腎臓における新規の血圧調節機構ＷＮＫ－ＮＣＣシグナル伝達系の解明

2013 年度 研究成果報告書

• 研究課題/領域番号: 22390168
• 研究種目: 基盤研究(B)
• 配分区分: 補助金
• 応募区分: 一般
• 研究分野: 腎臓内科学
• 研究機関: 東京医科歯科大学
• 研究代表者: 頼 建光 東京医科歯科大学, 医学部附属病院, 准教授 (80334431)
• 研究期間 (年度): 2010-04-01 – 2014-03-31
• キーワード: 偽性低アルドステロン血症II型 / KLHL3 / Cullin3 / ユビキチン / SPAK / Na-K-Cl共輸送体 / OSR1 / angiotensin II

研究概要

本研究では、新規血圧調節系であるWNK4-NCC刺激伝達系の詳細を明らかにし、これらの知見をもとに、高血圧の新しい治療戦略を立ち上げることを目的とした。野生型WNK4がNCCの正の制御因子である事を確認した。WNK4はKLHL3とCullin3と結合してユビキチン化されることで発現量を制御され、疾患起因性変異によるWNK4のユビキチン化障害がPHAII発症の共通の病態であることを明らかにした。WNK キナーゼとOSR1/SPAK との間の結合の阻害活性を持つ化合物のスクリーニングを行い、新規降圧薬開発へ向けて有望な化合物を得ることに成功した.

研究成果

• [雑誌論文] Impaired degradation of WNK1 and WNK4 kinases causes PHAII in mutant KLHL3 knock-in mice (2014)
  • 著者名/発表者名: Susa K, Sohara E, Rai T, Zeniya M, Mori Y, Mori T, Chiga M, Nomura N, Nishida H, Takahashi D, Isobe K, Inoue Y, Takeishi K, Takeda N, Sasaki S, Uchida S
  • 雑誌名: Hum Mol Genet 巻: (Epub ahead of print)
  • 査読あり
• [雑誌論文] WNK4 is the major WNK kinase positively regulating NCC in the mouse kidney (2014)
  • 著者名/発表者名: Takahashi D, Mori T, Nomura N, Khan MZ, Araki Y, Zeniya M, Sohara E, Rai T, Sasaki S, Uchida S
  • 雑誌名: Biosci Rep 巻: (Epub ahead of print)
  • 査読あり
• [雑誌論文] Regulation of with-no-lysine kinase signaling by Kelch-like proteins (2014)
  • 著者名/発表者名: Uchida S, Sohara E, Rai T, Sasaki S
  • 雑誌名: Biol Cell 巻: 106(2) ページ: 45-56
  • DOI: 10.1111/boc.201300069
  • 査読あり
• [雑誌論文] Development of enzyme-linked immunosorbent assays for urinary thiazide-sensitive Na-Cl cotransporter (NCC) measurement (2013)
  • 著者名/発表者名: Isobe K, Mori T, Asano T, Kawaguchi H, Nonoyama S, Kumagai N, Kamada F, Morimoto T, Hayashi M, Sohara E, Rai T, Sasaki S, Uchida S
  • 雑誌名: Am J Physiol Renal Physiol 巻: 305(9) ページ: F1374-81
  • DOI: 10.1152/ajprenal.00208.2013
  • 査読あり
• [雑誌論文] Dietary salt intake regulates WNK3-SPAK-NKCC1 phosphorylation cascade in mouse aorta through angiotensin II (2013)
  • 著者名/発表者名: Zeniya M, Sohara E, Kita S, Iwamoto T, Susa K, Mori T, Oi K, Chiga M, Takahashi D, Yang SS, Lin SH, Rai T, Sasaki S, Uchida S
  • 雑誌名: Hypertension 巻: 62(5) ページ: 872-8
  • DOI: 10.1161/HYPERTENSIONAHA.113.01543
  • 査読あり
• [雑誌論文] Chemical library screening for WNK signalling inhibitors using fluorescence correlation spectroscopy (2013)
  • 著者名/発表者名: Mori T, Kikuchi E, Watanabe Y, Fujii S, Ishigami-Yuasa M, Kagechika H, Sohara E, Rai T, Sasaki S, Uchida S
  • 雑誌名: Biochem J 巻: 455(3) ページ: 339-45
  • DOI: 10.1042/BJ20130597
  • 査読あり
• [雑誌論文] Decrease of WNK4 ubiquitination by disease-causing mutations of KLHL3 through different molecular mechanisms (2013)
  • 著者名/発表者名: Mori Y, Wakabayashi M, Mori T, Araki Y, Sohara E, Rai T, Sasaki S, Uchida S
  • 雑誌名: Biochem Biophys Res Commun 巻: 439(1) ページ: 30-4
  • DOI: 10.1016/j.bbrc.2013.08.035
  • 査読あり
