| In the developing nervous system, axons navigate long distances through a complex environment using intermediate targets to reach their final target. The midline of the central nervous system is one of the most highly studied intermediate targets. Commissural axons are first attracted to the ventral midline, but upon crossing the floor plate, they become repelled by the midline. We first took a genomic microarray approach to identify novel regulators of axon guidance at the midline. We next developed a method to test candidate genes using siRNA knockdown in whole mouse embryo culture. In this siRNA screen, the receptor tyrosine kinase, Kit, was found to be required for axon guidance in commissural neurons postcrossing. Kit and its ligand, Stem Cell Factor (SCF), are specifically expressed in commissural neurons and the floor plate, respectively. Dil Analysis in Kit and Steel mutant mice show the majority of commissural axons abnormally stalled in the floor plate. Furthermore, in vitro explant experiments show SCF promotes growth in commissural axons postcrossing but not precrossing. Unexpectedly, we find that very high levels of Netrin1 also promote robust growth postcrossing, and in Netrin1 mutants, the few commissural axons that do reach the midline behave like those in Kit and Steel mutants, lining up at the contralateral floor plate edge. Our findings identify SCF as a novel regulator of axon growth in the CNS, provide evidence that leaving intermediate targets requires not only activation of repulsive mechanisms but also the action of permissive factors for exit, and implicate SCF and Netrin1 in this function. |
