Deletion of Notch Signaling in the Developing Mouse Kidneys Results in Expanded Expression of the Small GTPase Arf6 and ZO-1, Reduced Expression of E-Cadherin, and Collecting Duct Dilations

Background: Notch signaling regulates proximal/distal patterning of the nephron in the mammalian kidney. There are four Notch receptors and five ligands, and during kidney development, Notch receptors are activated by binding to ligands on neighboring cells. This results in a series of proteolytic cleavages that lead to the expression of Notch target genes. Deletion of Notch signaling in the developing collecting duct results in collecting duct dilations and hydronephrosis. While cells lining the medullary cavity formed from hydronephrosis are highly proliferative, cells lining the cortical dilations do not label with markers of proliferation. Methods: To determine whether the dilated collecting ducts in Notch mutant kidneys, called RBPJ/kCD, resulted from changes in cell-cell interactions, we evaluated the expression of the ADP-ribosylation factor GTPase Arf6, receptor tyrosine kinase EGFR and phosphorylated EGFR, and cell-cell adhesion proteins ZO-1, E-cadherin, and N-cadherin. To further determine effects of reduced Notch signaling, we also assessed expression of alpha smooth muscle actin. Results: Newborn RBPJ/kCD mice exhibited medullary and cortical dilations. Staining with DBA lectin and cytokeratin confirmed the collecting duct origin of the dilations. Postnatal day 14 (P14), RBPJ/kCDmice exhibited hydronephrosis and cortical dilations. Cells lining the dilated collecting ducts exhibited expanded expression of the ADP-ribosylation factor GTPase Arf6 and cell-cell adhesion protein ZO-1, which are normally restricted to the apical membrane. EGFR, phosphorylated EGFR, and alpha smooth muscle actin expression were increased RBPJ/kCD mice. There was a reduction in the basolateral expression of E-cadherin in the collecting ducts of RBPJ/kCD mice. No differences in N-cadherin expression were observed. Conclusion: Taken together, our results suggest that Notch signaling is required for normal kidney collecting duct development, and that reduced Notch signaling may disrupt cell-cell interactions and EGFR signaling leading to hydronephrosis.