Phosphoproteomics of Acute Cell Stressors Targeting Exercise Signaling Networks Reveal Drug Interactions Regulating Protein Secretion

• Published in: Cell reports (Cambridge) 2019-11, Vol.29 (6), p.1524-1538.e6
• Main Authors: Needham, Elise J., Humphrey, Sean J., Cooke, Kristen C., Fazakerley, Daniel J., Duan, Xiaowen, Parker, Benjamin L., James, David E.
• Format: Article
• Language: English
• Subjects: Adrenergic beta-Agonists - metabolism; AMPK; Animals; Aripiprazole - metabolism; Aripiprazole - pharmacology; berberine; Calcium - agonists; Calcium - metabolism; Drug Interactions; EasyPhos; exercise; Exercise - physiology; Humans; Isoproterenol - metabolism; Isoproterenol - pharmacology; Mass Spectrometry; Mice; Muscle, Skeletal - metabolism; myotubes; Phosphoproteins - chemistry; Phosphoproteins - metabolism; phosphoproteomics; Phosphorylation; Plasminogen Activator Inhibitor 1 - genetics; Plasminogen Activator Inhibitor 1 - metabolism; Protein Kinases - metabolism; Protein Translocation Systems - genetics; Protein Translocation Systems - metabolism; Proteome - chemistry; Proteome - metabolism; proteomics; Rats; RNA-Binding Proteins - genetics; RNA-Binding Proteins - metabolism; secretome; signal transduction; Signal Transduction - physiology; Stress, Physiological - genetics; Stress, Physiological - physiology; Thapsigargin - metabolism; Thapsigargin - pharmacology
• ISSN: 2211-1247; 2211-1247
• DOI: 10.1016/j.celrep.2019.10.001

Exercise engages signaling networks to control the release of circulating factors beneficial to health. However, the nature of these networks remains undefined. Using high-throughput phosphoproteomics, we quantify 20,249 phosphorylation sites in skeletal muscle-like myotube cells and monitor their responses to a panel of cell stressors targeting aspects of exercise signaling in vivo. Integrating these in-depth phosphoproteomes with the phosphoproteome of acute aerobic exercise in human skeletal muscle suggests that co-administration of β-adrenergic and calcium agonists would activate complementary signaling relevant to this exercise context. The phosphoproteome of cells treated with this combination reveals a surprising divergence in signaling from the individual treatments. Remarkably, only the combination treatment promotes multisite phosphorylation of SERBP1, a regulator of Serpine1 mRNA stability, a pro-fibrotic secreted protein. Secretome analysis reveals that the combined treatments decrease secretion of SERPINE1 and other deleterious factors. This study provides a framework for dissecting phosphorylation-based signaling relevant to acute exercise. [Display omitted] •Global phosphoproteomes of 9 acute cell stressors integrated with human acute exercise•Comparing AMPK activators reveals potential AMPK substrates and aripiprazole mechanism•Co-administering β-adrenergic and calcium agonists causes extensive interactions•Global secretomes show potential links between exercise benefits and upstream signaling Needham et al. measure phosphoproteomes of acute cell stressors targeting different aspects of exercise signaling. This reveals treatment actions and creates a resource of treatments that regulate phosphosites regulated in acute aerobic exercise in human skeletal muscle. Combining isoproterenol and thapsigargin causes extensive interactions within the phosphoproteome and secretome.