Proteolytic cleavage of Slit by the Tolkin protease converts an axon repulsion cue to an axon growth cue in vivo

Slit is a secreted protein that has a canonical function of repelling growing axons from the CNS midline. The full-length Slit (Slit-FL) is cleaved into Slit-N and Slit-C fragments, which have potentially distinct functions via different receptors. Here we report that the BMP-1/Tolloid family metalloprotease, Tolkin (Tok), is responsible for Slit proteolysis in vivo and in vitro. In tok mutants lacking Slit cleavage, midline repulsion of axons occurs normally, confirming that Slit-FL is sufficient to repel axons. However, longitudinal axon guidance is highly disrupted in tok mutants and can be rescued by midline expression of Slit-N, suggesting that Slit is the primary substrate for Tok in the embryonic CNS. Transgenic restoration of Slit-N or Slit-C does repel axons in Slit-null animals. Slit-FL and Slit-N are both biologically active cues with distinct axon guidance functions in vivo. Slit signaling is used in diverse biological processes, thus differentiating between Slit-FL and Slit fragments will be essential for evaluating Slit function in broader contexts.