Role of Fat1 in cell-cell contact formation of podocytes in puromycin aminonucleoside nephrosis and neonatal kidney

Abstract

Background. Fat1 is a member of the cadherin superfamily characterized by its 34 cadherin repeats in the extracellular domain. Fat1 was originally found as a component of the slit diaphragm of podocytes, but its function in podocytes remains obscure. To gain insight into its role in podocytes, we expanded our study of Fat1 expression to puromycin aminonucleoside (PAN) nephrosis, the neonatal kidney, and the primary podocyte culture, where slit diaphragms are absent or disappear. Methods. Expression of Fat1 was examined in isolated glomeruli of PAN nephrosis by the ribonuclease protection assay and Western blot analysis and in the neonatal kidney by in situ hybridization. Fat1 localization in glomeruli and in the primary culture was confirmed by immunofluorescence or immunoelectron microscopy. Results. In PAN nephrotic rats, glomerular expression of Fat1 increased rather than decreased at both transcript and protein levels in comparison with normal rats. Immunofluorescence microscopy revealed distinct staining for Fat1 along the glomerular capillary wall, where nephrin staining was weakened or disappeared. Immunoelectron microscopy demonstrated significant accumulation of immunogold particles for Fat1 at intercellular junctions newly formed between podocytes in the nephrosis. In the primary culture of podocytes, Fat1 was mainly localized at cell-cell contact sites and in tips of cellular processes. In the neonatal kidney, immature podocytes expressed Fat1 more intensely than mature podocytes as shown by in situ hybridization. Double-labeled immunostaining using anti-pan cadherin antibody revealed that Fat1 in podocytes colocalized with cadherin in immature glomeruli, indicating that junctional complexes of developing podocytes contain Fat1. Conclusion. These findings suggest that Fat1 may be a fundamental component of intercellular junctions of podocytes, and may be involved in the initial step of cell contacts of podocytes. © 2005 by the International Society of Nephrology.