The glomerular filtration barrier in the kidney is formed in part by a specialized intercellular junction known as the slit diaphragm, which connects adjacent actin-based foot processes of kidney epithelial cells (podocytes)1. Mutations affecting a number of slit diaphragm proteins, including nephrin (encoded by NPHS1)2, lead to renal disease owing to disruption of the filtration barrier and rearrangement of the actin cytoskeleton3, although the molecular basis for this is unclear. Here we show that nephrin selectively binds the Src homology 2 (SH2)/SH3 domain-containing Nck adaptor proteins4, which in turn control the podocyte cytoskeleton in vivo. The cytoplasmic tail of nephrin has multiple YDxV sites that form preferred binding motifs for the Nck SH2 domain once phosphorylated by Src-family kinases. We show that this Nck–nephrin interaction is required for nephrin-dependent actin reorganization. Selective deletion of Nck from podocytes of transgenic mice results in defects in the formation of foot processes and in congenital nephrotic syndrome. Together, these findings identify a physiological signalling pathway in which nephrin is linked through phosphotyrosine-based interactions to Nck adaptors, and thus to the underlying actin cytoskeleton in podocytes. Simple and widely expressed SH2/SH3 adaptor proteins can therefore direct the formation of a specialized cellular morphology in vivo.
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We thank T. Huber, G. Rivera, B. Mayer, P. Mundel, P. Soriano, M. Kirschner, K. Tryggvason and D. Hedley for providing reagents and technical advice, D. Holmyard for electron microscopy, and N. Woody for technical contributions. N.J. was supported by fellowships from the Canadian Institutes for Health Research (CIHR) and the National Cancer Institute of Canada (NCIC), with funds from the Terry Fox Run. This work was supported by grants from the CIHR (to S.S.-C.L., T.T., S.E.Q. and T.P.), the NCIC (to S.S.-C.L., S.E.Q. and T.P.) and the Kidney Foundation of Canada (to T.T.). S.S.-C.L. is an NCIC Scientist with funds from the Canadian Cancer Society. L.L. and T.T. hold scholarships from the Fonds de la Recherche en Santé du Québec. S.E.Q. holds a Tier 2 Canada Research Chair. T.P. is a Distinguished Investigator of the CIHR.
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Scientific Reports (2016)