Phosphoproteomics of three exercise modalities identifies canonical signaling and C18ORF25 as an AMPK substrate regulating skeletal muscle function

Cell Metab. 2022 Oct 4;34(10):1561-1577.e9. doi: 10.1016/j.cmet.2022.07.003.

Ronnie Blazev ¹, Christian S Carl ², Yaan-Kit Ng ¹, Jeffrey Molendijk ¹, Christian T Voldstedlund ², Yuanyuan Zhao ³, Di Xiao ⁴, Andrew J Kueh ⁵, Paula M Miotto ³, Vanessa R Haynes ³, Justin P Hardee ¹, Jin D Chung ¹, James W McNamara ⁶, Hongwei Qian ¹, Paul Gregorevic ¹, Jonathan S Oakhill ⁷, Marco J Herold ⁵, Thomas E Jensen ², Leszek Lisowski ⁸, Gordon S Lynch ¹, Garron T Dodd ³, Matthew J Watt ³, Pengyi Yang ⁹, Bente Kiens ¹⁰, Erik A Richter ¹¹, Benjamin L Parker ¹²

Affiliations

1 Department of Anatomy & Physiology, The University of Melbourne, Parkville, VIC, Australia; Centre for Muscle Research, The University of Melbourne, Parkville, VIC, Australia.
2 August Krogh Section for Molecular Physiology, Department of Nutrition, Exercise and Sports, Faculty of Science, The University of Copenhagen, Copenhagen, Denmark.
3 Department of Anatomy & Physiology, The University of Melbourne, Parkville, VIC, Australia.
4 Children's Medical Research Institute, The University of Sydney, Camperdown, NSW, Australia; School of Mathematics and Statistics, The University of Sydney, Camperdown, NSW, Australia.
5 The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia.
6 Department of Anatomy & Physiology, The University of Melbourne, Parkville, VIC, Australia; Centre for Muscle Research, The University of Melbourne, Parkville, VIC, Australia; Murdoch Children's Research Institute and Melbourne Centre for Cardiovascular Genomics and Regenerative Medicine, The Royal Children's Hospital, Parkville, VIC, Australia.
7 St Vincent's Institute of Medical Research, Fitzroy, VIC, Australia.
8 Children's Medical Research Institute, The University of Sydney, Camperdown, NSW, Australia; Military Institute of Medicine, Warsaw, Poland.
9 Children's Medical Research Institute, The University of Sydney, Camperdown, NSW, Australia; The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia.
10 August Krogh Section for Molecular Physiology, Department of Nutrition, Exercise and Sports, Faculty of Science, The University of Copenhagen, Copenhagen, Denmark. Electronic address: bkiens@nexs.ku.dk.
11 August Krogh Section for Molecular Physiology, Department of Nutrition, Exercise and Sports, Faculty of Science, The University of Copenhagen, Copenhagen, Denmark. Electronic address: erichter@nexs.ku.dk.
12 Department of Anatomy & Physiology, The University of Melbourne, Parkville, VIC, Australia; Centre for Muscle Research, The University of Melbourne, Parkville, VIC, Australia. Electronic address: ben.parker@unimelb.edu.au.

PMID: 35882232 DOI: 10.1016/j.cmet.2022.07.003

Abstract

Exercise induces signaling networks to improve muscle function and confer health benefits. To identify divergent and common signaling networks during and after different exercise modalities, we performed a phosphoproteomic analysis of human skeletal muscle from a cross-over intervention of endurance, sprint, and resistance exercise. This identified 5,486 phosphosites regulated during or after at least one type of exercise modality and only 420 core phosphosites common to all exercise. One of these core phosphosites was S67 on the uncharacterized protein C18ORF25, which we validated as an AMPK substrate. Mice lacking C18ORF25 have reduced skeletal muscle fiber size, exercise capacity, and muscle contractile function, and this was associated with reduced phosphorylation of contractile and CA²⁺ handling proteins. Expression of C18ORF25 S66/67D phospho-mimetic reversed the decreased muscle force production. This work defines the divergent and canonical exercise phosphoproteome across different modalities and identifies C18ORF25 as a regulator of exercise signaling and muscle function.

Keywords

AMPK; C18ORF25; exercise; phosphoproteomics; signaling; skeletal muscle.

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