title: Identification of AMPK Phosphorylation Sites Reveals a Network of Proteins Involved in Cell Invasion and Facilitates Large-Scale Substrate Prediction.
abstract: AMP-activated protein kinase (AMPK) is a central energy gauge that regulates metabolism and has been increasingly involved in non-metabolic processes and diseases. However, AMPK's direct substrates in non-metabolic contexts are largely unknown. To better understand the AMPK network, we use a chemical genetics screen coupled to a peptide capture approach in whole cells, resulting in identification of direct AMPK phosphorylation sites. Interestingly, the high-confidence AMPK substrates contain many proteins involved in cell motility, adhesion, and invasion. AMPK phosphorylation of the RHOA guanine nucleotide exchange factor NET1A inhibits extracellular matrix degradation, an early step in cell invasion. The identification of direct AMPK phosphorylation sites also facilitates large-scale prediction of AMPK substrates. We provide an AMPK motif matrix and a pipeline to predict additional AMPK substrates from quantitative phosphoproteomics datasets. As AMPK is emerging as a critical node in aging and pathological processes, our study identifies potential targets for therapeutic strategies.
authors.affiliation: Cancer Biology Program, Stanford University, Stanford, CA 94305, USA; Department of Genetics, Stanford University, Stanford, CA 94305, USA.
authors.forename: Bethany E
authors.initials: BE
authors.lastname: Schaffer
authors.affiliation: Department of Cellular and Molecular Pharmacology, Howard Hughes Medical Institute, University of California, San Francisco, CA 94158, USA.
authors.forename: Rebecca S
authors.initials: RS
authors.lastname: Levin
authors.affiliation: Department of Cellular and Molecular Pharmacology, Howard Hughes Medical Institute, University of California, San Francisco, CA 94158, USA.
authors.forename: Nicholas T
authors.initials: NT
authors.lastname: Hertz
authors.affiliation: Department of Genetics, Stanford University, Stanford, CA 94305, USA.
authors.forename: Travis J
authors.initials: TJ
authors.lastname: Maures
authors.affiliation: Department of Genetics, Stanford University, Stanford, CA 94305, USA.
authors.forename: Michael L
authors.initials: ML
authors.lastname: Schoof
authors.affiliation: Molecular and Cell Biology Laboratory, Salk Institute for Biological Studies, La Jolla, CA 92037, USA.
authors.forename: Pablo E
authors.initials: PE
authors.lastname: Hollstein
authors.affiliation: Department of Genetics, Stanford University, Stanford, CA 94305, USA.
authors.forename: Bérénice A
authors.initials: BA
authors.lastname: Benayoun
authors.affiliation: Department of Genetics, Stanford University, Stanford, CA 94305, USA.
authors.forename: Max R
authors.initials: MR
authors.lastname: Banko
authors.affiliation: Molecular and Cell Biology Laboratory, Salk Institute for Biological Studies, La Jolla, CA 92037, USA.
authors.forename: Reuben J
authors.initials: RJ
authors.lastname: Shaw
authors.affiliation: Department of Cellular and Molecular Pharmacology, Howard Hughes Medical Institute, University of California, San Francisco, CA 94158, USA.
authors.forename: Kevan M
authors.initials: KM
authors.lastname: Shokat
authors.affiliation: Cancer Biology Program, Stanford University, Stanford, CA 94305, USA; Department of Genetics, Stanford University, Stanford, CA 94305, USA; Glenn Laboratories for the Biology of Aging, Stanford, CA 94305, USA. Electronic address: abrunet1@stanford.edu.
authors.forename: Anne
authors.initials: A
authors.lastname: Brunet
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002448:Cell Adhesion / Q000235:genetics*; D002465:Cell Movement / Q000235:genetics; D005109:Extracellular Matrix / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015513:Oncogene Proteins / Q000235:genetics* / Q000378:metabolism; D010455:Peptides / Q000378:metabolism; D010766:Phosphorylation; D060066:Protein Interaction Maps / Q000235:genetics*; D059010:Single-Cell Analysis; D013379:Substrate Specificity
chemical_list: C099763:NET1 protein, human; D015513:Oncogene Proteins; D010455:Peptides; D055372:AMP-Activated Protein Kinases