Impaired PTH-induced endocytotic down-regulation of the renal type IIa Na+/Pi-cotransporter in RAP-deficient mice with reduced megalin expression.

Inorganic phosphate (P(i)) reabsorption in the renal proximal tubule occurs mostly via the Na(+)/P(i) cotransporter type IIa (NaP(i)-IIa) located in the brush-border membrane (BBM) and is regulated, among other factors, by dietary P(i) intake and parathyroid hormone (PTH). The PTH-induced inhibition of P(i) reabsorption is mediated by endocytosis of Na/P(i)-IIa from the BBM and subsequent lysosomal degradation. Megalin is involved in receptor-mediated endocytosis of proteins from the urine in the renal proximal tubule. The recently identified receptor-associated protein (RAP) is a novel type of chaperone responsible for the intracellular transport of endocytotic receptors such as megalin. Gene disruption of RAP leads to a decrease of megalin in the BBM and to a disturbed proximal tubular endocytotic machinery. Here we investigated whether the distribution of NaP(i)-IIa and/or its regulation by dietary P(i) intake and PTH is affected in the proximal tubules of RAP-deficient mice as a model for megalin loss. In RAP-deficient mice megalin expression was strongly reduced and restricted to a subapical localization. NaP(i)-IIa protein distribution and abundance in the kidney was not altered. The localization and abundance of the NaP(i)-IIa interacting proteins MAP17, PDZK-1, D-AKAP2, and NHE-RF1 were also normal. Other transport proteins expressed in the BBM such as the Na(+)/H(+) exchanger NHE-3 and the Na(+)/sulphate cotransporter NaSi were normally expressed. In whole animals and in isolated fresh kidney slices the PTH-induced internalization of NaP(i)-IIa was strongly delayed in RAP-deficient mice. PTH receptor expression in the proximal tubule was not affected by the RAP knock-out. cAMP, cGMP or PKC activators induced internalization which was delayed in RAP-deficient mice. In contrast, both wildtype and RAP-deficient mice were able to adapt to high-, normal, and low-P(i) diets appropriately as indicated by urinary P(i) excretion and NaP(i)-IIa protein abundance