Forskolin stimulates phosphorylation and membrane accumulation of UT-A3

• Published in: American journal of physiology. Renal physiology 2007-10, Vol.293 (4), p.F1308-F1313
• Main Authors: Blount, Mitsi A, Klein, Janet D, Martin, Christopher F, Tchapyjnikov, Dmitry, Sands, Jeff M
• Format: Article
• Language: English
• Subjects: Animals; Antidiuretic Agents - pharmacology; Cell Membrane - metabolism; Cells; Colforsin - pharmacology; Kidney Medulla - cytology; Kidney Medulla - metabolism; Kidney Tubules, Collecting - cytology; Kidney Tubules, Collecting - metabolism; Kidneys; Membrane Transport Proteins - metabolism; Membranes; Phosphorylation - drug effects; Proteins; Rats; Rats, Sprague-Dawley; Rodents; Studies; Urea Transporters; Vasopressins - pharmacology
• ISSN: 1931-857X; 1522-1466
• DOI: 10.1152/ajprenal.00197.2007

UT-A1 is regulated by vasopressin and is localized to the apical membrane and intracellular compartment of inner medullary collecting duct (IMCD) cells. UT-A3 is also expressed in the IMCD and is regulated by forskolin in heterologous systems. The goal of the present study is to investigate mechanisms by which vasopressin regulates UT-A3 in rat IMCD. In fresh suspensions of rat IMCD, forskolin increases the phosphorylation of UT-A3, similar to UT-A1. Biotinylation studies indicate that UT-A3 is located in the plasma membrane. Forskolin treatment increases the abundance of UT-A3 in the plasma membrane similar to UT-A1. However, these two transporters do not form a complex through a protein-protein interaction, suggesting that transporter function is unique to each protein. While immunohistochemistry localized UT-A3 to the basal and lateral membranes, a majority of the staining was cytosolic. Immunohistochemistry of vasopressin-treated rat kidney sections also localized UT-A3 primarily to the cytosol with basal and lateral membrane staining but also showed some apical membrane staining in some IMCD cells. This suggests that under normal conditions, UT-A3 functions as the basolateral transporter but in a high cAMP environment, the transporter may move from the cytosol to all plasma membranes to increase urea flux in the IMCD. In summary, this study confirms that UT-A3 is located in the inner medullary tip where it is expressed in the basolateral membrane, shows that UT-A3 is a phosphoprotein in rat IMCD that can be trafficked to the plasma membrane independent of UT-A1, and suggests that vasopressin may induce UT-A3 expression in the apical plasma membrane of IMCD.