KLHL2/3 as molecular targets to modulate WNK kinases

• Project/Area Number: 15K15327
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Kidney internal medicine
• Research Institution: Tokyo Medical and Dental University
• Principal Investigator: UCHIDA SHINICIH 東京医科歯科大学, 大学院医歯学総合研究科, 教授 (50262184)
• Project Period (FY): 2015-04-01 – 2017-03-31
• Project Status: Completed (Fiscal Year 2016)
• Budget Amount: ¥3,510,000 (Direct Cost: ¥2,700,000、Indirect Cost: ¥810,000); Fiscal Year 2016: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000); Fiscal Year 2015: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
• Keywords: 高血圧 / 水・電荷室 / キナーゼ / E3リガーゼ / ユビキチンE3リガーゼ / WNKキナーゼ / ノックアウトマウス / Kelch-like protein / WNK kinase

Outline of Final Research Achievements

We evaluated the effects of overexpression of KLHL2 and KLHL3 on WNK protein abundance in cultured cells. Overexpression of KLHL2 or KLHL3 along with WNK kinases dramatically decreased WNK protein abundance. Thus, it would be possible to modulate WNK signaling in vivo by overexpressing KLHL2 or KLHL3. Further study using KLHL2/3 transgenic mice would be necessary. We also evaluated the in vivo roles of KLHL2 and KLHL3 by generating and analyzing KLHL2 and KLHL3 knockout mice. We found that KLHL2 knockout induced WNK4 protein level not in cortex but in medulla. In KLHL3 knockout mice, the major changes of WNK protein observed in various organs was WNK1 and WNK4 in the kidney. Although the abundant expression of KLHL3 in brain was observed, WNK proteins in the whole brain was not increased, suggesting the compensation by KLHL2 or the increased in specific cell populations. We plan further study using the double knockout of KLHL2 and KLHL3.

