Abstract
Polymorphisms in the gene encoding sterile 20/SPS1-related proline/alanine-rich kinase (SPAK) associate with hypertension susceptibility in humans. SPAK interacts with WNK kinases to regulate the Na(+)-K(+)-2Cl(-) and Na(+)-Cl(-) co-transporters [collectively, N(K)CC]. Mutations in WNK1/4 and N(K)CC can cause changes in BP and dyskalemia in humans, but the physiologic role of SPAK in vivo is unknown. We generated and analyzed SPAK-null mice by targeting disruption of exons 9 and 10 of SPAK. Compared with SPAK(+/+) littermates, SPAK(+/-) mice exhibited hypotension without significant electrolyte abnormalities, and SPAK(-/-) mice not only exhibited hypotension but also recapitulated Gitelman syndrome with hypokalemia, hypomagnesemia, and hypocalciuria. In the kidney tissues of SPAK(-/-) mice, the expression of total and phosphorylated (p-)NCC was markedly decreased, but that of p-OSR1, total NKCC2, and p-NKCC2 was significantly increased. We observed a blunted response to thiazide but normal response to furosemide in SPAK(-/-) mice. In aortic tissues, total NKCC1 expression was increased but p-NKCC1 was decreased in SPAK-deficient mice. Both SPAK(+/-) and SPAK(-/-) mice had impaired responses to the selective α(1)-adrenergic agonist phenylephrine and the NKCC1 inhibitor bumetanide, suggesting that impaired aortic contractility may contribute to the hypotension of SPAK-null mice. In summary, SPAK-null mice have defects of NCC in the kidneys and NKCC1 in the blood vessels, leading to hypotension through renal salt wasting and vasodilation. SPAK may be a promising target for antihypertensive therapy.
Highlights
Because there was no difference in gross appearance and phenotype between D7 and D10 SPS1-related proline/alanine-rich kinase (SPAK)-null mice strains, the D7 mice were adopted in this study
SPAK-null mice created by targeting disruption of exons 9 and 10 of SPAK (Stk39) exhibited low BP and prominent defects in the distal convoluted tubules (DCTs) with Gitelman syndrome (GS) phenotype, including salt wasting, hypokalemia with renal Kϩ wasting, hypomagnesemia, and hypocalciuria.[12,21]
These results indicate that SPAK is crucial in the regulation of vasoconstriction and renal tubular salt reabsorption
Summary
Sterile 20/SPS1-related proline/alanine-rich kinase (SPAK)[1,2] and oxidative stress-responsive kinase 1 (OSR1)[3] are serine/threonine kinases that share high homology in both their N-terminal catalytic and Cterminal regulatory domains and are widely distributed in the brain, pancreas, heart and kidney.[2,3,4,5] SPAK and OSR1 are downstream substrates of WNK [WithNo-Lysine (K)] 1 and 4 kinases and upstream regulators of the cation-chloride co-transporters (Naϩ-Kϩ2ClϪ co-transporter [NKCC] 1 and 2 and Naϩ-ClϪ co-transporter [NCC]).[6,7,8,9] phosphorylation and activation of SPAK and OSR1 by WNK1/4 can in turn phosphorylate and activate NCC/ NKCC1.10,11. A recent human study showed that genetic variations in the intron regions of STK39, the gene encoding SPAK, could enhance its expression and increase susceptibility to hypertension.[16] These findings suggest that SPAK and OSR1 play important roles in BP and renal tubular electrolyte regulation. The generation and analysis of individual SPAK- or OSR1-deficient mice may provide better platforms to study this issue. For this purpose, we generated SPAK- and OSR1null mice by disrupting exons 9 and 10 of the Stk[39] (SPAK) and Oxsr[1] (OSR1) genes, respectively. Results to be reported indicate that SPAKϪ/Ϫ mice exhibited hypotension but the phenotype of GS
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