Nkx6 genes pattern the frog neural plate and Nkx6.1 is necessary for motoneuron axon projection

Neuronal subtypes originate from an undifferentiated neural epithelium that is progressively divided into progenitor domains by homeodomain transcription factors such as members of the Nkx family. Here we report the functional analysis of Nkx6.1 and Nkx6.2 in Xenopus. While Nkx6.2 is expressed early in a large region of the medial neural plate, Nkx6.1 is restricted to a region overlapping with the region of motor neuron formation. By mRNA injection we show that both can inhibit primary neurogenesis as well as expression of intermediate neural plate markers. However, they do not form auto-regulatory loops and fail to induce ectopic motor neurons as they do in the chick. Using morpholino-mediated knockdown in Xenopus laevis and Xenopus tropicalis we show that Nkx6.1 knockdown results in paralyzed tadpoles. Using DiI labeling and immunohistochemistry we show that the underlying mechanism is a failure of spinal motor neurons to extend axons to their targets. Analysis of neural pattern reveals that ventral Lhx3+ and Pax2+ interneurons are dependent on Nkx6.1 function, but overall neural patterning is not. This study illustrates that while important aspects of Nkx6 gene function are conserved in vertebrate neural patterning, others are not.