The long-term objective of our work is directed toward a complete understanding of the cellular mechanisms and regulation of calcium transport by renal tubular epithelial cells. Although most calcium absorption proceeds in proximal tubules, distal tubules are the site of the physiological regulation of calcium transport by parathyroid hormone (PTH), calcitonin and vitamin D3.
The specific aims of the present proposal are to: 1) characterize the mechanism of calcium entry across apical plasma membranes of distal convoluted tubule (DCT) cells; 2) evaluate the participation and regulation of Na+/Ca2+ exchange in mediating calcium efflux; and, 3) define the signaling pathways activated by PTH and calcitonin in DCT cells, and to examine the coordinate regulation of PTH-dependent calcium transport by 1,25(OH)2 vitamin D3 and estradiol. We identified and partially characterized a novel calcium channel that mediates calcium entry. To acquire additional information about these channels we will: 1) characterize the properties of these channels with regard to their ion selectivity, pharmacologic sensitivity and voltage- dependence; 2) determine the regulation of these calcium channels by protein kinases; 3) define the participation of G-proteins in regulating calcium entry channels; and, 4) identify and clone calcium channel transcripts. Extrusion of calcium across basolateral plasma membranes of distal tubules is mediated by Na+/Ca2+ exchange and by /ca2+-ATPase. Pilot studies suggest that Na+/Ca2+ exchange is the dominant efflux mechanism in DCT cells. We will: 1) Test the hypothesis that thiazide diuretics can inhibit Na+/Ca2+ exchange; and 2) Characterize the stimulatory effects of PTH on Na+/Ca2+ exchange. Calcium transport in distal nephrons is regulated by PTH, calcitonin and 1,25(OH)2 vitamin D3. The following studies will evaluate the mechanism mediating the hormonal regulation of calcium transport in DCT cells. The goals of these experiments are to: 1) Characterize the signaling pathways with particular regard to the phospholipases responsible for activating PKA and PKC and characterize the temporal sequence in which PKA and PKC are activated. We will test the hypothesis that PTH and calcitonin activate PKC via phospholipase D. 2) Identify the mechanism by which PTH activates C1- channels, a primary event in membrane hyperpolarization and stimulation of calcium transport in DCT cells. We will test the hypothesis that these chloride channels are regulated by the PKA limb of the PTH-activated signaling pathway. 3) Examine the regulation of vitamin D3 and estradiol accelerate PTH dependent calcium transport by up-regulating PTH receptor expression.
The specific aims will be achieved by applying single cell fluorescence, patch clamp, tracer flux measurements, biochemical and molecular techniques to a DCT cell line that we developed, which expresses an appropriate phenotype. In selected studies, primary cell cultures of proximal or distal tubule cells will be used to verify results in transformed cells or as controls. All procedures are established in our lab. Results from the proposed experiments will provide new information on the mechanism and regulation of calcium transport in the kidney under normal conditions, but also in calcium-wasting syndromes including hyperparathyroidism, renal failure, osteoporosis and malignancy- associated hypercalcemia.
| Pavlos, Nathan J; Friedman, Peter A (2017) GPCR Signaling and Trafficking: The Long and Short of It. Trends Endocrinol Metab 28:213-226 |
| Pereira, Renata C; Andersen, Thomas L; Friedman, Peter A et al. (2016) Bone Canopies in Pediatric Renal Osteodystrophy. PLoS One 11:e0152871 |
| McGarvey, Jennifer C; Xiao, Kunhong; Bowman, Shanna L et al. (2016) Actin-Sorting Nexin 27 (SNX27)-Retromer Complex Mediates Rapid Parathyroid Hormone Receptor Recycling. J Biol Chem 291:10986-1002 |
| Walsh, Dustin R; Nolin, Thomas D; Friedman, Peter A (2015) Drug Transporters and Na+/H+ Exchange Regulatory Factor PSD-95/Drosophila Discs Large/ZO-1 Proteins. Pharmacol Rev 67:656-80 |
| Stubbs, Jason R; Zhang, Shiqin; Friedman, Peter A et al. (2014) Decreased conversion of 25-hydroxyvitamin D3 to 24,25-dihydroxyvitamin D3 following cholecalciferol therapy in patients with CKD. Clin J Am Soc Nephrol 9:1965-73 |
| Alonso, Veronica; Friedman, Peter A (2013) Minireview: ubiquitination-regulated G protein-coupled receptor signaling and trafficking. Mol Endocrinol 27:558-72 |
| Wheeler, D S; Barrick, S R; Grubisha, M J et al. (2011) Direct interaction between NHERF1 and Frizzled regulates ?-catenin signaling. Oncogene 30:32-42 |
| Liu, Li; Schlesinger, Paul H; Slack, Nicole M et al. (2011) High capacity Na+/H+ exchange activity in mineralizing osteoblasts. J Cell Physiol 226:1702-12 |
| Alonso, Verónica; Magyar, Clara E; Wang, Bin et al. (2011) Ubiquitination-deubiquitination balance dictates ligand-stimulated PTHR sorting. J Bone Miner Res 26:2923-34 |
| Blair, Harry C; Robinson, Lisa J; Huang, Christopher L-H et al. (2011) Calcium and bone disease. Biofactors 37:159-67 |
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