Vasopressin (VP) binds to the vasopressin type 2 receptor (V2R) to trigger physiological effects including body fluid homeostasis and blood pressure regulation. Signaling is terminated by receptor downregulation involving clathrin-mediated endocytosis and V2R degradation. We report here that both native and epitope-tagged V2R is internalized from the plasma membrane of LLC-PK1 kidney epithelial cells in the presence of another ligand, transferrin (Tf). The presence of iron-saturated Tf (holo-Tf) (4 h) reduced V2R binding sites at the cell surface by up to 33% while iron-free (apo-Tf) had no effect. However, no change in V2R-GFP distribution was observed in the presence of bovine serum albumin, atrial natriuretic peptide or angiotensin II. In contrast to the effect of VP, Tf did not increase intracellular cAMP or modify aquaporin type 2 (AQP2) distribution in these cells, although addition of VP and Tf together augmented VP-induced V2R internalization. Tf receptor co-immunoprecipitated with V2R, suggesting that they interact closely and this may explain the additive effect of VP and Tf on V2R endocytosis. Furthermore, Tf-induced V2R internalization was abolished in cells expressing a dominant negative dynamin (K44A) mutant, indicating the involvement of clathrin coated pits. We conclude that Tf can induce heterologous downregulation of the V2R and this might desensitize VP target cells without activating downstream V2R signaling events. It also provides new insights into urine concentrating defects observed in rat models of hemochromatosis.
- G-protein coupled receptor
- antidiurectic hormone
- kidney epithelial cells
- Copyright © 2011, American Journal of Physiology - Renal Physiology