Research Products

• [Journal Article] Salt suppresses IFNγ inducible chemokines through the IFNγ-JAK1-STAT1 signaling pathway in proximal tubular cells. (2017)
  • Author(s): Arai Y, Takahashi D, Asano K, Tanaka M, Oda M, Ko SBH, Ko MSH, Mandai S, Nomura N, Rai T, Uchida S, Sohara E.
  • Journal Title: Sci Rep. Volume: 7 Issue: 1 Pages: 46580-46580
  • DOI: 10.1038/srep46580
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Loop diuretics affect skeletal myoblast differentiation and exercise-induced muscle hypertrophy. (2017)
  • Author(s): Mandai S, Furukawa S, Kodaka M, Hata Y, Mori T, Nomura N, Ando F, Mori Y, Takahashi D, Yoshizaki Y, Kasagi Y, Arai Y, Sasaki E, Yoshida S, Furuichi Y, Fujii NL, Sohara E, Rai T, Uchida S.
  • Journal Title: Sci Rep. Volume: 7 Issue: 1 Pages: 46369-46369
  • DOI: 10.1038/srep46369
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Impaired degradation of medullary WNK4 in the kidneys of KLHL2 knockout mice. (2017)
  • Author(s): Kasagi Y, Takahashi D, Aida T, Nishida H, Nomura N, Zeniya M, Mori T, Sasaki E, Ando F, Rai T, Uchida S, Sohara E.
  • Journal Title: Biochem Biophys Res Commun. Volume: 487 Issue: 2 Pages: 368-374
  • DOI: 10.1016/j.bbrc.2017.04.068
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Calcineurin inhibitors block sodium-chloride cotransporter dephosphorylation in response to high potassium intake. (2017)
  • Author(s): Shoda W, Nomura N, Ando F, Mori Y, Mori T, Sohara E, Rai T, Uchida S.
  • Journal Title: Kidney Int. Volume: 91 Issue: 2 Pages: 402-411
  • DOI: 10.1016/j.kint.2016.09.001
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] WNK4 is an Adipogenic Factor and Its Deletion Reduces Diet-Induced Obesity in Mice. (2017)
  • Author(s): Takahashi D, Mori T, Sohara E, Tanaka M, Chiga M, Inoue Y, Nomura N, Zeniya M, Ochi H, Takeda S, Suganami T, Rai T, Uchida S.
  • Journal Title: EBioMedicine Volume: 18 Pages: 118-27
  • DOI: 10.1016/j.ebiom.2017.03.011
  • Peer Reviewed / Open Access
• [Journal Article] KLHL3 Knockout Mice Reveal the Physiological Role of KLHL3 and the Pathophysiology of Pseudohypoaldosteronism Type II Caused by Mutant KLHL3. (2017)
  • Author(s): Sasaki E, Susa K, Mori T, Isobe K, Araki Y, Inoue Y, Yoshizaki Y, Ando F, Mori Y, Mandai S, Zeniya M, Takahashi D, Nomura N, Rai T, Uchida S, Sohara E.
  • Journal Title: Mol Cell Biol. Volume: 27 Issue: 7 Pages: 1-13
  • DOI: 10.1128/mcb.00508-16
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Wnt5a induces renal AQP2 expression by activating calcineurin signalling pathway. (2016)
  • Author(s): Ando F, Sohara E, Morimoto T, Yui N, Nomura N, Kikuchi E, Takahashi D, Mori T, Vandewalle A, Rai T, Sasaki S, Kondo Y, Uchida S.
  • Journal Title: Nat Commun. Volume: 7 Issue: 1 Pages: 13636-13636
  • DOI: 10.1038/ncomms13636
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Generation and analysis of knock-in mice carrying pseudohypoaldosteronism type II-causing mutations in the cullin 3 gene. (2015)
  • Author(s): Araki Y, Rai T, Sohara E, Mori T, Inoue Y, Isobe K, Kikuchi E, Ohta A, Sasaki S, Uchida S.
  • Journal Title: Biol Open Volume: 4 Issue: 11 Pages: 1509-1517
  • DOI: 10.1242/bio.013276
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Genetic knockout and pharmacologic inhibition of NCX2 cause natriuresis and hypercalciuria. (2015)
  • Author(s): Gotoh Y, Kita S, Fujii M, Tagashira H, Horie I, Arai Y, Uchida S, Iwamoto T
  • Journal Title: Biochem Biophys Res Commun Volume: 456 (2) Issue: 2 Pages: 670-675
  • DOI: 10.1016/j.bbrc.2014.12.016
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Kelch-Like Protein 2 Mediates Angiotensin II-With No Lysine 3 Signaling in the Regulation of Vascular Tonus (2015)
  • Author(s): Zeniya M, Morimoto N, Takahashi D, Mori Y, Mori T, Ando F, Araki Y, Yoshizaki Y, Inoue Y, Isobe K, Nomura N, Oi K, Nishida H, Sasaki S, Sohara E, Rai T, Uchida S
  • Journal Title: J Am Soc Nephrol Volume: 26 Issue: 9 Pages: 2129-2138
  • DOI: 10.1681/asn.2014070639
  • Peer Reviewed
• [Journal Article] Involvement of selective autophagy mediated by p62/SQSTM1 in KLHL3-dependent WNK4 degradation. (2015)
  • Author(s): Mori Y, Mori T, Wakabayashi M, Yoshizaki Y, Zeniya M, Sohara E, Rai T, Uchida S.
  • Journal Title: Biochem J. Volume: 472 Issue: 1 Pages: 33-41
  • DOI: 10.1042/bj20150500
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Impaired degradation of WNK by Akt and PKA phosphorylation of KLHL3. (2015)
