Cell junction-associated proteins IQGAP1, MAGI-2, CASK, spectrins, and alpha-actinin are components of the nephrin multiprotein complex.
Nephrin is a cell surface receptor of the Ig superfamily that localizes to slit diaphragms, the specialized junctions between the interdigitating foot processes of the glomerular epithelium (podocytes) in the kidney. Mutations in the NPHS1 gene encoding nephrin lead to proteinuria and congenital nephrotic syndrome, indicating that nephrin is essential for normal glomerular development and function. To identify nephrin-binding proteins, we performed mass spectrometry on proteins obtained from pull-down assays with GST-nephrin cytoplasmic domain. Nephrin specifically pulled down six proteins from glomerular lysates, MAGI-2/S-SCAM (membrane-associated guanylate kinase inverted 2/synaptic scaffolding molecule), IQGAP1 (IQ motif-containingGTPase-activatingprotein1),CASK(calcium/calmodulin-dependent serine protein kinase), alpha-actinin, alphaII spectrin, and betaII spectrin. All of these scaffolding proteins are often associated with cell junctions. By immunofluorescence these proteins are expressed in glomerular epithelial cells, where they colocalize with nephrin in the foot processes. During glomerular development, IQGAP1 is expressed in the junctional complexes between the earliest identifiable podocytes, MAGI-2/S-SCAM is first detected in junctional complexes in podocytes after their migration to the base of the cells. Thus, the nephrin-slit diaphragm protein complex contains a group of scaffolding proteins that function to connect junctional membrane proteins to the actin cytoskeleton and signaling cascades. Despite their special morphology and function, there is considerable compositional similarity between the podocyte slit diaphragm and typical junctional complexes of other epithelial cells.
Lehtonen
Sanna
S
Departments of Cellular and Molecular Medicine and Pathology, and Ludwig Institute for Cancer Research, University of California at San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA.
Ryan
Jennifer J
JJ
Kudlicka
Krystyna
K
Iino
Noriaki
N
Zhou
Huilin
H
Farquhar
Marilyn G
MG
D000185:Actinin / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D005982:Glutathione Transferase; D051528:Guanylate Kinases; D007365:Intercellular Junctions / Q000378:metabolism*; D007678:Kidney Glomerulus / Q000166:cytology / Q000378:metabolism*; D008297:Male; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D017207:Rats, Sprague-Dawley; D013049:Spectrin / Q000378:metabolism; D020703:ras GTPase-Activating Proteins / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; C101203:IQ motif containing GTPase activating protein 1; C114103:Magi2 protein, rat; D008565:Membrane Proteins; D046912:Multiprotein Complexes; C113267:nephrin; D020703:ras GTPase-Activating Proteins; D000185:Actinin; D013049:Spectrin; D005982:Glutathione Transferase; C106868:CASK kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D051528:Guanylate Kinases