Inherited mutations in SLC12A1 and SLC12A2, the genes encoding the renal electroneutral Na-Cl and Na-K-2Cl co-transporters, result in salt wasting disorders associated with changes in blood pressure. The syndromes, called Gittleman and Bartter, are due to decreased Na+-re-absorption in distal convoluted tubule and thick ascending limb of Henle, respectively. Two Sterile20 kinase, SPAK, and OSR1, are involved in regulating the co-transporters. The kinase directly binds to the N-terminal tails of the co-transporter and phosphorylates them. A whole-genome study found an association between SPAK and increased blood pressure, and SPAK knockout mice exhibit a Gittleman-like phenotype. Surprisingly, NKCC2 phosphorylation is increased in SPAK knockout animals, but is blunted in phosphorylation-deficient knock-in SPAK mice, suggesting a possible interaction between SPAK and OSR1 in the thick ascending limb. The purpose of this 2 PD/PI application is to understand the molecular details leading to increased Na-K-2Cl co-transporter phosphorylation in the SPAK knockout and to define the role of each kinase in modulating Na+-re-absorption in the two tubule segments. In this application, we propose to 1) examine the mechanisms of SPAK/OSR1 activation, determine the role of dimerization, and establish the existence of SPAK-OSR1 interactions;2) examine the function of putative inhibitory isoforms and of Cab39, a scaffold calcium binding protein;and 4) establish the role of SPAK and OSR1 in regulating renal salt transport and maintaining blood pressure. The first two aims will involve functional studies using heterologous expression in both Xenopus laevis oocytes and mammalian kidney cells, as well as molecular studies involving protein-protein interaction both in vitro and in vivo.
The third aim will consist of detailed phenotypic analysis of genetically-modified mice under different diet regiments, and the development and study of a novel constitutively active OSR1 knock-in mouse. This application is a logical extension of extensive studies of cation-chloride co-transporters and their regulation by the submitting PI and a natural extension of a successful ongoing AARA-funded Challenge grant collaboration between the two PD/PIs. Upon completion, the proposed studies will clarify the role of two kinase and demonstrate that they are integrated into precise signaling networks in the thick ascending limb of Henle and the distal convoluted tubule.

Public Health Relevance

The proposed research will address the role and interdependency of two related kinases, SPAK and OSR1, in the modulation of transport mechanisms involved in renal salt transport and the maintenance of blood pressure. The studies will examine the molecular details of kinase activation and examine the possible interaction between the two kinases and/or their isoforms. The studies will make use of novel genetically-modified mouse models to probe the role of the kinase in two different segments of the renal tubule.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
Project #
Application #
Study Section
Special Emphasis Panel (KMBD)
Program Officer
Ketchum, Christian J
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Vanderbilt University Medical Center
Schools of Medicine
United States
Zip Code
Kahle, Kristopher T; Delpire, Eric (2016) Kinase-KCC2 coupling: Cl- rheostasis, disease susceptibility, therapeutic target. J Neurophysiol 115:8-18
Welling, Paul A (2016) Roles and Regulation of Renal K Channels. Annu Rev Physiol 78:415-35
Mathieu, Véronique; Chantôme, Aurélie; Lefranc, Florence et al. (2015) Sphaeropsidin A shows promising activity against drug-resistant cancer cells by targeting regulatory volume increase. Cell Mol Life Sci 72:3731-46
Feng, Xiuyan; Zhang, Yiqian; Shao, Ningjun et al. (2015) Aldosterone modulates thiazide-sensitive sodium chloride cotransporter abundance via DUSP6-mediated ERK1/2 signaling pathway. Am J Physiol Renal Physiol 308:F1119-27
Wade, James B; Liu, Jie; Coleman, Richard et al. (2015) SPAK-mediated NCC regulation in response to low-K+ diet. Am J Physiol Renal Physiol 308:F923-31
Nguyen, Thanh T; Lam, Humphrey V; Phillips, Maxie et al. (2015) Intraoperative optimization to decrease postoperative PRBC transfusion in children undergoing craniofacial reconstruction. Paediatr Anaesth 25:294-300
Kamat, Nikhil V; Thabet, Salim R; Xiao, Liang et al. (2015) Renal transporter activation during angiotensin-II hypertension is blunted in interferon-γ-/- and interleukin-17A-/- mice. Hypertension 65:569-76
Grimm, P Richard; Lazo-Fernandez, Yoskaly; Delpire, Eric et al. (2015) Integrated compensatory network is activated in the absence of NCC phosphorylation. J Clin Invest 125:2136-50
Rengarajan, Srinivas; Lee, Donna H; Oh, Young Taek et al. (2014) Increasing plasma [K+] by intravenous potassium infusion reduces NCC phosphorylation and drives kaliuresis and natriuresis. Am J Physiol Renal Physiol 306:F1059-68
Markadieu, Nicolas; Delpire, Eric (2014) Physiology and pathophysiology of SLC12A1/2 transporters. Pflugers Arch 466:91-105

Showing the most recent 10 out of 22 publications