Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Nck adaptor proteins link nephrin to the actin cytoskeleton of kidney podocytes

Nature volume 440, pages 818–823 (2006)Cite this article

Abstract

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.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: The kidney slit diaphragm protein nephrin has multiple YDxV motifs that bind the Nck SH2 domain.
Figure 2: Identification of a phosphotyrosine-dependent interaction between nephrin and the Nck SH2/SH3 adaptor.
Figure 3: Glomerulosclerosis and foot process fusion in mice lacking Nck expression in podocytes.
Figure 4: Nck recruitment to phosphorylated nephrin facilitates localized actin reorganization.

Similar content being viewed by others

References

  1. Pavenstadt, H., Kriz, W. & Kretzler, M. Cell biology of the glomerular podocyte. Physiol. Rev. 83, 253–307 (2003)

    Article  CAS  Google Scholar 

  2. Kestila, M. et al. Positionally cloned gene for a novel glomerular protein—nephrin—is mutated in congenital nephrotic syndrome. Mol. Cell 1, 575–582 (1998)

    Article  CAS  Google Scholar 

  3. Oh, J., Reiser, J. & Mundel, P. Dynamic (re)organization of the podocyte actin cytoskeleton in the nephrotic syndrome. Pediatr. Nephrol. 19, 130–137 (2004)

    Article  Google Scholar 

  4. Buday, L., Wunderlich, L. & Tamas, P. The Nck family of adapter proteins: regulators of actin cytoskeleton. Cell. Signal. 14, 723–731 (2002)

    Article  CAS  Google Scholar 

  5. Ruotsalainen, V. et al. Nephrin is specifically located at the slit diaphragm of glomerular podocytes. Proc. Natl Acad. Sci. USA 96, 7962–7967 (1999)

    Article  ADS  CAS  Google Scholar 

  6. Benzing, T. Signaling at the slit diaphragm. J. Am. Soc. Nephrol. 15, 1382–1391 (2004)

    Article  Google Scholar 

  7. Shih, N. Y. et al. Congenital nephrotic syndrome in mice lacking CD2-associated protein. Science 286, 312–315 (1999)

    Article  CAS  Google Scholar 

  8. Roselli, S. et al. Early glomerular filtration defect and severe renal disease in podocin-deficient mice. Mol. Cell. Biol. 24, 550–560 (2004)

    Article  CAS  Google Scholar 

  9. Lehtonen, S. et al. Cell junction-associated proteins IQGAP1, MAGI-2, CASK, spectrins, and α-actinin are components of the nephrin multiprotein complex. Proc. Natl Acad. Sci. USA 102, 9814–9819 (2005)

    Article  ADS  CAS  Google Scholar 

  10. Verma, R. et al. Fyn binds to and phosphorylates the kidney slit diaphragm component Nephrin. J. Biol. Chem. 278, 20716–20723 (2003)

    Article  CAS  Google Scholar 

  11. Li, H., Lemay, S., Aoudjit, L., Kawachi, H. & Takano, T. SRC-family kinase Fyn phosphorylates the cytoplasmic domain of nephrin and modulates its interaction with podocin. J. Am. Soc. Nephrol. 15, 3006–3015 (2004)

    Article  Google Scholar 

  12. Lahdenpera, J. et al. Clustering-induced tyrosine phosphorylation of nephrin by Src family kinases. Kidney Int. 64, 404–413 (2003)

    Article  CAS  Google Scholar 

  13. Yu, C. C., Yen, T. S., Lowell, C. A. & DeFranco, A. L. Lupus-like kidney disease in mice deficient in the Src family tyrosine kinases Lyn and Fyn. Curr. Biol. 11, 34–38 (2001)

    Article  CAS  Google Scholar 

  14. Putaala, H., Sainio, K., Sariola, H. & Tryggvason, K. Primary structure of mouse and rat nephrin cDNA and structure and expression of the mouse gene. J. Am. Soc. Nephrol. 11, 991–1001 (2000)

    CAS  PubMed  Google Scholar 

  15. Huber, T. B. et al. Nephrin and CD2AP associate with phosphoinositide 3-OH kinase and stimulate AKT-dependent signaling. Mol. Cell. Biol. 23, 4917–4928 (2003)

    Article  CAS  Google Scholar 

  16. Chen, M. et al. Identification of Nck family genes, chromosomal localization, expression, and signaling specificity. J. Biol. Chem. 273, 25171–25178 (1998)

    Article  CAS  Google Scholar 

  17. Gruenheid, S. et al. Enteropathogenic E. coli Tir binds Nck to initiate actin pedestal formation in host cells. Nature Cell Biol. 3, 856–859 (2001)

    Article  CAS  Google Scholar 

  18. Lu, W., Katz, S., Gupta, R. & Mayer, B. J. Activation of Pak by membrane localization mediated by an SH3 domain from the adaptor protein Nck. Curr. Biol. 7, 85–94 (1997)

    Article  CAS  Google Scholar 

  19. Rohatgi, R., Nollau, P., Ho, H. Y., Kirschner, M. W. & Mayer, B. J. Nck and phosphatidylinositol 4,5-bisphosphate synergistically activate actin polymerization through the N-WASP-Arp2/3 pathway. J. Biol. Chem. 276, 26448–26452 (2001)

    Article  CAS  Google Scholar 

  20. Bladt, F. et al. The murine Nck SH2/SH3 adaptors are important for the development of mesoderm-derived embryonic structures and for regulating the cellular actin network. Mol. Cell. Biol. 23, 4586–4597 (2003)

    Article  CAS  Google Scholar 

  21. Rivera, G. M., Briceno, C. A., Takeshima, F., Snapper, S. B. & Mayer, B. J. Inducible clustering of membrane-targeted SH3 domains of the adaptor protein Nck triggers localized actin polymerization. Curr. Biol. 14, 11–22 (2004)

