RhoA Controls Axon Extension Independent of Specification in the Developing Brain

Sebastian Dupraz1, Brett Hilton1, Andreas Husch1, Telma Santos1, Charlotte Coles2, Sina Stern1, Cord Brakebusch3, Frank Bradke1

1 Axonal Growth and Regeneration Group, German Center for Neurodegenerative Diseases (DZNE), Venusberg-Campus 1, Building 99, 53127 Bonn, Germany
2 Present address: Immunocore, Ltd., 101 Park Drive, Milton Park, Abingdon, OX14 4RY Oxfordshire, UK
3 Biotech Research & Innovation Centre, University of Copenhagen, Ole Maaløes Vej 5, 2200 Copenhagen, Denmark

The specification of an axon and its subsequent outgrowth are key steps during neuronal polarization, a prerequisite to wire the brain. The Rho-guanosine triphosphatase (GTPase) RhoA is believed to be a central player in these processes. However, its physiological role has remained undefined. Here, genetic loss- and gain-of-function experiments combined with time-lapse microscopy, cell culture, and in vivo analysis show that RhoA is not involved in axon specification but confines the initiation of neuronal polarization and axon outgrowth during development. Biochemical analysis and super-resolution microscopy together with molecular and pharmacological manipulations reveal that RhoA restrains axon growth by activating myosin-II-mediated actin arc formation in the growth cone to prevent microtubules from protruding toward the leading edge. Through this mechanism, RhoA regulates the duration of axon growth and pause phases, thus controlling the tightly timed extension of developing axons. Thereby, this work unravels physiologically relevant players coordinating actin-microtubule interactions during axon growth.