Vasopressin regulates urea transport by activating two cyclic AMP (cAMP) dependent signaling pathways: protein kinase A and exchange protein activated by cAMP. This results in increases of both the phosphorylation and apical plasma membrane accumulation of the UT-A1 urea transporter. The hypertonicity present in the inner medulla can act independently of vasopressin as a powerful stimulus of urea transport. Hypertonicity stimulates urea transport by increasing both UT-A1 phosphorylation and plasma membrane accumulation. However, the signaling pathway by which this occurs is unknown. We have new preliminary data showing that hypertonicity stimulates urea permeability via protein kinase C (PKC). We also have preliminary data suggesting that PKC1 is the specific PKC isozyme involved since: 1) hypertonicity activates PKC1 in rat inner medullary collecting ducts (IMCDs);and 2) PKC1 knock-out mice have a urine concentrating defect and a reduction in UT-A1 protein abundance. Thus, maximal stimulation of urea transport and UT-A1 activity in the terminal IMCD requires stimulation by both cAMP and PKC1. Our overall goal is to investigate the regulation of UT-A1 by PKC. We will test the hypothesis that PKC1 stimulates UT-A1 function through changes in UT-A1 phosphorylation.
Aim 1 will determine the mechanism by which PKC regulates UT-A1 phosphorylation.
Aim 2 will determine the site(s) in UT-A1 and the cellular location of phosphorylation by PKC.
Aim 3 will determine the interdependence of PKC and PKA in the regulation of UT-A1 membrane accumulation, activity, phosphorylation, and phosphatase-mediated dephosphorylation.
Aim 4 will determine the mechanism for the urine concentrating defect in PKC" knock-out mice. Our proposed studies are highly significant as they are likely to yield new information on mechanisms underlying dysregulation of water homeostasis. Elucidation of non-cAMP mechanisms for increasing urea transport, which in turn would increase urine concentrating ability, could form the basis for future translational studies of novel therapeutic approaches to congenital nephrogenic diabetes insipidus.

Public Health Relevance

Urea plays a critical role in the urinary concentrating mechanism, and therefore, in the regulation of water balance. Our studies into the regulation of urea transport will advance our understanding of the urinary concentrating mechanism. This could yield new therapies for nephrogenic diabetes insipidus and will become increasingly important with the development of protein kinase C inhibitors for the treatment of diabetes mellitus.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK089828-03
Application #
8516032
Study Section
Cellular and Molecular Biology of the Kidney Study Section (CMBK)
Program Officer
Ketchum, Christian J
Project Start
2011-07-01
Project End
2015-06-30
Budget Start
2013-07-01
Budget End
2014-06-30
Support Year
3
Fiscal Year
2013
Total Cost
$325,326
Indirect Cost
$115,438
Name
Emory University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
066469933
City
Atlanta
State
GA
Country
United States
Zip Code
30322
Wang, Hong; Ferraris, Joan D; Klein, Janet D et al. (2015) PKC-? contributes to high NaCl-induced activation of NFAT5 (TonEBP/OREBP) through MAPK ERK1/2. Am J Physiol Renal Physiol 308:F140-8
Hoban, Carol A; Black, Lauren N; Ordas, Ronald J et al. (2015) Vasopressin regulation of multisite phosphorylation of UT-A1 in the inner medullary collecting duct. Am J Physiol Renal Physiol 308:F49-55
Weiner, I David; Mitch, William E; Sands, Jeff M (2015) Urea and Ammonia Metabolism and the Control of Renal Nitrogen Excretion. Clin J Am Soc Nephrol 10:1444-58
Li, Xuechen; Yang, Baoxue; Chen, Minguang et al. (2015) Activation of protein kinase C-? and Src kinase increases urea transporter A1 ?-2, 6 sialylation. J Am Soc Nephrol 26:926-34
Bao, Hui-Fang; Thai, Tiffany L; Yue, Qiang et al. (2014) ENaC activity is increased in isolated, split-open cortical collecting ducts from protein kinase C* knockout mice. Am J Physiol Renal Physiol 306:F309-20
Ren, Huiwen; Wang, Yanhua; Xing, Yongning et al. (2014) Thienoquinolins exert diuresis by strongly inhibiting UT-A urea transporters. Am J Physiol Renal Physiol 307:F1363-72
Sands, Jeff M; Blount, Mitsi A (2014) Genes and proteins of urea transporters. Subcell Biochem 73:45-63
Sands, Jeff M; Blount, Mitsi A (2014) Novel activators of aquaporin 2 membrane expression for the treatment of nephrogenic diabetes insipidus: less is more. Focus on "High-throughput chemical screening identifies AG-490 as a stimulator of aquaporin 2 membrane expression and urine concentratio Am J Physiol Cell Physiol 307:C595-6
Sands, Jeff M; Layton, Harold E (2014) Advances in understanding the urine-concentrating mechanism. Annu Rev Physiol 76:387-409
Du, Jie; Klein, Janet D; Hassounah, Faten et al. (2014) Aging increases CCN1 expression leading to muscle senescence. Am J Physiol Cell Physiol 306:C28-36

Showing the most recent 10 out of 17 publications