Kidney formation requires the coordinated growth of multiple cell types including the collecting ducts, nephrons, vasculature and interstitium. There is a long-held belief that interactions between progenitors of the collecting ducts and nephrons are primarily responsible for kidney development. However, over the last several years, it has become increasingly clear that multiple aspects of kidney development require signaling from the interstitium. How the interstitium orchestrates these various roles is poorly understood. Here, we show that during development the interstitium is a highly heterogeneous patterned population of cells that occupies distinct positions correlated to the adjacent parenchyma. Our analysis indicates that the heterogeneity is not a mere reflection of different stages in a linear developmental trajectory but instead represents several novel differentiated cell states. Further, we find that β-catenin has a cell autonomous role in the development of a medullary subset of the interstitium and that this non-autonomously affects the development of the adjacent epithelia. These findings suggest the intriguing possibility that the different interstitial subtypes may create microenvironments that play unique roles in development of the adjacent epithelia and endothelia.

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statement: This study describes a novel function for TGFb signaling in the 15 developing renal interstitium. Mice with Foxd1-Cre-mediated deletion of Smad4 have interstitial 16 expansion and activated Wnt signaling. 17 18 McCarthy et al. preprint uploaded to Bioarxiv 04.11.20 ABSTRACT 1Expansion of interstitial cells in the adult kidney is a hallmark of chronic disease, whereas their 2 proliferation during fetal development is necessary for organ formation. An intriguing difference 3 between adult and neonatal kidneys is that the neonatal kidney has the capacity to control 4 interstitial cell proliferation when the target number has been reached. In this study, we define 5 the consequences of inactivating the TGFb/Smad response in the interstitial cell lineage. We find 6 that pathway inactivation through loss of Smad4 leads to over-proliferation of interstitial cells 7 regionally in the kidney medulla. Genetic and molecular interaction studies showed that Smad3/4 8 participates in the Wnt/b-catenin signaling pathway, which is responsible for promoting 9 proliferation of interstitial cells. Specifically, Smad4 is required for the expression of the Wnt 10 feedback inhibitor Apcdd1, and based on these findings we propose a model for interstitial cell 11 proliferation control in which the Wnt/b-catenin proliferative signal is attenuated by TGFb/Smad 12 signaling to ensure that proliferation ceases when the target number of interstitial cells has been 13 reached in the neonatal medulla.