mutation in wnt4 and wnt9b, or treatment with the canonical Wnt inhibitor IWR1 blocks the formation of basal protrusions in forming nephrons. Mutation in the Wnt receptor frizzled9b reveals a fusion-associated non-canonical Wnt pathway that acts to (1) restrict canonical Wnt gene expression, (2) drive Rho kinase-dependent apical constriction of epithelial cells and (3) position basal protrusions to generate orthogonal tubule lumenal connections. Mutation in wnt9b phenocopies fzd9b mutants, indicating that a single ligand can induce canonical and non-canonical Wnt signaling in the same cells to orient and drive tubule interconnection in the regenerating zebrafish kidney.