title: Integrin Ligation Results in Nephrin Tyrosine Phosphorylation In Vitro.
abstract: Nephrin is expressed at the basolateral aspect of podocytes and is an important signaling protein at the glomerular slit diaphragm. In vitro studies have demonstrated that Nephrin phosphorylation-dependent signaling is able to assemble a protein complex that is able to polymerize actin. However, proximal signaling events that result in nephrin tyrosine phosphorylation are not well understood. Nephrin deletion in mice and human nephrin mutations result in developmental failure of the podocyte intercellular junction resutling in proteinuria. This has been presumed to be due to a failure to respond to an external polarized cue in the absence of nephrin or a failure to transduce an outside-in signal in patients with nephrin mutations. The nephrin extracellular domain binds to itself or neph1 across the foot process intercellular junction. Nephrin is tyrosine phosphorylation-silent in healthy glomeruli when presumably the nephrin extracellular domain is in an engaged state. These observations raise the possibility of an alternate proximal signaling mechanism that might be responsible for nephrin tyrosine phosphorylation. Here we present data showing that integrin engagement at the basal aspect of cultured podocytes results in nephrin tyrosine phosphorylation. This is abrogated by incubating podocytes with an antibody that prevents integrin β1 ligation and activation in response to binding to extracellular matrix. Furthermore, nephrin tyrosine phosphorylation was observed in podocytes expressing a membrane-targeted nephrin construct that lacks the extracellular domain. We propose, integrin-activation based signaling might be responsible for nephrin phosphorylation rather than engagment of the nephrin extracellular domain by a ligand.
authors: Division of Nephroloigy, Department of Internal Medicine, University of Michigan School of Medicine, Ann Arbor, Michigan, United States of America.
authors: Rakesh
authors: R
authors: Verma
authors: Division of Nephroloigy, Department of Internal Medicine, University of Michigan School of Medicine, Ann Arbor, Michigan, United States of America.
authors: Madhusudan
authors: M
authors: Venkatareddy
authors: Division of Nephroloigy, Department of Internal Medicine, University of Michigan School of Medicine, Ann Arbor, Michigan, United States of America.
authors: Anne
authors: A
authors: Kalinowski
authors: Division of Nephroloigy, Department of Internal Medicine, University of Michigan School of Medicine, Ann Arbor, Michigan, United States of America.
authors: Sanjeevkumar R
authors: SR
authors: Patel
authors: Division of Nephroloigy, Department of Internal Medicine, University of Michigan School of Medicine, Ann Arbor, Michigan, United States of America.
authors: Puneet
authors: P
authors: Garg
mesh_terms: D000818:Animals; D057809:HEK293 Cells; D006801:Humans; D019012:Integrin beta1 / Q000032:analysis / Q000378:metabolism; D016023:Integrins / Q000378:metabolism*; D008565:Membrane Proteins / Q000032:analysis / Q000378:metabolism*; D051379:Mice; D010766:Phosphorylation; D015398:Signal Transduction; D014443:Tyrosine / Q000378:metabolism*
chemical_list: D019012:Integrin beta1; D016023:Integrins; D008565:Membrane Proteins; C113267:nephrin; D014443:Tyrosine