title: Increased SHP-1 protein expression by high glucose levels reduces nephrin phosphorylation in podocytes.
abstract: Nephrin, a critical podocyte membrane component that is reduced in diabetic nephropathy, has been shown to activate phosphotyrosine signaling pathways in human podocytes. Nephrin signaling is important to reduce cell death induced by apoptotic stimuli. We have shown previously that high glucose level exposure and diabetes increased the expression of SHP-1, causing podocyte apoptosis. SHP-1 possesses two Src homology 2 domains that serve as docking elements to dephosphorylate tyrosine residues of target proteins. However, it remains unknown whether SHP-1 interacts with nephrin and whether its elevated expression affects the nephrin phosphorylation state in diabetes. Here we show that human podocytes exposed to high glucose levels exhibited elevated expression of SHP-1, which was associated with nephrin. Coexpression of nephrin-CD16 and SHP-1 reduced nephrin tyrosine phosphorylation in transfected human embryonic kidney 293 cells. A single tyrosine-to-phenylalanine mutation revealed that rat nephrin Tyr(1127) and Tyr(1152) are required to allow SHP-1 interaction with nephrin. Overexpression of dominant negative SHP-1 in human podocytes prevented high glucose-induced reduction of nephrin phosphorylation. In vivo, immunoblot analysis demonstrated that nephrin expression and phosphorylation were decreased in glomeruli of type 1 diabetic Akita mice (Ins2(+/C96Y)) compared with control littermate mice (Ins2(+/+)), and this was associated with elevated SHP-1 and cleaved caspase-3 expression. Furthermore, immunofluorescence analysis indicated increased colocalization of SHP-1 with nephrin in diabetic mice compared with control littermates. In conclusion, our results demonstrate that high glucose exposure increases SHP-1 interaction with nephrin, causing decreased nephrin phosphorylation, which may, in turn, contribute to diabetic nephropathy.
authors.affiliation: From the Research Center of the Centre Hospitalier Universitaire de Sherbrooke and Division of Endocrinology, Departments of Medicine, Université de Sherbrooke, Sherbrooke, Québec J1H 5N4, Canada.
authors.forename: Benoit
authors.initials: B
authors.lastname: Denhez
authors.affiliation: From the Research Center of the Centre Hospitalier Universitaire de Sherbrooke and Division of Endocrinology, Departments of Medicine, Université de Sherbrooke, Sherbrooke, Québec J1H 5N4, Canada.
authors.forename: Farah
authors.initials: F
authors.lastname: Lizotte
authors.affiliation: From the Research Center of the Centre Hospitalier Universitaire de Sherbrooke and Division of Endocrinology, Departments of Medicine, Université de Sherbrooke, Sherbrooke, Québec J1H 5N4, Canada.
authors.forename: Marie-Odile
authors.initials: MO
authors.lastname: Guimond
authors.affiliation: the Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario N1G 2W1, Canada, and.
authors.forename: Nina
authors.initials: N
authors.lastname: Jones
authors.affiliation: the McGill University Health Center, Montreal, Québec H3H 2R9, Canada.
authors.forename: Tomoko
authors.initials: T
authors.lastname: Takano
authors.affiliation: From the Research Center of the Centre Hospitalier Universitaire de Sherbrooke and Division of Endocrinology, Departments of Medicine, Université de Sherbrooke, Sherbrooke, Québec J1H 5N4, Canada, Pedro.Geraldes@USherbrooke.ca.
authors.forename: Pedro
authors.initials: P
authors.lastname: Geraldes
mesh_terms: D000818:Animals; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D003922:Diabetes Mellitus, Type 1 / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D003928:Diabetic Nephropathies / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D004195:Disease Models, Animal; D058851:GPI-Linked Proteins / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D005947:Glucose / Q000633:toxicity; D057809:HEK293 Cells; D006801:Humans; D007328:Insulin / Q000235:genetics / Q000378:metabolism; D007678:Kidney Glomerulus / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D050199:Podocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6 / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D017452:Receptors, IgG / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction
chemical_list: C428200:FCGR3B protein, human; D058851:GPI-Linked Proteins; C561967:Ins2 protein, mouse; D007328:Insulin; D008565:Membrane Proteins; D017452:Receptors, IgG; C113267:nephrin; D019000:Phosphotyrosine; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; C509250:Ptpn6 protein, rat; D053148:Caspase 3; D005947:Glucose
keywords: Diabetes; Nephrin; Nephrology; Phosphotyrosine; Podocyte; Protein-Tyrosine Phosphatase (Tyrosine Phosphatase); Src Homology 2 domain (SH2 domain)