• [雑誌論文] KLHL2 interacts with and ubiquitinates WNK kinases (2013)
  • 著者名/発表者名: Takahashi D, Mori T, Wakabayashi M, Mori Y, Susa K, Zeniya M, Sohara E, Rai T, Sasaki S, Uchida S
  • 雑誌名: Biochem Biophys Res Commun 巻: 437(3) ページ: 457-62
  • DOI: 10.1016/j.bbrc.2013.06.104
  • 査読あり
• [雑誌論文] Impaired KLHL3-mediated ubiquitination of WNK4 causes human hypertension (2013)
  • 著者名/発表者名: Wakabayashi M, Mori T, Isobe K, Sohara E, Susa K, Araki Y, Chiga M, Kikuchi E, Nomura N, Mori Y, Matsuo H, Murata T, Nomura S, Asano T, Kawaguchi H, Nonoyama S, Rai T, Sasaki S, Uchida S
  • 雑誌名: Cell Rep 巻: 3(3) ページ: 858-68
  • DOI: 10.1016/j.celrep.2013.02.024
  • 査読あり
• [雑誌論文] A minor role of WNK3 in regulating phosphorylation of renal NKCC2 and NCC co-transporters in vivo (2013)
  • 著者名/発表者名: Oi K, Sohara E, Rai T, Misawa M, Chiga M, Alessi DR, Sasaki S, Uchida S
  • 雑誌名: Biol Open 巻: 1(2) ページ: 120-7
  • DOI: 10.1242/bio.2011048
  • 査読あり
• [雑誌論文] WNK-OSR1/SPAK-NCC signal cascade has circadian rhythm dependent on aldosterone (2012)
  • 著者名/発表者名: Susa K, Sohara E, Isobe K, Chiga M, Rai T, Sasaki S, Uchida S
  • 雑誌名: Biochem Biophys Res Commun 巻: 427(4) ページ: 743-7
  • DOI: 10.1016/j.bbrc.2012.09.130
  • 査読あり
• [雑誌論文] Does a β(2)-adrenergic receptor - WNK4 - Na-Cl co-transporter signal cascade exist in the in vivo kidney? (2012)
  • 著者名/発表者名: Uchida S, Chiga M, Sohara E, Rai T, Sasaki S
  • 雑誌名: Nat Med 巻: 18(9) ページ: 1324-5
  • DOI: 10.1038/nm.2809
  • 査読あり
• [雑誌論文] Phosphatidylinositol 3-kinase/Akt signaling pathway activates the WNK - OSR1/SPAK - NCC phosphorylation cascade in hyperinsulinemic db/db Mice (2012)
  • 著者名/発表者名: Nishida H, Sohara E, Nomura N, Chiga M, Alessi DR, Rai T, Sasaki S, Uchida S
  • 雑誌名: Hypertension 巻: 60(4) ページ: 981-90
  • DOI: 10.1161/HYPERTENSIONAHA
  • 査読あり
• [雑誌論文] Phosphorylation of Na-Cl cotransporter by OSR1 and SPAK kinases regulates its ubiquitination (2012)
  • 著者名/発表者名: Hossain Khan MZ, Sohara E, Ohta A, Chiga M, Inoue Y, Isobe K, Wakabayashi M, Oi K, Rai T, Sasaki S, Uchida S
  • 雑誌名: Biochem Biophys Res Commun 巻: 425(2) ページ: 456-61
  • DOI: 10.1016/j.bbrc.2012.07.124
  • 査読あり
• [雑誌論文] Effect of heterozygous deletion of WNK1 on the WNK-OSR1 / SPAK-NCC / NKCC1 / NKCC2 signal cascade in the kidney and blood vessels (2012)
  • 著者名/発表者名: Susa K, Kita S, Iwamoto T, Yang SS, Lin SH, Ohta A, Sohara E, Rai T, Sasaki S, Alessi DR, Uchida S
  • 雑誌名: Clin Exp Nephrol 巻: 16(4) ページ: 530-8
  • 査読あり
• [雑誌論文] Acute insulin stimulation induces phosphorylation of the Na-Cl cotransporter in cultured distal mpkDCT cells and mouse kidney (2011)
  • 著者名/発表者名: Sohara E, Rai T, Yang SS, Ohta A, Naito S, Chiga M, Nomura N, Lin SH, Vandewalle A, Ohta E, Sasaki S, Uchida S
  • 雑誌名: PLos One 巻: 6(8) ページ: 6(8)
  • DOI: 10.1371/journal.pone.0024277
  • 査読あり
