Abstract
Growth cones navigate axonal projection in response to guidance cues. However, it is unclear how they can decide the migratory direction by transducing the local spatial cues into protrusive forces. Here we show that knockout mice of Shootin1 display abnormal projection of the forebrain commissural axons, a phenotype similar to that of the axon guidance molecule netrin-1. Shallow gradients of netrin-1 elicited highly polarized Pak1-mediated phosphorylation of shootin1 within growth cones. We demonstrate that netrin-1-elicited shootin1 phosphorylation increases shootin1 interaction with the cell adhesion molecule L1-CAM; this, in turn, promotes F-actin-adhesion coupling and concomitant generation of forces for growth cone migration. Moreover, the spatially regulated shootin1 phosphorylation within growth cones is required for axon turning induced by netrin-1 gradients. Our study defines a mechano-effector for netrin-1 signaling and demonstrates that shootin1 phosphorylation is a critical readout for netrin-1 gradients that results in a directional mechanoresponse for axon guidance.
Highlights
Axon guidance is a critical step for the formation and regeneration of neuronal networks
Using shootin1a (1-125) as a dominant negative mutant, we examined whether the shootin1a–L1-CAM interaction is involved in netrin-1–induced mechanical coupling between F-actin flow and the substrate
These results demonstrate that the disturbance of axon turning caused by depletion of shootin1a (Figure 7D and Figure 7—figure supplement 2D) or by dominant negative shootin1a (Figure 7—figure supplement 3B) is not attributed only to the inhibited axon outgrowth, and suggest that the polarized phosphorylation of shootin1a within growth cones is required for the directional axon guidance induced by netrin-1 gradients (Figure 8E)
Summary
Axon guidance is a critical step for the formation and regeneration of neuronal networks. If the internal skeleton is the engine of the axon, L1-CAMs are the wheels, and shootin1a the clutch It is not clear whether shootin1a is involved in guiding growing axons, and how it could help neurons ‘understand’ and react to gradients of netrin-1. We show that shootin1a is expressed at high levels in developing forebrain commissural axons and that Shootin knockout mice display abnormal guidance of these axons, a phenotype similar to that of Netrin-1 knockout mice. Disturbance of the spatially regulated shootin1a phosphorylation within growth cones inhibited axon turning, but not axon outgrowth, induced by netrin-1 gradients. Our data demonstrate that shootin1a, through its spatially regulated phosphorylation within growth cones, mediates the gradient reading and mechanoresponse for netrin-1–induced axon guidance
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