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. [ABSTRACT FROM AUTHOR]