• [雑誌論文] Phenotypes of pseudohypoaldosteronism type II caused by the WNK4 D561A missense mutation are dependent on the WNK-OSR1/ SPAK kinase cascade (2011)
  • 著者名/発表者名: Chiga M, Rafiqi FH, Alessi DR, Sohara E, Ohta A, Rai T, Sasaki S, Uchida S
  • 雑誌名: J Cell Sci 巻: 124(Pt9) ページ: 1391-5
  • DOI: 10.1242/jcs.084111
  • 査読あり
• [雑誌論文] Regulation of WNK1 kinase by extracellular potassium (2011)
  • 著者名/発表者名: Naito S, Ohta A, Sohara E, Ohta E, Rai T, Sasaki S, Shinichi Uchida
  • 雑誌名: Clin Exp Nephrol 巻: 15(2) ページ: 195-202
  • DOI: 10.1007/s10157-010-0378-9
  • 査読あり
• [雑誌論文] Mechanisms for hypercalciuria in pseudohypoaldosteronism type II-causing WNK4 knock-in mice (2010)
  • 著者名/発表者名: Yang SS, Hsu YJ, Chiga M, Rai T, Sasaki S, Uchida S, Lin SH
  • 雑誌名: Endocrinology 巻: 151(4) ページ: 1829-36
  • DOI: 10.1210/en.2009-0951
  • 査読あり
• [雑誌論文] Effect of angiotensin II on the WNK-OSR1/SPAK-NCC phosphorylation cascade in cultured mpkDCT cells and in vivo mouse kidney (2010)
  • 著者名/発表者名: Talati G, Ohta A, Rai T, Sohara E, Naito S, Vandewalle A, Sasaki S, Uchida S
  • 雑誌名: Biochem Biophys Res Commun 巻: 393(4) ページ: 844-8
  • DOI: 10.1016/j.bbrc.2010.02.096
  • 査読あり
• [学会発表] Clinical significance of urinary thiazide-sensitive Na-Cl cotransporter (NCC) measurement by newly developed enzyme-linked immunosorbent assays (2013)
  • 著者名/発表者名: Isobe K, Mori T, Sohara E, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2013
  • 発表場所: Atlanta, U.S.A.
  • 年月日: 20131105-10
• [学会発表] Low salt intake decreased transcription and protein level of KLHL3 in mouse kidney (2013)
  • 著者名/発表者名: Susa K, Sohara E, Zeniya M, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2013
  • 発表場所: Atlanta, U.S.A.
  • 年月日: 20131105-10
• [学会発表] Chemical library screening for WNK signaling inhibitors by using fluorescent correlation spectroscopy (2013)
  • 著者名/発表者名: Mori T, Kikuchi E, Sohara E, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2013
  • 発表場所: Atlanta, U.S.A.
  • 年月日: 20131105-10
• [学会発表] Impaired KHLH3-mediated ubiquitination of WNK4 activates OSR1 and SPAK kinases - NaCl cotransporter (NCC) signaling and causes hypertension (2013)
  • 著者名/発表者名: Wakabayashi M, Mori T, Isobe K, Sohara E, Susa K, Araki Y, Chiga M, Kikuchi E, Nomura N, Mori Y, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2013
  • 発表場所: Atlanta, U.S.A.
  • 年月日: 20131105-10
• [学会発表] Analyses of KHLH3 mutants that cause Pseudohypoaldosteronism type II (2013)
  • 著者名/発表者名: Mori Y, Wakabayashi M, Mori T, Araki Y, Sohara E, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2013
  • 発表場所: Atlanta, U.S.A.
  • 年月日: 20131105-10
• [学会発表] KHLH2 interacts with and ubiquitinates WNK4 kinases (2013)
  • 著者名/発表者名: Takahashi D, Mori T, Wakabayashi M, Mori Y, Susa K, Zeniya M, Rai T, Sohara E, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2013
  • 発表場所: Atlanta, U.S.A.
