Phosphorylation of slit diaphragm proteins NEPHRIN and NEPH1 upon binding of HGF promotes podocyte repair

Ashish K. Solanki, Ehtesham Arif, Pankaj Srivastava, Christopher M. Furcht, Bushra Rahman, Pei Wen, Avinash Singh, Lawrence B. Holzman, Wayne R. Fitzgibbon, Milos N. Budisavljevic, Glenn P. Lobo, Sang Ho Kwon, Zhe Han, Matthew J. Lazzara, Joshua H. Lipschutz, Deepak Nihalani

Research output: Contribution to journalArticlepeer-review

Abstract

Phosphorylation (activation) and dephosphorylation (deactivation) of the slit diaphragm proteins NEPHRIN and NEPH1 are critical for maintaining the kidney epithelial podocyte actin cytoskeleton and, therefore, proper glomerular filtration. However, the mechanisms underlying these events remain largely unknown. Here we show that NEPHRIN and NEPH1 are novel receptor proteins for hepatocyte growth factor (HGF) and can be phosphorylated independently of the mesenchymal epithelial transition receptor in a ligand-dependent fashion through engagement of their extracellular domains by HGF. Furthermore, we demonstrate SH2 domain-containing protein tyrosine phosphatase-2-dependent dephosphorylation of these proteins. To establish HGF as a ligand, purified baculovirus-expressed NEPHRIN and NEPH1 recombinant proteins were used in surface plasma resonance binding experiments. We report high-affinity interactions of NEPHRIN and NEPH1 with HGF, although NEPHRIN binding was 20-fold higher than that of NEPH1. In addition, using molecular modeling we constructed peptides that were used to map specific HGF-binding regions in the extracellular domains of NEPHRIN and NEPH1. Finally, using an in vitro model of cultured podocytes and an ex vivo model of Drosophila nephrocytes, as well as chemically induced injury models, we demonstrated that HGF-induced phosphorylation of NEPHRIN and NEPH1 is centrally involved in podocyte repair. Taken together, this is the first study demonstrating a receptor-based function for NEPHRIN and NEPH1. This has important biological and clinical implications for the repair of injured podocytes and the maintenance of podocyte integrity.

Original languageEnglish (US)
Article number101079
JournalJournal of Biological Chemistry
Volume297
Issue number3
DOIs
StatePublished - Sep 1 2021

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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