Cloning of rat nephrin: expression in developing glomeruli and in proteinuric states.
BACKGROUND
Nephrin is identified as a product of the gene mutated in a patient with congenital nephrotic syndrome of the Finnish type. However, its precise localization and function are not yet fully clarified.


METHODS
To clone the rat homologue of nephrin, polymerase chain reaction (PCR) was employed. To elucidate the localization and expression of nephrin, immunohistological analysis with a specific antirat nephrin antibody, reverse transcription-PCR, and RNase protection assay were performed.


RESULTS
Amino acid sequences of rat and human nephrin are highly homologous (82.2% identity). The domain structure of nephrin is also highly conserved between rats and humans. The rat nephrin was detected only in kidney glomeruli along glomerular capillary walls, and its localization was always identical to that of the anti-slit diaphragm monoclonal antibody (mAb) 5-1-6-recognized antigen in normal matured and fetal rat glomeruli and in the glomeruli of proteinuric states. The nephrin staining pattern was clearly distinguished from that of zonula occludens-1 (ZO-1), alpha3-integrin, or podocalyxin. mRNA expression for nephrin was first detected in the fetal rat kidneys at 18.5 embryonic days. Nephrin mRNA expression decreased just after injection of mAb 5-1-6 (47.4%) or puromycin aminonucleoside (51.2%), and the staining pattern of nephrin shifted from a linear to a granular pattern in both proteinuric states.


CONCLUSIONS
Nephrin is localized in slit diaphragm in the matured glomeruli and is identical with mAb 5-1-6 antigen. Nephrin is involved in the development of proteinuria not only in mAb 5-1-6 nephropathy, but also in puromycin aminonucleoside nephropathy.
Kawachi
H
H
Department of Cell Biology and Department of Pathology, Institute of Nephrology, Niigata University School of Medicine, Japan. kawachi@med.niigata-u.ac.jp
Koike
H
H
Kurihara
H
H
Yaoita
E
E
Orikasa
M
M
Shia
M A
MA
Sakai
T
T
Yamamoto
T
T
Salant
D J
DJ
Shimizu
F
F
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001483:Base Sequence; D003001:Cloning, Molecular; D005333:Fetus / Q000378:metabolism; D006801:Humans; D007678:Kidney Glomerulus / Q000737:chemistry / Q000196:embryology / Q000378:metabolism*; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D009928:Organ Specificity; D011506:Proteins / Q000032:analysis / Q000235:genetics*; D011507:Proteinuria / Q000209:etiology / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction
D000911:Antibodies, Monoclonal; D008565:Membrane Proteins; D011506:Proteins; D012333:RNA, Messenger; C113267:nephrin