Genetic evidence for a contribution of EphA:EphrinA reverse signaling to motor axon guidance

Repulsive Eph forward signaling from limb-derived ephrins guides the axons of lateral motor column (LMC) motor neurons. LMC axons also express ephrinAs, while theirEphAreceptors are expressed in the limb mesenchyme. In vitro studies have suggested that reverse signaling from limb-derived EphA4 to axonal ephrinAs might result in attraction ofLMCaxons. However, genetic evidence for this function is lacking. Here we use theDunnchamber turning assay to show thatEphAproteins are chemoattractants and elicit fast turning responses inLMCneurons in vitro. Moreover, ectopic expression ofEphA4in chick hindlimb changes the limb trajectory ofLMCaxons. Nervous system-specific deletion ofEphA4 in mice resulted in fewer LMC axon projection errors than the ubiquitous deletion of EphA4. Additionally, a signaling-incompetent EphA4 mutant partially rescued guidance errors in the hindlimb, suggesting that limb-derived EphA4 contributes to the establishment ofLMCprojections. In summary, we provide evidence for a role of EphA:ephrinA attractive reverse signaling in motor axon guidance and in vivo evidence of in-parallel forward Eph and reverse ephrin signaling function in the same neuronal population.