The renal Na+/phosphate cotransporter NaPi-IIa is internalized via the receptor-mediated endocytic route in response to parathyroid hormone

Abstract

The major renal Na(+)/phosphate cotransporter, NaPi-IIa, is regulated by a number of factors including parathyroid hormone (PTH), dopamine, and dietary phosphate intake. PTH induces the acute internalization of NaPi-IIa from the brush border membrane (BBM) and its routing to and subsequent degradation in lysosomes. Previous work indicated that megalin, part of the apical receptor-mediated endocytic apparatus, may play a role in the PTH-induced removal of NaPi-IIa. Here we examined in rats the time-dependent internalization route of NaPi-IIa after acute PTH application using immunohistochemistry and markers of several endocytic compartments. NaPi-IIa removal from the BBM was detectable as early as 5 min after PTH injection. After 10-15 min, NaPi-IIa was localized in subapical compartments positive for clathrin. Shortly thereafter, NaPi-IIa appeared in endosomes stained for EEA1 (early endosomal antigen 1). After 45-60 min, NaPi-IIa was found in late endosomes/lysosomes marked with lgp120. In contrast, no change in the subcellular localization of megalin and the Na(+)/H(+) exchanger NHE3 was detected up to 60 min after PTH injection. To further characterize the internalization route, insulin, as a marker for receptor-mediated endocytosis, and horseradish peroxidase (HRP) and fluorescein isothiocyanate (FITC)-dextran (10 kDa), as markers for fluid-phase mediated endocytosis, were used. NaPi-IIa colocalized with insulin 5-30 min after PTH injection but did not overlap with HRP or FITC-dextran. These results demonstrate a distinct internalization route of NaPi-IIa in response to acute PTH application that may involve the receptor-mediated endocytic pathway including clathrin-coated vesicles and EEA1-positive early endosomes, and routes NaPi-IIa to lysosomes for degradation.

The phosphate transporter NaPi-IIa determines the rapid renal adaptation to dietary phosphate intake in mouse irrespective of persistently high FGF23 levels

“…In rat, mouse, and rabbit, NaPi-IIa is upregulated by low Pi diet and decreased by high Pi intake both acutely and chronically [16-17, 29, 57]. This regulation occurs essentially by trafficking of the protein, lysosomal degradation, recruitment of newly synthesized transporters [5,26,[29][30] and may also include transcriptional regulation [34]. Much less is known about the regulation of NaPi-IIc being regulated also by PTH, FGF23, and Pi intake with a slower change in brush border membrane abundance upon high Pi intake or PTH application [8,28,[43][44].…”

“…Mouse kidneys were perfusion fixed through the right ventricule with a 4% paraformaldehyde fixative/PBS solution and the subsequent immunohistochemistry were performed as described previously [5]. Slices of fixed kidneys were frozen in liquid propane and cooled with liquid nitrogen.…”

NaPi-IIa interacting proteins and regulation of renal reabsorption of phosphate

“…We have evidences that PTHinduced endocytosis of NaPi-IIa takes place at least partially via clathrin-coated pits [1], opening the question of whether the absence of GABARAP would blunt the hormonal effect. However PTH downregulates the expression of NaPi-IIa to comparable levels in wild type and GABARAP deficient mice.…”

“…Since P i is freely filtered in the glomerulus, an efficient renal reabsorption is required to prevent a massive urinary lost of P i that would have otherwise fatal consequences. In this regard, hypophosphatemia increases intrarenal production of 1,25 dihydroxyvitamin D 3 (1,25(OH) 2 A. Renal reabsorption of phosphate Reabsorption of P i from the primary urine takes place preferentially along the proximal tubule with very little contribution of the distal segments. Several Na-dependent P i cotransporters from the SLC34 and SLC20 families of solute carriers are expressed in the brush border membrane (BBM) of proximal tubular cells, namely NaPi-IIa (SLC34A1) [8], NaPi-IIc (SLC34A3) [25] and PiT2 (SLC20A2) [30].…”

Effects of Calcium Administration on Parathyroid Gland, NaPi 2a Cotransporter and PTH1R in an Animal Model of the Andropause / EFEKTI TRETMANA KALCIJUMOM NA PARATIREOIDNU ŽLEZDU, NAPI 2A KOTRANSPORTER I PTH1R U ANIMALNOM MODELU ANDROPAUZE

“…In addition, the ob ser ved increase in the immunofluorescent signal of PTH1R in Orx animals' epithelial cells of the kidney proximal tubules could also point to reduced Pi uptake from primary urine. Namely, since PTH has a stimulatory effect on PTH1R mRNA stability (26), an increased presence of PTH1R could provide the inhibitory effect of PTH on abundance of NaPi 2a (27,28). After Ca 2+ treatment, the expected increase in serum Ca 2+ occurred, together with a rise in serum Pi concentration, the latter implying enhancement of Pi reabsorption rate from the urine.…”