Ducommun S Deak M Zeigerer A Göransson O Seitz S Collodet C Madsen AB Jensen TE Viollet B Foretz M Gut P Sumpton D Sakamoto K 2019 AMP-activated protein kinase (AMPK) is a key regulator of cellular energy homeostasis, acting as a sensor of energy and nutrient status. As such, AMPK is considered a promising drug target for treatment of medical conditions particularly associated with metabolic dysfunctions. To better understand the downstream effectors and physiological consequences of AMPK activation, we have employed a chemical genetic screen in mouse primary hepatocytes in an attempt to identify novel AMPK targets. Treatment of hepatocytes with a potent and specific AMPK activator 991 resulted in identification of 65 proteins phosphorylated upon AMPK activation, which are involved in a variety of cellular processes such as lipid/glycogen metabolism, vesicle trafficking, and cytoskeleton organisation. Further characterisation and validation using mass spectrometry followed by immunoblotting analysis with phosphorylation site-specific antibodies identified AMPK-dependent phosphorylation of Gapex-5 (also known as GTPase-activating protein and VPS9 domain-containing protein 1 (GAPVD1)) on Ser902 in hepatocytes and starch-binding domain 1 (STBD1) on Ser175 in multiple cells/tissues. As new promising roles of AMPK as a key metabolic regulator continue to emerge, the substrates we identified could provide new mechanistic and therapeutic insights into AMPK-activating drugs in the liver. https://sonar.ch/global/documents/209830 eng info:eu-repo/semantics/altIdentifier/doi/10.1016/j.cellsig.2019.02.001 info:eu-repo/semantics/altIdentifier/pmid/30772465 Cellular signalling. - 2019 GTPase activating protein and VPS9 domains 1 Shokat Starch-binding domain 1 AMP-Activated Protein Kinases Animals Guanine Nucleotide Exchange Factors Hepatocytes Homeostasis Lipid Metabolism Liver Mass Spectrometry Membrane Proteins Mice, Knockout Muscle Proteins Phosphorylation Substrate Specificity Chemical genetic screen identifies Gapex-5/GAPVD1 and STBD1 as novel AMPK substrates. http://purl.org/coar/resource_type/c_6501