Vasopressin increases plasma membrane accumulation of urea transporter UT-A1 in rat inner medullary collecting ducts

Emory University School of Medicine, Renal Division, WMB Room 3319B, 1639 Pierce Drive NE, Atlanta, GA 30322, USA.
Journal of the American Society of Nephrology (Impact Factor: 9.47). 11/2006; 17(10):2680-6. DOI: 10.1681/ASN.2006030246
Source: PubMed

ABSTRACT Urea transport, mediated by the urea transporter A1 (UT-A1) and/or UT-A3, is important for the production of concentrated urine. Vasopressin rapidly increases urea transport in rat terminal inner medullary collecting ducts (IMCD). A previous study showed that one mechanism for rapid regulation of urea transport is a vasopressin-induced increase in UT-A1 phosphorylation. This study tests whether vasopressin or directly activating adenylyl cyclase with forskolin also increases UT-A1 accumulation in the plasma membrane of rat IMCD. Inner medullas were harvested from rats 45 min after injection with vasopressin or vehicle. UT-A1 abundance in the plasma membrane was significantly increased in the membrane fraction after differential centrifugation and in the biotinylated protein population. Vasopressin and forskolin each increased the amount of biotinylated UT-A1 in rat IMCD suspensions that were treated ex vivo. The observed changes in the plasma membrane are specific, as the amount of biotinylated UT-A1 but not the calcium-sensing receptor was increased by forskolin. Next, whether forskolin or the V(2)-selective agonist dDAVP would increase apical membrane expression of UT-A1 in MDCK cells that were stably transfected with UT-A1 (UT-A1-MDCK cells) was tested. Forskolin and dDAVP significantly increased UT-A1 abundance in the apical membrane in UT-A1-MDCK cells. It is concluded that vasopressin and forskolin increase UT-A1 accumulation in the plasma membrane in rat IMCD and in the apical plasma membrane of UT-A1-MDCK cells. These findings suggest that vasopressin regulates urea transport by increasing UT-A1 accumulation in the plasma membrane and/or UT-A1 phosphorylation.

  • [Show abstract] [Hide abstract]
    ABSTRACT: Vasopressin signaling is critical for regulation of urea transport in the inner medullary collecting duct (IMCD). Increased urea permeability is driven by a vasopressin-mediated elevation of cAMP that results in the direct phosphorylation of the urea transporter, UT-A1. Identification of the cAMP-sensitive phosphorylation sites, S486 and S499, in the rat UT-A1 sequence was the first step in understanding the mechanism of vasopressin action on the phosphorylation-dependent modulation of urea transport. To investigate the significance of multisite phosphorylation of UT-A1 in response to elevated cAMP, we employed highly specific and sensitive phosphosite antibodies to S486 and S499 to determine cAMP action at each phosphorylation site. We found that phosphorylation at both sites was rapid and sustained. Furthermore, the rate of phosphorylation of the two sites was similar in both mIMCD3 cells and rat inner medullary tissue. The UT-A1 localized to the apical membrane in response to vasopressin was phosphorylated at S486 and S499. We confirmed that elevated cAMP resulted in increased phosphorylation of both sites by PKA but not through the vasopressin-sensitive Epac pathway. These results elucidate the multisite phosphorylation of UT-A1 in response to cAMP thus providing the beginning of understanding the intracellular factors underlying vasopressin stimulation of urea transport in the IMCD.
    American journal of physiology. Renal physiology 11/2014; 308(1):ajprenal.00642.2013. DOI:10.1152/ajprenal.00642.2013 · 3.30 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Aquaporins (AQPs) are a family of membrane water channels that basically function as regulators of intracellular and intercellular water flow. To date, thirteen aquaporins have been characterized. They are distributed wildly in specific cell types in multiple organs and tissues. Each AQP channel consists of six membrane-spanning alpha-helices that have a central water-transporting pore. Four AQP monomers assemble to form tetramers, which are the functional units in the membrane. Some of AQPs also transport urea, glycerol, ammonia, hydrogen peroxide, and gas molecules. AQP-mediated osmotic water transport across epithelial plasma membranes facilitates transcellular fluid transport and thus water reabsorption. AQP-mediated urea and glycerol transport is involved in energy metabolism and epidermal hydration. AQP-mediated CO2 and NH3 transport across membrane maintains intracellular acid-base homeostasis. AQPs are also involved in the pathophysiology of a wide range of human diseases (including water disbalance in kidney and brain, neuroinflammatory disease, obesity, and cancer). Further work is required to determine whether aquaporins are viable therapeutic targets or reliable diagnostic and prognostic biomarkers.
    Sub-cellular biochemistry 01/2014; 73:227-65. DOI:10.1007/978-94-017-9343-8_14
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Marine elasmobranchs maintain their body fluid isoosmotic or slightly hyperosmotic to the exter-nal medium by the retention of large urea con-centrations. This review focuses on the strate-gies adopted by these fishes to maintain a large outwardly direct concentration gradient of this osmolyte minimizing the loss across the main interfaces between body fluid and the external medium such as the gills, the kidney and the rectal gland, thus reducing the cost of making urea. The high plasma osmolarity, mainly main-tained by urea retention, is a challenge to vol-ume homeostasis when fish move from sea-water to water with a low salinity, since the high water permeability of branchial epithelium would cause a net flux of water into the animal. Since the renal regulation of urea retention in habitat with different salinities is crucial for the osmotic homeostasis of these species, the regulation of the activity and/or the expression of urea trans-porters in renal tubules will be also discussed. In addition attention will be paid on the urea– methylamine system involved in maintaining the stability and functioning of many proteins since it is known that the high urea concentration found in marine elasmobranch fish, similar only to that found in mammalian kidney, has a desta-bilizing effect on many macromolecules and inhibits functions such as ligand binding.


Available from