title: A loss-of-function screen for phosphatases that regulate neurite outgrowth identifies PTPN12 as a negative regulator of TrkB tyrosine phosphorylation.
abstract: Alterations in function of the neurotrophin BDNF are associated with neurodegeneration, cognitive decline, and psychiatric disorders. BDNF promotes axonal outgrowth and branching, regulates dendritic tree morphology and is important for axonal regeneration after injury, responses that largely result from activation of its tyrosine kinase receptor TrkB. Although intracellular neurotrophin (NT) signaling presumably reflects the combined action of kinases and phosphatases, little is known about the contributions of the latter to TrkB regulation. The issue is complicated by the fact that phosphatases belong to multiple independently evolved families, which are rarely studied together. We undertook a loss-of-function RNA-interference-based screen of virtually all known (254) human phosphatases to understand their function in BDNF/TrkB-mediated neurite outgrowth in differentiated SH-SY5Y cells. This approach identified phosphatases from diverse families, which either positively or negatively modulate BDNF-TrkB-mediated neurite outgrowth, and most of which have little or no previously established function related to NT signaling. "Classical" protein tyrosine phosphatases (PTPs) accounted for 13% of the candidate regulatory phosphatases. The top classical PTP identified as a negative regulator of BDNF-TrkB-mediated neurite outgrowth was PTPN12 (also called PTP-PEST). Validation and follow-up studies showed that endogenous PTPN12 antagonizes tyrosine phosphorylation of TrkB itself, and the downstream activation of ERK1/2. We also found PTPN12 to negatively regulate phosphorylation of p130cas and FAK, proteins with previously described functions related to cell motility and growth cone behavior. Our data provide the first comprehensive survey of phosphatase function in NT signaling and neurite outgrowth. They reveal the complexity of phosphatase control, with several evolutionarily unrelated phosphatase families cooperating to affect this biological response, and hence the relevance of considering all phosphatase families when mining for potentially druggable targets.
authors.affiliation: Department of Biomedical Sciences and Biotechnology Research and Innovation Centre (BRIC), Faculty of Health Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Malene
authors.initials: M
authors.lastname: Ambjørn
authors.forename: Véronique
authors.initials: V
authors.lastname: Dubreuil
authors.forename: Federico
authors.initials: F
authors.lastname: Miozzo
authors.forename: Fabienne
authors.initials: F
authors.lastname: Nigon
authors.forename: Bente
authors.initials: B
authors.lastname: Møller
authors.forename: Shohreh
authors.initials: S
authors.lastname: Issazadeh-Navikas
authors.forename: Jacob
authors.initials: J
authors.lastname: Berg
authors.forename: Michael
authors.initials: M
authors.lastname: Lees
authors.forename: Jan
authors.initials: J
authors.lastname: Sap
mesh_terms: D019208:Brain-Derived Neurotrophic Factor / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D050719:Crk-Associated Substrate Protein / Q000378:metabolism; D004353:Drug Evaluation, Preclinical / Q000379:methods; D051417:Focal Adhesion Kinase 1 / Q000378:metabolism; D055785:Gene Knockdown Techniques; D006801:Humans; D016501:Neurites / Q000187:drug effects / Q000502:physiology*; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D010641:Phenotype; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D054594:Protein Tyrosine Phosphatase, Non-Receptor Type 12 / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D020813:Receptor, trkB / Q000378:metabolism*; D015203:Reproducibility of Results; D015398:Signal Transduction; D014212:Tretinoin / Q000494:pharmacology
chemical_list: D019208:Brain-Derived Neurotrophic Factor; D050719:Crk-Associated Substrate Protein; D014212:Tretinoin; D020813:Receptor, trkB; D051417:Focal Adhesion Kinase 1; C496050:PTK2 protein, human; D010744:Phosphoric Monoester Hydrolases; D054594:Protein Tyrosine Phosphatase, Non-Receptor Type 12