Trio and CRMP2 regulate axon branching and Semaphorin3A signaling

Trio is a neuronally expressed, Rac1- and RhoA-activating RhoGEF, that is required for neurodevelopment. Mutations affecting the Rac1-activating GEF domain of Trio are associated with profound neurodevelopmental delay and Trio knock-out is embryonic lethal. Although there are studies showing a role for Trio in axon patterning, our understanding of the mechanistic underpinnings of Trio function is incomplete. We have now taken an unbiased approach to identifying the interactome of Trio in embryonic axonal compartments. Using immunoprecipitation-mass spectrometry, we identified the Collapsin Response Mediator Protein 2 (CRMP2) as a robust association partner of growth cone-localized Trio. Like Trio, CRMP2 has a well-known role in shaping the cytoskeleton, particularly during axon patterning. In the current study, we demonstrate Trio preferentially interacts with phosphorylated CRMP2 (pCRMP2) and is recruited by pCRMP2 to limit filopodial motility and axon branching. By introducing a GEF1-ablating disease-related mutation, we further demonstrate that Trio-GEF1 signaling is required for pCRMP2-mediated axon branch suppression. Finally, we show that Semaphorin3A invokes pCRMP2-Trio signaling to limit axon branching in vitro, revealing a developmental role for pCRMP2-Trio signaling.