Interaction of the type IIa Na/P-i cotransporter with PDZ proteins

The type IIa Na+-dependent inorganic phosphate (Na/P-i) cotransporter is lo calized in the apical membrane of proximal tubular cells and is regulated b y an endocytotic pathway. Because molecular processes such as apical sortin g, internalization, or subsequent degradation might be assisted by associat ed proteins, a yeast two-hybrid screen against the C-terminal, cytosolic ta il of type IIa cotransporter was designed. Most of the potential proteins f ound belonged to proteins with multiple PDZ modules and were either identic al/related to PDZK1 or identical to NHERF-1. Yeast trap truncation assays c onfined the peptide-protein association to the C-terminal amino acid residu es TRL of type IIa cotransporter and to single PDZ domains of each identifi ed protein, respectively. The specificity of these interactions were confir med in yeast by testing other apical localized transmembraneous proteins. M oreover, the type IIa protein was recovered in vitro by glutathione S-trans ferase-fused PDZ proteins from isolated renal brush border membranes or fro m type IIa-expressing oocytes. Further, these PDZ proteins are immunohistoc hemically detected either in the microvilli or in the subapical compartment of proximal tubular cells. Our results suggest that the type IIa Na/P-i co transporter interacts with various PDZ proteins that might be responsible f or the apical sorting, parathyroid hormone controlled endocytosis or the ly sosomal sorting of internalized type IIa cotransporter.