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

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

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Department of Cell Biology, Kidney Research Center, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Japan.
Department of Comparative and Experimental Medicine, Brain Research Institute, Niigata University, Niigata, Japan.
Department of Pediatrics, Niigata City General Hospital, Niigata, Japan; and.
Department of Internal Medicine IV, Teikyo University School of Medicine, Kawasaki, Japan.
Department of Cell Biology, Kidney Research Center, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Japan;


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.


Cell Signaling; cell adhesion; glomerular filtration barrier; nephrin; podocyte; proteinuria

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