  • Author(s): Yoshizaki Y, Mori Y, Tsuzaki Y, Mori T, Nomura N, Wakabayashi M, Takahashi D, Zeniya M, Kikuchi E, Araki Y, Ando F, Isobe K, Nishida H, Ohta A, Susa K, Inoue Y, Chiga M, Rai T, Sasaki S, Uchida S, Sohara E.
  • Journal Title: Biochem Biophys Res Commun. Volume: 467 Issue: 2 Pages: 229-234
  • DOI: 10.1016/j.bbrc.2015.09.184
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Associated STK39 Polymorphism Knockin Cell Lines With the Clustered Regularly Interspaced Short Palindromic Repeats/Cas9 System. (2015)
  • Author(s): Mandai S, Mori T, Sohara E, Rai T, Uchida S.
  • Journal Title: Hypertension Volume: 66 Issue: 6 Pages: 1199-1206
  • DOI: 10.1161/hypertensionaha.115.05872
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Discovery of Novel SPAK Inhibitors That Block WNK Kinase Signaling to Cation Chloride Transporters (2014)
  • Author(s): Kikuchi E, Mori T, Zeniya M, Isobe K, Ishigami-Yuasa M, Fujii S, Kagechika H, Ishihara T, Mizushima T, Sasaki S, Sohara E, Rai T, Uchida S
  • Journal Title: J Am Soc Nephrol Volume: 26 Issue: 7 Pages: 1525-1536
  • DOI: 10.1681/asn.2014060560
  • Peer Reviewed
• [Presentation] Generation and analysis of KLHL3 knockout mice (2016)
  • Author(s): Emi Sasaki, Koichiro Susa, Takayasu Mori, Kiyoshi Isobe, Yuya Araki, Yuichi Inoue, Tatemitsu Rai, Shinichi Uchida, Eisei Sohara
  • Organizer: Annual Meeting of the American Society of Nephrology, Kidney Week 2016
  • Place of Presentation: Chicago（米国）
  • Year and Date: 2016-11-17
  • Int'l Joint Research
• [Presentation] Sodium chloride suppresses interferon-gamma inducible chemokines through the JAK/STAT signaling pathway in proximal tubular epithelial cells (2016)
  • Author(s): Yohei Arai, Daiei Takahashi, Kenichi Asano, Masato Tanaka, Mayumi Oda, Shigeru B.H. Ko, Minoru S.H. Ko, Tatemitsu Rai, Shinichi Uchida, Eisei Sohara
  • Organizer: Annual Meeting of the American Society of Nephrology, Kidney Week 2016
  • Place of Presentation: Chicago（米国）
  • Year and Date: 2016-11-17
  • Int'l Joint Research
• [Presentation] Drug Repositioning Screening for the Discovery of Inhibitors of Keap1-Nrf2 Interaction Inhibitors by Using Fluorescent Correlation Spectroscopy (2016)
  • Author(s): Yuki Yoshizaki, Takayasu Mori, Eriko Kikuchi, Daiei Takahashi, Moko Zeniya, Yuya Araki, Yutaro Mori, Fumiaki Ando, Tatemitsu Rai, Shinichi Uchida, Eisei Sohara.
  • Organizer: Annual Meeting of the American Society of Nephrology, Kidney Week 2016
  • Place of Presentation: Chicago（米国）
  • Year and Date: 2016-11-17
  • Int'l Joint Research
• [Presentation] The KLHL proteins, the WNKs and renal Na+ transport. (2015)
  • Author(s): Uchida S
  • Organizer: 8Th FAOPS (Federation of the Asian and Oceanian Physiological Societies) Congress 2015
  • Place of Presentation: Bangkok, Thailand
  • Year and Date: 2015-11-23
  • Int'l Joint Research / Invited
• [Presentation] Generation and Analysis of Knock-In Mice Carrying Pseudohypoaldosteronism Type II-Causing Mutations in the Cullin 3 Gene. (2015)
  • Author(s): Araki Y, Rai T, Sohara E, Mori T, Inoue Y, Kikuchi E, Uchida S.
  • Organizer: The 48th Annual Meeting of the American Society of Nephrology
  • Place of Presentation: San Diego, USA
  • Year and Date: 2015-11-05
  • Int'l Joint Research
• [Presentation] Potassium-induced dephosphorylation of renal sodium-chloride cotransporter is NOT dependent on the anions. (2015)
  • Author(s): Nomura N, Shoda W, Sohara E, Rai T, Uchida S.
  • Organizer: The 48th Annual Meeting of the American Society of Nephrology
  • Place of Presentation: San Diego
  • Year and Date: 2015-11-05
  • Int'l Joint Research
• [Presentation] Generation of Hypertension-Associated STK39 Polymor- phism Knockin Cell Lines with the CRISPR/Cas9 System. (2015)
  • Author(s): Mandai S, Mori T, Sohara E, Rai T, Uchida S.
  • Organizer: The 48th Annual Meeting of the American Society of Nephrology
  • Place of Presentation: San Diego
  • Year and Date: 2015-11-05
  • Int'l Joint Research
• [Presentation] The Major Contribution of WNK4 to the Pathogenesis of Pseudohypoaldosteronism Type II (PHAII) Caused by the KLHL3 Mutation R528H. (2015)
  • Author(s): Susa K, Sohara E, Takahashi D, Rai T, Uchida S.
  • Organizer: The 48th Annual Meeting of the American Society of Nephrology
  • Place of Presentation: San Diego
  • Year and Date: 2015-11-05
  • Int'l Joint Research
• [Remarks]
  • URL: http://www.tmd.ac.jp/grad/kid/index.html
• [Remarks] 東京医科歯科大学腎臓内科学
  • URL: http://www.tmd.ac.jp/grad/kid/index.html