Ion Channels – Membrane Transport – Integrative Physiology
Kidney International (2003) 64, 1733–1745; doi:10.1046/j.1523-1755.2003.00266.x
PDZK1: I. A major scaffolder in brush borders of proximal tubular cells1
Serge M Gisler, Sandra Pribanic, Desa Bacic, Patrik Forrer, Andrea Gantenbein, Luc A Sabourin, Akira Tsuji, Zhuo-Shen Zhao, Edward Manser, Jürg Biber and Heini Murer
Department of Physiology and Biochemistry, University of Zürich, Zürich, Switzerland; Neuroscience Research Institute and Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, Canada; Faculty of Pharmaceutical Sciences, Kanazawa University, Takara-machi, Kanazawa, Japan; and Glaxo-IMCB Laboratory, Institute of Molecular and Cell Biology, Singapore
Correspondence: Dr Jürg Biber, Institute of Physiology, University of Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland. E-mail: JuergBiber@access.unizh.ch
1See Editorial by Moe, p. 1916.
Received 23 January 2003; Revised 4 April 2003; Accepted 23 June 2003.
Abstract
PDZK1: I. A major scaffolder in brush borders of proximal tubular cells.
Background
In proximal tubular cells, PDZK1 (NaPi-Cap1) has been implicated in apical expression of the Na+-dependent phosphate cotransporter (NaPi-IIa) via interaction with its C-terminus. PDZK1 represents a multidomain protein consisting of four PDZ domains and thus is believed to have a broader specificity besides NaPi-IIa.
Methods
We subjected single PDZ domains derived from PDZK1 either to yeast two-hybrid screens or yeast trap assays. Different pull-down assays and blot overlays were applied to corroborate the PDZK1-mediated interactions in vitro. Co-localization of interacting proteins with PDZK1 in proximal tubular cells was assessed by immunohistochemistry.
Results
In the yeast screens, the most abundant candidate protein to interact with PDZK1 was the membrane-associated protein of 17 kD (MAP17). Besides MAP17, C-terminal parts of following transporters were also identified: NaPi-IIa, solute carrier SLC17A1 (NaPi-I), Na+/H+ exchanger (NHE-3), organic cation transporter (OCTN1), chloride-formate exchanger (CFEX), and urate-anion exchanger (URAT1). In addition, other regulatory factors were found among the clones, such as a protein kinase A (PKA)-anchoring protein (D-AKAP2) and N+/H+ exchanger regulator factor (NHERF-1). All interactions of itemized proteins with PDZK1 were affirmed by in vitro techniques. Apart from PDZK1, strong in vitro interactions of NHERF-1 were also observed with the solute transporters (excluding MAP17) and D-AKAP2. All identified proteins were immunolocalized in proximal tubular cells, wherein all membrane proteins co-localized with PDZK1 in brush borders.
Conclusion
We hypothesize that PDZK1 and NHERF-1 establish an extended network beneath the apical membrane to which membrane proteins and regulatory components are anchored.
Keywords:
renal transport of phosphate, NaPi-IIa, NaPi-I, PDZK1, NHERF-1, D-AKAP2, PDZ proteins, MAP17, URAT1, CFEX, yeast two-hybrid
