Role of PDZ domain-containing proteins and ERM proteins in regulation of renal function and dysfunction.

Hormonal, dietary, and metabolic factors play an important role in regulation of renal proximal tubular apical membrane sodium-hydrogen (Na/H) exchange and sodium-phosphate (Na/Pi) cotransport and basolateral membrane sodium-potassium-ATPase (Na-K-ATPase) activity by diverse cellular mechanisms including transcriptional, translational, and posttranslational mechanisms. Recent work in this area indicates novel protein interactions that play an important role in the fine regulation of these important physiologic processes. Parathyroid hormone (PTH) is well known to cause rapid inhibition of Na/H exchange activity and Na/Pi cotransport activity by inducing endocytic internalization of Na/H exchange protein (NHE3) (1) and Na/Pi cotransport protein (NaPi IIa) (2) from the apical brush border membrane. PTH also causes rapid inhibition of Na-K-ATPase activity and endocytic internalization of Na-K-ATPase protein (3) from the basolateral membrane. PTH activates a number of protein kinases including protein kinase A (PKA), C (PKC), G (PKG), and ERK/MAPK kinase. Although PTH and/or the signaling intermediates that activate these protein kinases have been shown to induce phosphorylation of NHE3 and Na-K-ATPase before or in parallel with the inhibition of their activity (1,3), it has not been possible to date to demonstrate the direct phosphorylation of NaPi IIa (4). This has raised the possibility of closely interacting regulatory proteins that may be a target of PTH and regulate NaPi IIa independent of direct phosphorylation. Indeed biochemical and molecular (yeast two-hybrid) approaches have led to the identification of two novel classes of proteins, PDZ domain-containing proteins and ERM proteins that play an important role in the regulation of NHE3, NaPi IIa, and Na-K-ATPase activity and their close association with the actin cytoskeleton and the regulatory hormone regulated kinases, including PTH-activated PKA. PDZ domains are modular protein interaction domains that often occur in scaffolding proteins that bind in a sequence-specific fashion the C-terminal peptide sequence or at times the internal peptide sequences of target proteins. These domains of approximately 90 amino acids are known by an acronym of the first three PDZ-containing proteins identified including the postsynaptic protein PSD-95/SAP90, the Drosophila septate junction protein Discs-large, and the tight junction protein ZO-1 (5–7). In recent years, several proteins that contain PDZ domains including NHERF-1, NHERF-2, PDZK1, CAL, and ZO-1 have been identified in the kidney that play an important role in the following: (a) the regulation of the expression and activity of renal proximal tubular brush border membrane transport proteins, including NHE-3 (5) and NaPi IIa (4), basolateral membrane transport proteins, including Na-K-ATPase, and Na-HC03 cotransporter (NBC) (8); (b) the regulation of the expression and activity of cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-regulated chloride channel and channel regulator (9–13); (c) PTH 1 receptor signaling (14) as well as endocytic sorting of the 2-adrenergic receptor (15) and platelet-derived growth factor receptor (PDGFR) (16,17); (d) epithelial cell polarity and formation of tight junctions (18,19); and (e) maintaining the integrity of the glomerular barrier to proteins through interactions with podocalyxin, negatively charged sialoprotein expressed on the surface of the glomerular visceral epithelial cells or podocytes (20–25). In addition to their interaction with membrane proteins and receptors, the PDZ domain-containing proteins also interact with F-actin cytoskeleton through their interactions with the ezrinradixin-moesin (ERM) proteins (26). ERM proteins are typically located peripherally in the membrane and link the cytoplasmic tails of membrane proteins and receptors to the cortical actin cytoskeleton. The ERM proteins play an important role in the formation of microvilli, cell-cell junctions, and membrane ruffles and also participate in signal transduction pathways. The ERM proteins contain an F-actin binding site within their carboxyterminal 30 residues. In addition, the ERM proteins have a FERM (four-point one, ezrin, radixin, moesin) domain, which are generally located at or near the amino terminal and act as multifunctional protein and lipid binding sites. The FERM domain of ezrin interacts strongly with NHERF-1 and NHERF-2. NHERF-1 and NHERF-2 have two PDZ domains and have a carboxy-terminal sequence of 30 amino acids that bind ezrin. NHERF-1 (the sodium-hydrogen exchanger regulatory factor-1, also called EBP50) was isolated initially as a co-factor necessary in PTH and cAMP-induced inhibition of NHE3 (27–29). A second member of this family NHERF-2 (also called E3KARP) was also subsequently cloned. NHERF-1 and NHERF-2 are docking proteins that assemble multi-protein signaling complexes, including ezrin, NHE3, and PKA, through their multiple PDZ domains. These protein-signaling Correspondence to Dr, Moshe Levi, Professor of Medicine, Physiology and Biophysics, Head Division of Renal Diseases and Hypertension, University of Colorado Health Sciences Center, 4200 East 9th Avenue, Denver, Colorado 80262. Phone: 303-329-5415 or 5416; Fax: 303-393-4611; E-mail: [email protected]