title: A signaling mechanism coupling netrin-1/deleted in colorectal cancer chemoattraction to SNARE-mediated exocytosis in axonal growth cones.
abstract: Directed cell migration and axonal guidance are essential steps in neural development. Both processes are controlled by specific guidance cues that activate the signaling cascades that ultimately control cytoskeletal dynamics. Another essential step in migration and axonal guidance is the regulation of plasmalemma turnover and exocytosis in leading edges and growth cones. However, the cross talk mechanisms linking guidance receptors and membrane exocytosis are not understood. Netrin-1 is a chemoattractive cue required for the formation of commissural pathways. Here, we show that the Netrin-1 receptor deleted in colorectal cancer (DCC) forms a protein complex with the t-SNARE (target SNARE) protein Syntaxin-1 (Sytx1). This interaction is Netrin-1 dependent both in vitro and in vivo, and requires specific Sytx1 and DCC domains. Blockade of Sytx1 function by using botulinum toxins abolished Netrin-1-dependent chemoattraction of axons in mouse neuronal cultures. Similar loss-of-function experiments in the chicken spinal cord in vivo using dominant-negative Sytx1 constructs or RNAi led to defects in commissural axon pathfinding reminiscent to those described in Netrin-1 and DCC loss-of-function models. We also show that Netrin-1 elicits exocytosis at growth cones in a Sytx1-dependent manner. Moreover, we demonstrate that the Sytx1/DCC complex associates with the v-SNARE (vesicle SNARE) tetanus neurotoxin-insensitive vesicle-associated membrane protein (TI-VAMP) and that knockdown of TI-VAMP in the commissural pathway in the spinal cord results in aberrant axonal guidance phenotypes. Our data provide evidence of a new signaling mechanism that couples chemotropic Netrin-1/DCC axonal guidance and Sytx1/TI-VAMP SNARE proteins regulating membrane turnover and exocytosis.
authors: Developmental Neurobiology and Regeneration Unit, Institut de Recerca Biomèdica Barcelona, Parc Cientific de Barcelona, Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (Instituto de Salud Carlos III), Department of Cell Biology, University of Barcelona, E-08028 Barcelona, Spain.
authors: Tiziana
authors: T
authors: Cotrufo
authors: Francesc
authors: F
authors: Pérez-Brangulí
authors: Ashraf
authors: A
authors: Muhaisen
authors: Oriol
authors: O
authors: Ros
authors: Rosa
authors: R
authors: Andrés
authors: Thomas
authors: T
authors: Baeriswyl
authors: Giulia
authors: G
authors: Fuschini
authors: Teresa
authors: T
authors: Tarrago
authors: Marta
authors: M
authors: Pascual
authors: Jesús
authors: J
authors: Ureña
authors: Joan
authors: J
authors: Blasi
authors: Ernest
authors: E
authors: Giralt
authors: Esther T
authors: ET
authors: Stoeckli
authors: Eduardo
authors: E
authors: Soriano
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D000831:Animals, Newborn; D001369:Axons / Q000187:drug effects / Q000502:physiology; D001896:Boron Compounds / Q000378:metabolism; D019274:Botulinum Toxins, Type A / Q000494:pharmacology; D019208:Brain-Derived Neurotrophic Factor / Q000494:pharmacology; D002478:Cells, Cultured; D002633:Chemotaxis / Q000187:drug effects; D002522:Chlorocebus aethiops; D003172:Complement C1 / Q000494:pharmacology; D000075389:DCC Receptor; D004622:Embryo, Mammalian; D005089:Exocytosis / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D020439:Growth Cones / Q000187:drug effects / Q000502:physiology*; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D047468:Immunoprecipitation; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D051938:Munc18 Proteins / Q000235:genetics / Q000378:metabolism; D009414:Nerve Growth Factors / Q000235:genetics / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D000075388:Netrin-1; D009465:Neuromuscular Agents / Q000494:pharmacology; D009474:Neurons / Q000166:cytology*; D009924:Organ Culture Techniques; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D050600:SNARE Proteins / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics*; D020349:Surface Plasmon Resonance / Q000379:methods; D013744:Tetanus Toxin / Q000494:pharmacology; D014162:Transfection / Q000379:methods; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*; D050988:Vesicle-Associated Membrane Protein 2 / Q000378:metabolism
chemical_list: C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D001896:Boron Compounds; D019208:Brain-Derived Neurotrophic Factor; D003172:Complement C1; D000075389:DCC Receptor; C501910:Dcc protein, mouse; D020662:Guanine Nucleotide Exchange Factors; D051938:Munc18 Proteins; C000617351:NTN1 protein, human; D009414:Nerve Growth Factors; D009419:Nerve Tissue Proteins; D009465:Neuromuscular Agents; C000617353:Ntn1 protein, mouse; D011956:Receptors, Cell Surface; D050600:SNARE Proteins; C511269:Syx protein, mouse; D013744:Tetanus Toxin; D025521:Tumor Suppressor Proteins; D050988:Vesicle-Associated Membrane Protein 2; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D000075388:Netrin-1; D019274:Botulinum Toxins, Type A