  • 年月日: 20131105-10
• [学会発表] WNK3 regulates blood pressure through the regulation of vascular OSR1 / SPAK-NKCC1 phosphorylation cascade (2013)
  • 著者名/発表者名: Zeniya M, Sohara E, Oi K, Chiga M, Susa K, Mori T, Takahashi D, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2013
  • 発表場所: Atlanta, U.S.A.
  • 年月日: 20131105-10
• [学会発表] Chemical library screening for direct SPAK inhibitors by a newly devoloped ELISA (2012)
  • 著者名/発表者名: Kikuchi E, Mori T, Isobe K, Sohara E, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2012
  • 発表場所: San Diego, U.S.A.
  • 年月日: 20121030-1104
• [学会発表] Development of sandwich enzyme linked immunosorbent assay for measurement of urinary total and phosphorylated Na-Cl cotransporter protein (2012)
  • 著者名/発表者名: Isobe K, Sohara E, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2012
  • 発表場所: San Diego, U.S.A.
  • 年月日: 20121030-1104
• [学会発表] Dietary salt intake and angiotensin II regulates WNK - SPAK - NKCC1 phosphorylation cascade in mouse aorta (2012)
  • 著者名/発表者名: Zeniya M, Sohara E, Oi K, Chiga M, Susa K, Mori T, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2012
  • 発表場所: San Diego, U.S.A.
  • 年月日: 20121030-1104
• [学会発表] Phosphorylation of Na-Cl cotransporter by OSR1 and SPAK kinases regulates its ubiquitination (2012)
  • 著者名/発表者名: Kahn MZH, Sohara E, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2012
  • 発表場所: San Diego, U.S.A.
  • 年月日: 20121030-1104
• [学会発表] Generation and analysis of WT-WNK4 transgenic mice reveal the physiological role of WNK4 (2012)
  • 著者名/発表者名: Mori T, Sohara E, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2012
  • 発表場所: San Diego, U.S.A.
  • 年月日: 20121030-1104
• [学会発表] WNK4 - OSR1/SPAK - NCC signal cascade has circadian rhythm dependent on aldosterone (2012)
  • 著者名/発表者名: Susa K, Sohara E, Isobe K, Chiga M, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2012
  • 発表場所: San Diego, U.S.A.
  • 年月日: 20121030-1104
• [学会発表] Increased Na-Cl cotransporter phosphorylation in hyperinsulinemic db/db mice is regulated by insulin/PI3K pathway (2011)
  • 著者名/発表者名: Nishida H, Sohara E, Alessi D, Nomura N, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2011
  • 発表場所: Philadelphia, U.S.A
  • 年月日: 20111108-13
• [学会発表] Increased protein abundance of the mutant WNK4 may be a cause of the increased WNK4 kinase activity in the mouse model of pseudohypoaldosteronism type II(PHA II). (2011)
  • 著者名/発表者名: Wakabayashi M, Naito S, Sohara E, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2011
  • 発表場所: Philadelphia, U.S.A.
  • 年月日: 20111108-13
• [学会発表] Acute insulin stimulation induces phosphorylation of the Na-Cl cotransporter in cultured distal mpkDCT cells and mouse kidney (2011)
  • 著者名/発表者名: Sohara E, Rai T, Yang SS, Ohta A, Lin SH, Vandewalle A, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2011
  • 発表場所: Philadelphia, U.S.A.
  • 年月日: 20111108-13
• [学会発表] Development of new systems to measure total and phosphorylated Na-Cl cotransporter (NCC) protein in human urine (2011)
  • 著者名/発表者名: Isobe K, Sohara E, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2011
  • 発表場所: Philadelphia, U.S.A.
  • 年月日: 20111108-13
• [学会発表] Urinary excretion of Na-Cl cotransporter in exosomes is increased by high salt diet as well as low salt diet (2011)
  • 著者名/発表者名: Kahn MZH, Sohara E, Ohta A, Naito S, Chiga M, Rai T, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2011
  • 発表場所: Philadelphia, U.S.A.
  • 年月日: 20111108-13
• [学会発表] WNK - OSR1/SPAK - SLC12A phosphorylation cascade in the WNK1 (+/-) mice (2011)
  • 著者名/発表者名: Susa K, Sohara E, Rai T, Kita S, Iwamoto T, Alessi D, Sasaki S, Uchida S
  • 学会等名: American Society of Nephrology Kidney Week 2011
  • 発表場所: Philadelphia, U.S.A
  • 年月日: 20111108-13