Rac1 and RhoA promote neurite outgrowth through formation and stabilization of growth cone point contacts.
Growth cone advance depends on coordinated membrane protrusion and adhesion to the extracellular matrix. Although many studies have addressed the mechanisms responsible for membrane protrusion, the assembly of integrin-dependent adhesion sites known as point contacts remains poorly understood in growth cones. We show balanced Rac1 activity controls both leading edge protrusion and point contact dynamics during neurite outgrowth. Immunocytochemistry and live imaging of paxillin-green fluorescent protein (GFP) showed that inhibiting Rac1 blocked point contact formation, whereas Rac1 overactivation produced small, unstable point contacts. Both inhibition and overactivation of Rac1 reduced the persistence of lamellar protrusions and neurite outgrowth. Inhibition of ROCK (Rho kinase), a RhoA effector, perturbed protrusion and point contact dynamics similar to Rac1 overactivation. Moreover, the repulsive guidance cue Semaphorin 3A, which signals through Rac1, destabilizes point contacts. Together, our data suggest that coordinated Rho GTPase activities regulate neurite outgrowth through point contact formation and stabilization of membrane protrusion.
Woo
Stephanie
S
Department of Anatomy, Program in Cellular and Molecular Biology, University of Wisconsin, Madison, Wisconsin 53706, USA.
Gomez
Timothy M
TM
D000818:Animals; D002448:Cell Adhesion / Q000187:drug effects; D002478:Cells, Cultured; D049452:Green Fluorescent Proteins / Q000235:genetics; D020439:Growth Cones / Q000201:enzymology* / Q000648:ultrastructure; D047908:Intracellular Signaling Peptides and Proteins; D007797:Laminin / Q000494:pharmacology; D008856:Microscopy, Fluorescence; D024462:Myosin Type II / Q000378:metabolism; D016501:Neurites / Q000201:enzymology* / Q000648:ultrastructure; D051419:Paxillin / Q000032:analysis / Q000235:genetics; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors; D011554:Pseudopodia / Q000648:ultrastructure; D011993:Recombinant Fusion Proteins / Q000032:analysis; D040121:Semaphorin-3A / Q000494:pharmacology; D014982:Xenopus laevis; D020830:rac1 GTP-Binding Protein / Q000037:antagonists & inhibitors / Q000378:metabolism*; D054460:rho-Associated Kinases; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D047908:Intracellular Signaling Peptides and Proteins; D007797:Laminin; D051419:Paxillin; D011993:Recombinant Fusion Proteins; D040121:Semaphorin-3A; D049452:Green Fluorescent Proteins; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D024462:Myosin Type II; D020830:rac1 GTP-Binding Protein; D020742:rhoA GTP-Binding Protein