Nephrin-Binding Ephrin-B1 at the Slit Diaphragm Controls Podocyte Function through the JNK Pathway

J Am Soc Nephrol. 2018 May;29(5):1462-1474. doi: 10.1681/ASN.2017090993. Epub 2018 Mar 30.


Background B-type ephrins are membrane-bound proteins that maintain tissue function in several organs. We previously reported that ephrin-B1 is localized at the slit diaphragm of glomerular podocytes. However, the function of ephrin-B1 at this location is unclear.Methods We analyzed the phenotype of podocyte-specific ephrin-B1 knockout mice and assessed the molecular association of ephrin-B1 and nephrin, a key molecule of the slit diaphragm, in HEK293 cells and rats with anti-nephrin antibody-induced nephropathy.Results Compared with controls, ephrin-B1 conditional knockout mice displayed altered podocyte morphology, disarrangement of the slit diaphragm molecules, and proteinuria. Ephrin-B1 expressed in HEK293 cells immunoprecipitated with nephrin, which required the basal regions of the extracellular domains of both proteins. Treatment of cells with an anti-nephrin antibody promoted the phosphorylation of nephrin and ephrin-B1. However, phosphorylation of ephrin-B1 did not occur in cells expressing a mutant nephrin lacking the ephrin-B1 binding site or in cells treated with an Src kinase inhibitor. The phosphorylation of ephrin-B1 enhanced the phosphorylation of nephrin and promoted the phosphorylation of c-Jun N-terminal kinase (JNK), which was required for ephrin-B1-promoted cell motility in wound-healing assays. Notably, phosphorylated JNK was detected in the glomeruli of control mice but not ephrin-B1 conditional knockout mice. In rats, the phosphorylation of ephrin-B1, JNK, and nephrin occurred in the early phase (24 hours) of anti-nephrin antibody-induced nephropathy.Conclusions Through interactions with nephrin, ephrin-B1 maintains the structure and barrier function of the slit diaphragm. Moreover, phosphorylation of ephrin-B1 and, consequently, JNK are involved in the development of podocyte injury.

Keywords: Cell Signaling; cell adhesion; glomerular filtration barrier; nephrin; podocyte; proteinuria.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies
  • Cell Movement
  • Ephrin-B1 / genetics*
  • Ephrin-B1 / metabolism*
  • HEK293 Cells
  • Humans
  • JNK Mitogen-Activated Protein Kinases / metabolism
  • Membrane Proteins / immunology
  • Membrane Proteins / metabolism*
  • Mice
  • Mice, Knockout
  • Nephrosis / immunology
  • Nephrosis / metabolism*
  • Phosphorylation
  • Podocytes / metabolism*
  • Podocytes / pathology
  • Rats


  • Antibodies
  • EFNB1 protein, human
  • Efnb1 protein, mouse
  • Efnb1 protein, rat
  • Ephrin-B1
  • Membrane Proteins
  • nephrin
  • JNK Mitogen-Activated Protein Kinases