Na/K-ATPase belongs to the family of P-type ATPases and was discovered as an energy transducing ion pump. A major difference between the Na/K-ATPase and other P-type ATPases is its ability to bind cardiotonic steroids (CIS) such as ouabain. While endogenous CTS regulate blood pressure via their effects on vasculature and renal salt handling, the plant-derived CTS have been used as drugs for more than 200 years. Recently, we have demonstrated that the Na/K-ATPase is an important receptor that resides in caveolae and interacts directly with Src, a non-receptor tyrosine kinase. We know now that the effects of low doses of ouabain on many cellular functions are not due to the simple inhibition of the ATPase; rather they require the activation of the Na/K-ATPase/Src receptor complex. In addition, the Na/K-ATPase contains multiple binding motifs (domains) and is capable of bringing Src and other signaling enzymes to their effectors such as ion channels. These findings have led the research field to look at the Na/K-ATPase not only as an ion pump, but also a classical receptor complex. This shift of the paradigm has brought about an important question: which regulatory purposes does this signaling Na/K-ATPase serve in regulation of cellular functions that are relevant to the physiology of endogenous CTS (e.g. blood pressure control)? This application is proposed to bridge this gap by studying the most likely target of this receptor complex, namely the Ca2+-signaling module because changes in intracellular Ca2+ are known to play a key role in regulation of vascular function and renal salt handling. Specifically, we will investigate how the Na/K-ATPase integrates multiple constituents into a functional Ca2+-signaling module in renal epithelial cells. We propose to use a combined biochemical, cellular, genetic and dynamic imaging approach to 1) define the molecular mechanism by which the Na/K-ATPase integrates Src/PLC-y/PKC and IPS receptor into a dynamic Ca2+ signaling module; 2) reveal whether disruption of the interaction between the Na/K-ATPase and IPS receptors affects IPS receptor trafficking and ouabain-induced Ca2+ signaling; And 3) identify the plasma membrane channel (s) that interacts with the Na/K-ATPase and is responsible for ouabain-induced Ca2+ influx. These studies will not only relate the newly discovered receptor function of the Na/K-ATPase to renal physiology of CTS, but also provide detailed mechanistic information on the formation of a Ca2+-signaling module that will eventually give us a new target for developing therapeutic approaches to renal and cardiovascular diseases involving dysfunction of intracellular Ca regulation. Calcium is a universal second messenger that plays an essential role in control of kidney and cardiovascular function. Abnormality in intracellular calcium regulation will lead to both kidney and heart diseases such as hypertension. We are using a simple model to dissect the formation of a very important calcium controlling system in kidney cells and to investigate how we can manipulate this system to eventually develop new approaches to prevent renal and cardiovascular diseases. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM078565-02
Application #
7457662
Study Section
Special Emphasis Panel (ZRG1-RUS-A (02))
Program Officer
Chin, Jean
Project Start
2007-07-01
Project End
2011-06-30
Budget Start
2008-07-01
Budget End
2009-06-30
Support Year
2
Fiscal Year
2008
Total Cost
$323,661
Indirect Cost
Name
University of Toledo
Department
Physiology
Type
Schools of Medicine
DUNS #
807418939
City
Toledo
State
OH
Country
United States
Zip Code
43614
Ye, Qiqi; Lai, Fangfang; Banerjee, Moumita et al. (2013) Expression of mutant ?1 Na/K-ATPase defective in conformational transition attenuates Src-mediated signal transduction. J Biol Chem 288:5803-14
Yan, Yanling; Haller, Steven; Shapiro, Anna et al. (2012) Ouabain-stimulated trafficking regulation of the Na/K-ATPase and NHE3 in renal proximal tubule cells. Mol Cell Biochem 367:175-83
Gupta, Shalini; Yan, Yanling; Malhotra, Deepak et al. (2012) Ouabain and insulin induce sodium pump endocytosis in renal epithelium. Hypertension 59:665-72
Liu, Jiang; Yan, Yanling; Liu, Lijun et al. (2011) Impairment of Na/K-ATPase signaling in renal proximal tubule contributes to Dahl salt-sensitive hypertension. J Biol Chem 286:22806-13
Chen, Yiliang; Li, Xin; Ye, Qiqi et al. (2011) Regulation of alpha1 Na/K-ATPase expression by cholesterol. J Biol Chem 286:15517-24
Li, Zhichuan; Zhang, Zhongbing; Xie, Joe X et al. (2011) Na/K-ATPase mimetic pNaKtide peptide inhibits the growth of human cancer cells. J Biol Chem 286:32394-403
Ye, Qiqi; Li, Zhichuan; Tian, Jiang et al. (2011) Identification of a potential receptor that couples ion transport to protein kinase activity. J Biol Chem 286:6225-32
Zhang, Zhongbing; Li, Zhichuan; Tian, Jiang et al. (2010) Identification of hydroxyxanthones as Na/K-ATPase ligands. Mol Pharmacol 77:961-7
Quintas, Luis E M; Pierre, Sandrine V; Liu, Lijun et al. (2010) Alterations of Na+/K+-ATPase function in caveolin-1 knockout cardiac fibroblasts. J Mol Cell Cardiol 49:525-31
Liu, Jiang; Xie, Zi-Jian (2010) The sodium pump and cardiotonic steroids-induced signal transduction protein kinases and calcium-signaling microdomain in regulation of transporter trafficking. Biochim Biophys Acta 1802:1237-45

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