Decreased tyrosine phosphorylation of nephrin in rat and human nephrosis.
Phosphorylation of tyrosine residue (Y1204) of rat nephrin by Fyn kinase allows Nck adaptor protein binding to nephrin motifs, which include the phosphorylated tyrosine. This phosphorylation-dependent switch induces actin polymerization in a cell culture system. Here, we generated an antibody recognizing phosphorylated nephrin at the Nck binding sites pY1204 and pY1228 to determine the phosphorylation status of nephrin using a rat model of puromycin aminonucleoside-induced nephrosis. Changes in globular actin (G-actin) and filamentous actin (F-actin) contents in isolated glomeruli were measured by western blot. Before experimental nephrosis, both Y1204 and Y1228 were phosphorylated, and most of the actin was filamentous. Before the onset of overt proteinuria, however, phosphorylation of both Y1204 and Y1228 rapidly decreased and became almost undetectable. During this period, the amount of F-actin in glomeruli began to decrease, whereas G-actin increased. Phosphorylation of nephrin at Y1228 in glomeruli of patients with minimal change nephrosis was significantly decreased compared with that in normal glomeruli. Our study suggests that tyrosine phosphorylation of nephrin by regulating F-actin formation may be important for the maintenance of normal podocyte morphology and function.
Uchida
K
K
Department of Medicine, Kidney Center, Tokyo Women's Medical University, Tokyo, Japan. kuchida@kc.twmu.ac.jp
Suzuki
K
K
Iwamoto
M
M
Kawachi
H
H
Ohno
M
M
Horita
S
S
Nitta
K
K
D000199:Actins / Q000378:metabolism*; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification; D000964:Antimetabolites, Antineoplastic / Q000633:toxicity; D019556:COS Cells; D002522:Chlorocebus aethiops; D003599:Cytoskeleton / Q000378:metabolism; D006801:Humans; D007678:Kidney Glomerulus / Q000378:metabolism*; D008565:Membrane Proteins / Q000276:immunology / Q000378:metabolism*; D009401:Nephrosis / Q000139:chemically induced / Q000378:metabolism*; D010766:Phosphorylation; D050199:Podocytes / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011692:Puromycin Aminonucleoside / Q000633:toxicity; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology
D000199:Actins; D000906:Antibodies; D000964:Antimetabolites, Antineoplastic; D008565:Membrane Proteins; C113267:nephrin; D011692:Puromycin Aminonucleoside; D011505:Protein-Tyrosine Kinases