Switching from Shh-Mediated Attraction to Repulsion

Science Signaling  04 Dec 2012:
Vol. 5, Issue 253, pp. ec311
DOI: 10.1126/scisignal.2003836

In the developing nervous system, multiple signals mediate axon attraction or repulsion to guide axons to the final destinations where they make synaptic connections. Sonic hedgehog (Shh) functions as a morphogen and as a guidance cue in the developing nervous system. In the developing spinal cord, Shh produced by the floorplate initially directly attracts commissural axons to the midline and after midline crossing indirectly enables responsiveness to repulsive cues mediated by other guidance ligands, which prevents the axons from recrossing the midline. These axons then turn and migrate anteriorly. Using a spinal cord preparation called the open-book format, Yam et al. found that genetic disruption of Shh signaling in commissural neurons resulted in random movement of axons both anteriorly and posteriorly after crossing the midline, as well as increased frequency of axons that inappropriately recrossed the floorplate. As in the developing chick brain, Shh in the rodent brain was present in a gradient along the anterior-posterior axis, with the abundance highest at the posterior end. Rat commissural neurons cultured in vitro were initially attracted to Shh, then repulsed after 3 days in vitro (DIV). The activity of protein kinase A (PKA) and its inhibition by 14-3-3 proteins has been implicated in axon repulsion. Older cultured commissural neurons (3 DIV) and postcrossing axons in mouse spinal cord showed enriched abundance of the 14-3-3 isoforms β and γ and reduced PKA activity. In 3 DIV cultures, inhibition of 14-3-3 function by knockdown or a function-blocking peptide or stimulation of PKA activity converted the Shh-mediated repulsion to attraction, and inhibition of PKA activity in 2 DIV cultures switched their response to repulsion. Disruption of 14-3-3 activity in the open-book format preparations of rat or chick spinal cord reduced the frequency of anterior turning of axons that crossed the midline. In 2 DIV cultures, overexpression of 14-3-3β prevented Shh-mediated attraction, whereas overexpression of 14-3-3γ converted the response to Shh-mediated repulsion. In the developing chick, overexpression of either 14-3-3 isoform resulted in premature turning of axons before reaching the floorplate (the source of Shh), consistent with a premature switch to Shh-mediated repulsion. Thus, changes in the abundance of 14-3-3 to inhibit PKA activity appear to underlie the switch from Shh-mediated attraction to Shh-mediated repulsion in commissural axons.

P. T. Yam, C. B. Kent, S. Morin, W. T. Farmer, R. Alchini, L. Lepelletier, D. R. Colman, M. Tessier-Lavigne, A. E. Fournier, F. Charron, 14-3-3 proteins regulate a cell-intrinsic switch from Sonic hedgehog-mediated commissural axon attraction to repulsion after midline crossing. Neuron 76, 735–749 (2012). [PubMed]