title: Tyrosine phosphorylation of the Rho guanine nucleotide exchange factor Trio regulates netrin-1/DCC-mediated cortical axon outgrowth.
abstract: The chemotropic guidance cue netrin-1 mediates attraction of migrating axons during central nervous system development through the receptor Deleted in Colorectal Cancer (DCC). Downstream of netrin-1, activated Rho GTPases Rac1 and Cdc42 induce cytoskeletal rearrangements within the growth cone. The Rho guanine nucleotide exchange factor (GEF) Trio is essential for Rac1 activation downstream of netrin-1/DCC, but the molecular mechanisms governing Trio activity remain elusive. Here, we demonstrate that Trio is phosphorylated by Src family kinases in the embryonic rat cortex in response to netrin-1. In vitro, Trio was predominantly phosphorylated at Tyr(2622) by the Src kinase Fyn. Though the phospho-null mutant Trio(Y2622F) retained GEF activity toward Rac1, its expression impaired netrin-1-induced Rac1 activation and DCC-mediated neurite outgrowth in N1E-115 neuroblastoma cells. Trio(Y2622F) impaired netrin-1-induced axonal extension in cultured cortical neurons and was unable to colocalize with DCC in growth cones, in contrast to wild-type Trio. Furthermore, depletion of Trio in cortical neurons reduced the level of cell surface DCC in growth cones, which could be restored by expression of wild-type Trio but not Trio(Y2622F). Together, these findings demonstrate that Trio(Y2622) phosphorylation is essential for the regulation of the DCC/Trio signaling complex in cortical neurons during netrin-1-mediated axon outgrowth.
authors.affiliation: Department of Anatomy & Cell Biology, McGill University, Montreal, Quebec, Canada.
authors.forename: Jonathan
authors.initials: J
authors.lastname: DeGeer
authors.forename: Jérôme
authors.initials: J
authors.lastname: Boudeau
authors.forename: Susanne
authors.initials: S
authors.lastname: Schmidt
authors.forename: Fiona
authors.initials: F
authors.lastname: Bedford
authors.forename: Nathalie
authors.initials: N
authors.lastname: Lamarche-Vane
authors.forename: Anne
authors.initials: A
authors.lastname: Debant
mesh_terms: D000818:Animals; D001369:Axons / Q000502:physiology*; D002460:Cell Line; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000196:embryology / Q000502:physiology; D000075389:DCC Receptor; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D009414:Nerve Growth Factors / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000378:metabolism*; D000075388:Netrin-1; D016501:Neurites / Q000502:physiology; D010766:Phosphorylation; D051076:Proto-Oncogene Proteins c-fyn / Q000378:metabolism; D051381:Rats; D011956:Receptors, Cell Surface / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000378:metabolism*; D014443:Tyrosine / Q000737:chemistry / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism; D019061:src-Family Kinases / Q000378:metabolism
chemical_list: D000075389:DCC Receptor; C501912:Dcc protein, rat; D020662:Guanine Nucleotide Exchange Factors; C000617351:NTN1 protein, human; D009414:Nerve Growth Factors; D009419:Nerve Tissue Proteins; C000617354:Ntn1 protein, rat; D011956:Receptors, Cell Surface; C500736:Trio protein, rat; D025521:Tumor Suppressor Proteins; D000075388:Netrin-1; D014443:Tyrosine; C494961:Fyn protein, rat; D051076:Proto-Oncogene Proteins c-fyn; D019061:src-Family Kinases; C499134:Rac1 protein, rat; D020830:rac1 GTP-Binding Protein