title: FLIM FRET Visualization of Cdc42 Activation by Netrin-1 in Embryonic Spinal Commissural Neuron Growth Cones.
abstract: Netrin-1 is an essential extracellular chemoattractant that signals through its receptor DCC to guide commissural axon extension in the embryonic spinal cord. DCC directs the organization of F-actin in growth cones by activating an intracellular protein complex that includes the Rho GTPase Cdc42, a critical regulator of cell polarity and directional migration. To address the spatial distribution of signaling events downstream of netrin-1, we expressed the FRET biosensor Raichu-Cdc42 in cultured embryonic rat spinal commissural neurons. Using FLIM-FRET imaging we detected rapid activation of Cdc42 in neuronal growth cones following application of netrin-1. Investigating the signaling mechanisms that control Cdc42 activation by netrin-1, we demonstrate that netrin-1 rapidly enriches DCC at the leading edge of commissural neuron growth cones and that netrin-1 induced activation of Cdc42 in the growth cone is blocked by inhibiting src family kinase signaling. These findings reveal the activation of Cdc42 in embryonic spinal commissural axon growth cones and support the conclusion that src family kinase activation downstream of DCC is required for Cdc42 activation by netrin-1.
authors.affiliation: Program in NeuroEngineering, McGill University, Montreal, QC, H3A 2B4, Canada.|Department of Neurology and Neurosurgery, Montréal Neurological Institute, McGill University, Montreal, QC, H3A 2B4, Canada.|Department of Physics, McGill University, Montreal, QC, H3A 2T8, Canada.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Rappaz
authors.affiliation: Program in NeuroEngineering, McGill University, Montreal, QC, H3A 2B4, Canada.|Department of Neurology and Neurosurgery, Montréal Neurological Institute, McGill University, Montreal, QC, H3A 2B4, Canada.
authors.forename: Karen
authors.initials: K
authors.lastname: Lai Wing Sun
authors.affiliation: Program in NeuroEngineering, McGill University, Montreal, QC, H3A 2B4, Canada.|Department of Neurology and Neurosurgery, Montréal Neurological Institute, McGill University, Montreal, QC, H3A 2B4, Canada.
authors.forename: James P
authors.initials: JP
authors.lastname: Correia
authors.affiliation: Program in NeuroEngineering, McGill University, Montreal, QC, H3A 2B4, Canada.|Department of Physics, McGill University, Montreal, QC, H3A 2T8, Canada.|Department of Chemistry, McGill University, Montreal, QC, H3A 0B8, Canada.
authors.forename: Paul W
authors.initials: PW
authors.lastname: Wiseman
authors.affiliation: Program in NeuroEngineering, McGill University, Montreal, QC, H3A 2B4, Canada.|Department of Neurology and Neurosurgery, Montréal Neurological Institute, McGill University, Montreal, QC, H3A 2B4, Canada.
authors.forename: Timothy E
authors.initials: TE
authors.lastname: Kennedy
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D000075389:DCC Receptor; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D020439:Growth Cones / Q000378:metabolism / Q000648:ultrastructure*; D042282:Microdissection; D008856:Microscopy, Fluorescence / Q000379:methods*; D009414:Nerve Growth Factors / Q000032:analysis* / Q000378:metabolism; D000075388:Netrin-1; D017207:Rats, Sprague-Dawley; D011956:Receptors, Cell Surface / Q000032:analysis / Q000378:metabolism; D015398:Signal Transduction; D013116:Spinal Cord / Q000166:cytology / Q000196:embryology* / Q000378:metabolism / Q000648:ultrastructure; D025521:Tumor Suppressor Proteins / Q000032:analysis* / Q000378:metabolism; D020764:cdc42 GTP-Binding Protein / Q000032:analysis* / Q000378:metabolism; D019061:src-Family Kinases / Q000378:metabolism
chemical_list: D000075389:DCC Receptor; C501912:Dcc protein, rat; D009414:Nerve Growth Factors; C000617354:Ntn1 protein, rat; D011956:Receptors, Cell Surface; D025521:Tumor Suppressor Proteins; D000075388:Netrin-1; D019061:src-Family Kinases; D020764:cdc42 GTP-Binding Protein