Cardiovascular and Renal Actions of the PDZ protein SAP97

Abstract

Synapse Associated Protein-97 (SAP97) is a PDZ protein that is expressed in the CNS as well as in peripheral organs. SAP97 is a major compartmentalizing protein involved in organizing signaling complexes. We investigated the role of SAP97 in anterograde translocation of beta1-adrenergic receptors (beta1-AR) in adult rat ventricular myocytes (ARVM) and in translocation of aquaporin-2 forming channels in kidney cells. Interactions between SAP97 and other proteins are mediated by binding of one of its three PDZ domains with type-1 PDZ binding motifs in target proteins. We determined that SAP97 forms a ternary complex with the beta1-AR and with A-kinase anchoring protein 79/150 (AKAP5)-PKA, which targets the beta1-AR to AKAP-PKA. Activation of the beta1-AR by agonists promotes induces PKA phosphorylation of the beta1-AR at serine312 in the third intracellular loop and its internalization to endosomes. This phospho-serine along with the type- PDZ binding motif in the C-tail of the beta1-AR serve as trafficking barcodes which promote the recycling of the agonist-internalized beta1-AR back to the membrane. Similarly, SAP97 binds to AQP2 via its type1-PDZ binding motif and to an unidentified AKAP-PKA complex. Exposing renal collecting cells to vasopressin, promotes the phosphorylation of AQP2 on serine256 in the C-tail, which is a primary signal for translocating cytoplasmic AQP2 storage vesicles to the apical membrane. These two pathways were dependent on targeting these trafficking proteins to an AKAP-PKA complex via SAP97. In this regard we verified that anterograde translocation of beta1-AR and AQP2 was dependent on their type-1 PDZs, and on the SAP97-anchored complex to faithfully phosphorylate a specific trafficking serine. In addition, we determined that SAP97 co-localized with beta1-AR in ARVM and with AQP2 in the kidneys, and knockdown of SAP97 inhibited anterograde translocation of these proteins, without affecting their retrograde internalization. Here in we describe these pathways and will show that SAP97 provides a novel paradigm to decipher trafficking events localized around cis-acting barcodes and their trans-acting proteins that might ultimately be used for therapeutic advantage.