Kidney Week

Abstract: SA-PO830

WNK1 Regulates Skeletal Muscle Hypertrophy by Modulating Phosphorylation, Nuclear Localization, and Transcriptional Activity of FoxO4

Session Information

• Molecular Mechanisms of CKD - III
  October 27, 2018 | Location: Exhibit Hall, San Diego Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: CKD (Non-Dialysis)

• 1903 CKD (Non-Dialysis): Mechanisms

Authors

• Mandai, Shintaro, Tokyo Medical and Dental University, Tokyo, Japan

Shintaro Mandai,

• Mori, Takayasu, Tokyo Medical and Dental University, Tokyo, Japan

Takayasu Mori,

• Nomura, Naohiro, Tokyo Medical and Dental University, Tokyo, Japan

Naohiro Nomura,

• Furusho, Taisuke, Tokyo Medical and Dental University, Tokyo, Japan

Taisuke Furusho,

• Arai, Yohei, Tokyo Medical and Dental University, Tokyo, Japan

Yohei Arai,

• Kikuchi, Hiroaki, Tokyo Medical and Dental University, Tokyo, Japan

Hiroaki Kikuchi,

• Sasaki, Emi, Tokyo Medical and Dental University, Tokyo, Japan

Emi Sasaki,

• Sohara, Eisei, Tokyo Medical and Dental University, Tokyo, Japan

Eisei Sohara,

• Rai, Tatemitsu, Tokyo Medical and Dental University, Tokyo, Japan

Tatemitsu Rai,

• Uchida, Shinichi, Tokyo Medical and Dental University, Tokyo, Japan

Shinichi Uchida,

Background

With-no-lysine (K) (WNK) kinases, mutated in the inherited form of hypertension pseudohypoaldosteronism type II, are essential regulators of ion transporters. WNK kinases phosphorylate and activate SLC12A cation-chloride cotransporters via the intermediate kinases SPAK/OSR1. We previously showed that Na⁺-K⁺-2Cl⁻ cotransporter 1, a member of SLC12A family, promotes mouse skeletal myogenesis and its inhibitor loop diuretics are associated with risk for sarcopenia in patients with chronic kidney disease (CKD). We further aimed to investigate the physiological role of WNK1 that is the major WNK isoform in mammalian skeletal muscle.

Methods

In C2C12 mouse skeletal muscle cells, differentiation was induced after transfection of Wnk1-targeted siRNA, and the diameter of differentiated myotubes was evaluated. In C57BL/6J mice, WNK1 protein abundance was evaluated after six weeks of voluntary wheel running or four weeks of 0.25% adenine-containing diet feeding (CKD model).

Results

In C2C12 cells, silencing of WNK1, but unexpectedly not SPAK/OSR1, induced myotube atrophy and remarkable increases in the mRNA expression of the muscle atrophy ubiquitin ligases MAFbx and MuRF1 called ‘atrogenes’. These atrogenes are predominantly upregulated by transcription factors forkhead box protein O (FoxO)1/3/4 in CKD or other diseases causing sarcopenia. WNK1 silencing selectively increased FoxO4 nuclear localization, and co-transfection of FoxO4-targeted siRNA completely reversed the myotube atrophy and upregulation of atrogene transcription. Phos-tag SDS-PAGE revealed that WNK1 silencing decreased FoxO4 phosphorylation, which might be associated with nuclear localization and transcriptional activity of this protein. We further showed that WNK1 abundance in mouse skeletal muscle was increased by chronic exercise and decreased in adenine-induced CKD and sarcopenia. Moreover, to validate the role of WNK1 in muscle hypertrophy in vivo, a WNK kinase inhibitor WNK463 was administered by oral gavage (10 mg/kg), resulting in marked increase of atrogene expression in mouse skeletal muscle.

Conclusion

WNK1 physiologically regulates mammalian skeletal muscle hypertrophy via interactions with FoxO4. The WNK1-FoxO4 axis may be a novel therapeutic target in sarcopenia.