Role of PDZK1 Protein in Apical Membrane Expression of Renal Sodium-coupled Phosphate Transporters

Abstract

The sodium-dependent phosphate (Na/P i ) transporters NaPi-2a and NaPi-2c play a major role in the renal reabsorption of P i . The functional need for several transporters accomplishing the same role is still not clear. However, the fact that these transporters show differential regulation under dietary and hormonal stimuli suggests different roles in P i reabsorption. The pathways controlling this differential regulation are still unknown, but one of the candidates involved is the NHERF family of scaffolding PDZ proteins. We propose that differences in the molecular interaction with PDZ proteins are related with the differential adaptation of Na/P i transporters. Pdzk1 ؊/؊ mice adapted to chronic low P i diets showed an increased expression of NaPi-2a protein in the apical membrane of proximal tubules but impaired up-regulation of NaPi-2c. These results suggest an important role for PDZK1 in the stabilization of NaPi-2c in the apical membrane. We studied the specific protein-protein interactions of Na/P i transporters with NHERF-1 and PDZK1 by FRET. FRET measurements showed a much stronger interaction of NHERF-1 with NaPi-2a than with NaPi-2c. However, both Na/P i transporters showed similar FRET efficiencies with PDZK1. Interestingly, in cells adapted to low P i concentrations, there were increases in NaPi-2c/PDZK1 and NaPi-2a/NHERF-1 interactions. The differential affinity of the Na/P i transporters for NHERF-1 and PDZK1 proteins could partially explain their differential regulation and/or stability in the apical membrane. In this regard, direct interaction between NaPi-2c and PDZK1 seems to play an important role in the physiological regulation of NaPi-2c.The type II sodium-coupled phosphate (Na/P i ) 3 transporters are the molecules responsible for tubular reabsorption of P i and are the target of hormonal and nonhormonal mechanisms that control phosphate homeostasis. NaPi-2a (NaPiIIa) is responsible for ϳ70% of the P i reabsorbed in the adult kidney of mice, whereas NaPi-2c (NaPiIIc) handles the remaining 30% (1, 2). A low dietary P i intake induces an increase of P i reabsorption in the proximal tubule mediated by augmented apical expression of NaPi-2a and NaPi-2c transporters and consequent increased P i uptake (3-6).Despite sharing a very similar molecular structure, NaPi-2a and NaPi-2c exhibit an increasing list of physiological differences. For example, whereas NaPi-2a is electrogenic, NaPi-2c is electroneutral (4, 7). In addition, both transporters participate in the renal adaptation to changes in dietary P i with different characteristics. In response to a high P i intake, NaPi-2a abundance decreases quickly (less than 1 h), internalization takes place in a microtubule-independent way, and molecules are targeted to the lysosomes via endosomes (8, 9). In contrast, under a high P i intake, NaPi-2c abundance decreases slowly (4 h), molecules are internalized through a microtubule-dependent pathway, and rather than being degraded, they are accumulated in a subapical compartment (10). Similar differences...

Renal Phosphate Wasting in the Absence of Adenylyl Cyclase 6

“…In contrast to studies localizing Npt2c mainly to the S1 and weaker in the S2 segment, 6,17 our antibody 16,42,43 identified Npt2c along the entire length of proximal tubules, with clear transitions from a labeled S3 segment to an unlabeled thin descending limb of Henle's loop. Our antibody previously gave a strong signal in the outer stripe of the outer medulla in WT mice, consistent with Npt2c expression in S3 segments.…”

“…Our antibody previously gave a strong signal in the outer stripe of the outer medulla in WT mice, consistent with Npt2c expression in S3 segments. 16 The reason for this discrepancy remains unknown; however, it might relate to species differences. Why is FGF-23 increased and 1,25 (OH 2 )D suppressed in AC6 2/2 mice?…”

“…26,27 Proteins were transferred to nitrocellulose membranes and immunoblotted with Npt2a (1:1000) and Npt2c (1:2500). 16,42 Chemiluminescent detection was performed with ECL Plus (Amersham, Piscataway, NJ). Densitometric analysis was performed by ImageJ Software (National Institutes of Health).…”

“…6,14,15 Npt2c has a similar cellular distribution with greater abundance in S1 segments. 5,16,17 Genetic defects in G sa result in impaired PTH-induced retrieval of Npt2a, resulting in a failure to respond with increased urinary P i excretion: a disease commonly referred to as pseudohypoparathyroidism type Ia or Ib (depending on the presence or absence of skeletal defects, respectively). [18][19][20] In comparison, less is known about the role of AC proteins in mediating the signaling between PTH 1 R and Npt2a and/or Npt2c.…”

Na+/H+ Exchanger Regulatory Factor 3 Is Critical for Multidrug Resistance Protein 4–Mediated Drug Efflux in the Kidney

“…NHERF3 has been reported to regulate membrane proteins in the digestive organs and the kidney, 7,8,10,19 and therefore, we first examined expression patterns of NHERF3 using tissues from wild-type (Nherf3 +/+ ) and Nherf3 2/2 mice. Immunoblotting experiments confirmed the absence of Nherf3 protein in the Nherf3 2/2 mice (Supplemental Figure 3).…”