Flavin monooxygenases regulate Caenorhabditis elegans axon guidance and growth cone protrusion with UNC-6/Netrin signaling and Rac GTPases

• Published in: PLoS genetics 2017-08, Vol.13 (8), p.e1006998-e1006998
• Main Authors: Gujar, Mahekta R, Stricker, Aubrie M, Lundquist, Erik A
• Format: Article
• Language: English
• Subjects: Actin; Animal behavior; Animals; Animals, Genetically Modified; Axon guidance; Axon Guidance - genetics; Biology and Life Sciences; Caenorhabditis elegans - genetics; Caenorhabditis elegans - growth & development; Caenorhabditis elegans Proteins - genetics; Cones; Cytoskeleton; DCC protein; Depolymerization; Developmental biology; Dinitrocresols - metabolism; Flavin; Genomes; Growth cones; Guanosine triphosphatases; Guanosine triphosphate; Insects; Invertebrates; Medicine and Health Sciences; Mixed Function Oxygenases - genetics; Monooxygenase; Mutation; Nematodes; Nerve Tissue Proteins - genetics; Netrins; Oxidation; Phosphorylation; Pseudopodia - genetics; Pseudopodia - metabolism; rac GTP-Binding Proteins - genetics; Research and Analysis Methods; Signal Transduction
• ISSN: 1553-7404; 1553-7390; 1553-7404
• DOI: 10.1371/journal.pgen.1006998

The guidance cue UNC-6/Netrin regulates both attractive and repulsive axon guidance. Our previous work showed that in C. elegans, the attractive UNC-6/Netrin receptor UNC-40/DCC stimulates growth cone protrusion, and that the repulsive receptor, an UNC-5:UNC-40 heterodimer, inhibits growth cone protrusion. We have also shown that inhibition of growth cone protrusion downstream of the UNC-5:UNC-40 repulsive receptor involves Rac GTPases, the Rac GTP exchange factor UNC-73/Trio, and the cytoskeletal regulator UNC-33/CRMP, which mediates Semaphorin-induced growth cone collapse in other systems. The multidomain flavoprotein monooxygenase (FMO) MICAL (Molecule Interacting with CasL) also mediates growth cone collapse in response to Semaphorin by directly oxidizing F-actin, resulting in depolymerization. The C. elegans genome does not encode a multidomain MICAL-like molecule, but does encode five flavin monooxygenases (FMO-1, -2, -3, -4, and 5) and another molecule, EHBP-1, similar to the non-FMO portion of MICAL. Here we show that FMO-1, FMO-4, FMO-5, and EHBP-1 may play a role in UNC-6/Netrin directed repulsive guidance mediated through UNC-40 and UNC-5 receptors. Mutations in fmo-1, fmo-4, fmo-5, and ehbp-1 showed VD/DD axon guidance and branching defects, and variably enhanced unc-40 and unc-5 VD/DD axon guidance defects. Developing growth cones in vivo of fmo-1, fmo-4, fmo-5, and ehbp-1 mutants displayed excessive filopodial protrusion, and transgenic expression of FMO-5 inhibited growth cone protrusion. Mutations suppressed growth cone inhibition caused by activated UNC-40 and UNC-5 signaling, and activated Rac GTPase CED-10 and MIG-2, suggesting that these molecules are required downstream of UNC-6/Netrin receptors and Rac GTPases. From these studies we conclude that FMO-1, FMO-4, FMO-5, and EHBP-1 represent new players downstream of UNC-6/Netrin receptors and Rac GTPases that inhibit growth cone filopodial protrusion in repulsive axon guidance.