Parathyroid hormone leads to the lysosomal degradation of the renal type II Na/P _i cotransporter

Abstract

We have studied the involvement of proteolytic pathways in the regulation of the Na/P i cotransporter type II by parathyroid hormone (PTH) in opossum kidney cells. Inhibition of lysosomal degradation (by leupeptin, ammonium chloride, methylamine, chloroquine, l -methionine methyl ester) prevented the PTH-mediated degradation of the transporter, whereas inhibition of the proteasomal pathway (by lactacystin) did not. Moreover it was found ( i ) that whereas lysosomal inhibitors prevented the PTH-mediated degradation of the transporter they did not prevent the PTH-mediated inhibition of the Na/P i cotransport and ( ii ) that treating opossum kidney cells with lysosomal inhibitors led to an increased expression of the transporter without any concomitant increase in the Na/P i cotransport. Further analysis by subcellular fractionation and morphological techniques showed ( i ) that the Na/P i cotransporter is constitutively transported to and degraded within late endosomes/lysosomes and ( ii ) that PTH leads to the increased degradation of the transporter in late endosomes/lysosomes.

A dibasic motif involved in parathyroid hormone-induced down-regulation of the type IIa NaPi cotransporter

“…Opossum kidney cells retain the proximal tubular characteristics of PTH-induced regulation of brush-border membrane NaPi cotransport, associated with membrane retrieval and lysosomal degradation of type IIa NaPi cotransporters (10,11,(16)(17)(18)(30)(31)(32)(33). As a similar regulatory behavior was observed for the intrinsic (NaPi-4) and transfected (NaPi-2) transporter (10,11,14,19), OK cells represent an ideal in vitro system to study molecular determinants responsible for PTH-induced downregulation of the type IIa NaPi cotransporter.…”

“…OK cells were plated on glass coverslips and transfected as described above. After confluence, they were fixed in 3% paraformaldehyde and permeabilized with 0.1% saponine as described (11). They were then incubated 30 min in the presence of phalloidine-Texas Red to stain the actin.…”

“…PTH exerts its effect by an inhibition of proximal tubular brush-border NaPi cotransport associated with a membrane retrieval and lysosomal degradation of type IIa NaPi cotransporters (4, 6-9). These PTH-induced regulations are retained in cultured opossum kidney (OK) cells for both the endogenous (NaPi-4) and the transfected rat (NaPi-2) type IIa NaPi cotransporter (10)(11)(12)(13)(14). In proximal tubules and in OK cells, two signaling pathways are involved in PTH-induced downregulation of P i transport and of type IIa NaPi cotransporter: protein kinase A and protein kinase C (15-19).…”

Targeted disruption of the mouse NHERF-1 gene promotes internalization of proximal tubule sodium-phosphate cotransporter type IIa and renal phosphate wasting

“…Like NHE3, Npt2 function is inhibited by parathyroid hormone and cAMP. Despite the presence of multiple phosphorylation sites in the Npt2 protein (22), the predominant mechanism by which hormones regulate Npt2 activity is by internalization and subsequent lysosomal degradation of the transporter protein (42). At least four different PDZ proteins, including NHERF-1 and NHERF-2, bound Npt2 and may play a role in endocytosis and sorting of the renal phosphate transporter (7).…”

Disorders of Renal Tubular Phosphate Transport

“…Internalized transporters are then routed to the lysosomes for degradation (41,42). There is no evidence for recycling of the type IIa protein, although it cannot be excluded as a fine-tuning system in response to small fluctuations in PTH levels.…”