Renal type A intercalated cells (A-ICs) actively secrete protons and are key players in the regulation of blood pH. However, the mechanisms by which A-ICs detect sysmetic pH variations are poorly understood. Proton secretion is achieved by the V-ATPase, and severe diseases such as distal renal tubular acidosis and kidney stones result from dysfunctional V-ATPase. While searching for potential acid/base sensors, we found that A-ICs and similar proton-secreting cells in the epididymis (clear cells;CCs) sense luminal bicarbonate via sAC, a bicarbonate-activated adenylyl cyclase. This proposal now examines how A-ICs sense urinary pH in addition to bicarbonate to regulate proton secretion. Some purinergic receptors were proposed as pH sensors, and Aim 1 will test the hypothesis that A-ICs sense urinary pH variations via these receptors. The proposed experiments are based on our new data showing that luminal ATP stimulates proton secretion by CCs. We will identify P1 and P2 receptors that are exclusively expressed in A-ICs, and we will examine their role in regulating V-ATPase-dependent proton secretion. We will also examine ATP secretion by A-ICs and the role of CFTR in this process. A-ICs isolated by FACS from B1-EGFP mice, the C11 intercalated cell line, and microdissected OMCDs will be examined using complementary imaging, biochemical and electrophysiological procedures.
Aim 2 will explore the possibility that the V- ATPase itself might be part of the pH-sensing property of A-ICs. These experiments are based on exciting data showing that the V-ATPase a2 subunit is a pH sensor involved in intracellular targeting events. We will examine whether the """"""""plasma membrane"""""""" a4 is a pH sensor that regulates V-ATPase via association of its cytosolic V1 with its transmembrane V0 domains. We will use the epididymis as a model system in which luminal pH can be modulated in vivo. V-ATPase assembly/disassembly will be examined using confocal microscopy, EM, co-IP and cell fractionation. We will """"""""translate"""""""" our findings to A-ICs by using FACS isolated A-ICs and OMCDs perfused in vitro. These experiments will contribute to our better understanding of renal luminal acidification, which is central to the maintenance of blood pH within a very narrow viable range.

Public Health Relevance

One of the main functions of the kidney is to acidify the urine to control blood pH. Malfunction of this process leads to a life threatening disorder called acidosis, and the formation of kidney stones, which cause severe damage to the kidney architecture. We study how specialized kidney cells detect pH variations in our body and adapt their rate of acid secretion to maintain blood pH within a very narrow viable range.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
1R01DK097124-01
Application #
8419239
Study Section
Special Emphasis Panel (ZRG1-DKUS-L (03))
Program Officer
Ketchum, Christian J
Project Start
2012-12-15
Project End
2016-11-30
Budget Start
2012-12-15
Budget End
2013-11-30
Support Year
1
Fiscal Year
2013
Total Cost
$378,450
Indirect Cost
$160,950
Name
Massachusetts General Hospital
Department
Type
DUNS #
073130411
City
Boston
State
MA
Country
United States
Zip Code
02199
Kim, Bongki; Breton, Sylvie (2016) The MAPK/ERK-Signaling Pathway Regulates the Expression and Distribution of Tight Junction Proteins in the Mouse Proximal Epididymis. Biol Reprod 94:22
Roy, Jeremy; Kim, Bongki; Hill, Eric et al. (2016) Tyrosine kinase-mediated axial motility of basal cells revealed by intravital imaging. Nat Commun 7:10666
Breton, Sylvie; Ruan, Ye Chun; Park, Yoo-Jin et al. (2016) Regulation of epithelial function, differentiation, and remodeling in the epididymis. Asian J Androl 18:3-9
Azroyan, Anie; Cortez-Retamozo, Virna; Bouley, Richard et al. (2015) Renal intercalated cells sense and mediate inflammation via the P2Y14 receptor. PLoS One 10:e0121419
Kim, Bongki; Roy, Jeremy; Shum, Winnie W C et al. (2015) Role of testicular luminal factors on Basal cell elongation and proliferation in the mouse epididymis. Biol Reprod 92:9
Merkulova, Maria; Păunescu, Teodor G; Azroyan, Anie et al. (2015) Mapping the H(+) (V)-ATPase interactome: identification of proteins involved in trafficking, folding, assembly and phosphorylation. Sci Rep 5:14827
Ruan, Ye Chun; Wang, Yan; Da Silva, Nicolas et al. (2014) CFTR interacts with ZO-1 to regulate tight junction assembly and epithelial differentiation through the ZONAB pathway. J Cell Sci 127:4396-408
Păunescu, Teodor G; Shum, Winnie W C; Huynh, Chuong et al. (2014) High-resolution helium ion microscopy of epididymal epithelial cells and their interaction with spermatozoa. Mol Hum Reprod 20:929-37
Roy, Jeremy W; Hill, Eric; Ruan, Ye Chun et al. (2013) Circulating aldosterone induces the apical accumulation of the proton pumping V-ATPase and increases proton secretion in clear cells in the caput epididymis. Am J Physiol Cell Physiol 305:C436-46
Breton, Sylvie; Brown, Dennis (2013) Regulation of luminal acidification by the V-ATPase. Physiology (Bethesda) 28:318-29

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