    Article  CAS  Google Scholar 

  22. Putaala, H., Soininen, R., Kilpelainen, P., Wartiovaara, J. & Tryggvason, K. The murine nephrin gene is specifically expressed in kidney, brain and pancreas: inactivation of the gene leads to massive proteinuria and neonatal death. Hum. Mol. Genet. 10, 1–8 (2001)

    Article  CAS  Google Scholar 

  23. Rantanen, M. et al. Nephrin TRAP mice lack slit diaphragms and show fibrotic glomeruli and cystic tubular lesions. J. Am. Soc. Nephrol. 13, 1586–1594 (2002)

    Article  CAS  Google Scholar 

  24. Donoviel, D. B. et al. Proteinuria and perinatal lethality in mice lacking NEPH1, a novel protein with homology to NEPHRIN. Mol. Cell. Biol. 21, 4829–4836 (2001)

    Article  CAS  Google Scholar 

  25. Ciani, L., Patel, A., Allen, N. D. & ffrench-Constant, C. Mice lacking the giant protocadherin mFAT1 exhibit renal slit junction abnormalities and a partially penetrant cyclopia and anophthalmia phenotype. Mol. Cell Biol. 23, 3575–3582 (2003)

    Article  CAS  Google Scholar 

  26. Kreidberg, J. A. et al. Alpha 3 beta 1 integrin has a crucial role in kidney and lung organogenesis. Development 122, 3537–3547 (1996)

    CAS  PubMed  Google Scholar 

  27. Kaplan, J. M. et al. Mutations in ACTN4, encoding α-actinin-4, cause familial focal segmental glomerulosclerosis. Nature Genet. 24, 251–256 (2000)

    Article  ADS  CAS  Google Scholar 

  28. Welsch, T., Endlich, N., Kriz, W. & Endlich, K. CD2AP and p130Cas localize to different F-actin structures in podocytes. Am. J. Physiol. Renal Physiol. 281, F769–F777 (2001)

    Article  CAS  Google Scholar 

  29. Kos, C. H. et al. Mice deficient in α-actinin-4 have severe glomerular disease. J. Clin. Invest. 111, 1683–1690 (2003)

    Article  CAS  Google Scholar 

  30. Frischknecht, F. et al. Actin-based motility of vaccinia virus mimics receptor tyrosine kinase signalling. Nature 401, 926–929 (1999)

    Article  ADS  CAS  Google Scholar 

Download references

Acknowledgements

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.

Author information

Author notes

  1. Ivan M. Blasutig and Vera Eremina: *These authors contributed equally to this work

Authors and Affiliations

  1. Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 600 University Avenue, Ontario, M5G 1X5, Toronto, Canada

    Nina Jones, Ivan M. Blasutig, Vera Eremina, Julie M. Ruston, Friedhelm Bladt, Susan E. Quaggin & Tony Pawson

  2. Department of Molecular and Medical Genetics, University of Toronto, 4388 Medical Sciences Building, 1 King's College Circle, Ontario, M5S 1A8, Toronto, Canada

    Ivan M. Blasutig & Tony Pawson

  3. Institute of Medical Science, University of Toronto, 7213 Medical Sciences Building, 1 King's College Circle, Ontario, M5S 1A8, Toronto, Canada

    Vera Eremina & Susan E. Quaggin

  4. Department of Medicine, McGill University Health Centre, 3775 University Street, Québec, H3A 2B4, Montreal, Canada

    Hongping Li, Louise Larose & Tomoko Takano

  5. Department of Biochemistry, University of Western Ontario, Ontario, N6A 5C1, London, Canada

    Haiming Huang & Shawn S.-C. Li

  6. Division of Nephrology, St. Michael's Hospital, 30 Bond Street, Ontario, M5B 1W8, Toronto, Canada

    Susan E. Quaggin

Authors
  1. Nina Jones
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ivan M. Blasutig
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Vera Eremina
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Julie M. Ruston
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Friedhelm Bladt
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Hongping Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Haiming Huang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Louise Larose
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Shawn S.-C. Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Tomoko Takano
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Susan E. Quaggin
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Tony Pawson
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Tony Pawson.

Ethics declarations

Competing interests

Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Supplementary information

Supplementary Figure 1

This figure shows expression of Nck1 and Nck2 in podocytes (by lacZ staining and immunoblotting) as well as the presence of mature foot processes in mice lacking either Nck1 or Nck2. The specificity of Podocin-Cre is shown using the Z/EG reporter strain. (PDF 12859 kb)

Supplementary Figure 2

This figure shows that weaning age mice lacking Nck1 and Nck2 in podocytes are less robust than littermates and display massive albuminuria. (PDF 3882 kb)

Supplementary Figure 3

This figure shows by RNA in situ hybridization analysis of glomeruli that podocytes are progressively lost in mice lacking Nck1 and Nck2 expression in podocytes. (PDF 4436 kb)

Supplementary Figure 4

This figure shows by immunoblotting that N-WASp is recruited to phosphorylated Nephrin containing Nck binding sites. (PDF 695 kb)

Supplementary Figure 5

This figure shows by immunoblotting that Src family kinase activity is required to induce tyrosine phosphorylation of Nephrin. (PDF 1719 kb)

Supplementary Methods

This file contains additional details of the methods used in this study. This file also contains additional references. (DOC 33 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Jones, N., Blasutig, I., Eremina, V. et al. Nck adaptor proteins link nephrin to the actin cytoskeleton of kidney podocytes. Nature 440, 818–823 (2006). https://doi.org/10.1038/nature04662

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature04662

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing