Paradigm-shifting observations on aquaporin 2 (AQP2) and vasopressin receptor (V2R) trafficking, and vacuolar ATPase (V-ATPase) pH sensing and recycling have lead to new hypotheses to be addressed in this renewal. Project I will define VP-dependent and independent regulation of AQP2, and will identify proteins whose interaction with AQP2 is modified by phosphorylation to regulate trafficking. A novel role for AQP2 in regulating actin polymerization via interaction with RhoGAPS will be explored. Use of PDE5 inhibitors and statins to achieve VP-independent urine concentration will be tested as a potential strategy for future treatment of NDI. Project II will examine ligand induced conformational changes of the V2R at varying pH and tonicity using FRET techniques to dissect intra- and intermolecular protein interactions. Association of V2R with accessory proteins during internalization will be defined in cells expressing wild type and mutant V2R. Our novel observation that V2R interacts with the ESCRT protein Alix to accelerate V2R degradation will be pursued in these studies that address the regulation of body fluid homeostasis. Project III will pursue the breakthrough finding that the V-ATPase is an endosomal pH sensor by defining conformational changes in the V-ATPase tail that result in pH-dependent recruitment of small GTPases to membranes. It will identify pH sensitive residues on the luminal domains of the V-ATPase, and use albumin uptake to show relevance of the pH sensing mechanism to proximal tubule function. Project IV will elucidate downstream effectors (PKA, Epac) of soluble adenylate cyclase in modulating V-ATPase recycling and proton secretion in the epididymis, a """"""""model"""""""" epithelium in which luminal acidic pH is critical for sperm maturation and storage. It will examine the role of cGMP-induced proton secretion in this tissue, and will address the exciting hypothesis that the V-ATPase is also an extracellular pH sensor that provides feedback control of luminal pH. These studies will allow a better understanding of male fertility, and uncover mechanisms that regulate V-ATPase in acidifying cells in general, including renal intercalated cells. The Microscopy Core B facility has been a major contributor to the success of this PPG. All projects gain considerable added value from extensive intellectual and technical collaborations that characterize our efforts to understand the relationship between cellular signaling, protein trafficking, and the responses of urogenital epithelial cells to their environment.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Program Projects (P01)
Project #
Application #
Study Section
Special Emphasis Panel (ZDK1-GRB-S (J1))
Program Officer
Mullins, Christopher V
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Massachusetts General Hospital
United States
Zip Code
Li, Wei; Jin, William W; Tsuji, Kenji et al. (2017) Ezrin directly interacts with AQP2 and promotes its endocytosis. J Cell Sci 130:2914-2925
Arthur, Julian; Huang, Jianmin; Nomura, Naohiro et al. (2015) Characterization of the putative phosphorylation sites of the AQP2 C terminus and their role in AQP2 trafficking in LLC-PK1 cells. Am J Physiol Renal Physiol 309:F673-9
Rice, William L; Li, Wei; Mamuya, Fahmy et al. (2015) Polarized Trafficking of AQP2 Revealed in Three Dimensional Epithelial Culture. PLoS One 10:e0131719
Zhang, Ping L; Mashni, Joseph W; Sabbisetti, Venkata S et al. (2014) Urine kidney injury molecule-1: a potential non-invasive biomarker for patients with renal cell carcinoma. Int Urol Nephrol 46:379-88
Marshansky, Vladimir; Rubinstein, John L; GrĂ¼ber, Gerhard (2014) Eukaryotic V-ATPase: novel structural findings and functional insights. Biochim Biophys Acta 1837:857-79
Hosokawa, Hiroyuki; Dip, Phat Vinh; Merkulova, Maria et al. (2013) The N termini of a-subunit isoforms are involved in signaling between vacuolar H+-ATPase (V-ATPase) and cytohesin-2. J Biol Chem 288:5896-913
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
P?unescu, Teodor G; Lu, Hua A J; Russo, Leileata M et al. (2013) Vasopressin induces apical expression of caveolin in rat kidney collecting duct principal cells. Am J Physiol Renal Physiol 305:F1783-95
Feinstein, Timothy N; Yui, Naofumi; Webber, Matthew J et al. (2013) Noncanonical control of vasopressin receptor type 2 signaling by retromer and arrestin. J Biol Chem 288:27849-60

Showing the most recent 10 out of 278 publications