Open AccessIntraperitoneal Administration of Recombinant Receptor-Associated Protein Causes Phosphaturia via an Alteration in Subcellular Distribution of the Renal Sodium Phosphate Co-Transporter
Author(s) -
Masayo Yamagata,
Keiichi Ozono,
Yuta Hashimoto,
Yoshiteru Miyauchi,
Hiroki Kondou,
Toshimi Michigami
Publication year - 2005
Publication title -
journal of the american society of nephrology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 4.451
H-Index - 279
eISSN - 1533-3450
pISSN - 1046-6673
DOI - 10.1681/asn.2004070599
Subject(s) - internalization , reabsorption , endocrinology , medicine , chemistry , receptor , endocytic cycle , kidney , endocytosis , renal physiology , recombinant dna , western blot , biochemistry , biology , gene
Megalin is a multifunctional endocytic receptor that is expressed in renal proximal tubules and plays critical roles in the renal uptake of various proteins. It was hypothesized that megalin-dependent endocytosis might play a role in renal phosphate reabsorption. For addressing the short-term effects of altered megalin function, a recombinant protein for the soluble form of 39-kD receptor-associated protein (RAP) was administered intraperitoneally to 7-wk-old mice. Histidine (His)-tagged soluble RAP (amino acids 39 to 356) lacking the amino-terminal signal peptide and the carboxy-terminal endoplasmic reticulum retention signal was prepared by bacterial expression (designated His-sRAP). After the direct interaction between His-sRAP and megalin was confirmed, mice were given a single intraperitoneal administration of His-sRAP (3.5 mg/dose). Immunostaining and Western blot analyses demonstrated the uptake of His-sRAP and the accelerated internalization of megalin in proximal tubular cells 1 h after administration. In addition, internalization of the type II sodium/phosphate co-transporter (NaPi-II) was observed. The effects of three sequential administrations of His-sRAP (3.5 mg/dose, three doses at 4-h intervals) then were examined, and increased urinary excretion of low molecular weight proteins, including vitamin D-binding protein, was found, which is consistent with findings reported for megalin-deficient mice. It is interesting that urinary excretion of phosphate was also increased, and the protein level of NaPi-II in the brush border membrane was decreased. Serum concentration of 25-hydroxyvitamin D was decreased, whereas the plasma level of intact parathyroid hormone was not altered by the administration of His-sRAP. The results suggest that the His-sRAP-induced acceleration of megalin-mediated endocytosis caused phosphaturia via altered subcellular distribution of NaPi-II.