In the developing spinal cord, the ventral neural tube gives rise to motor neurons (MNs) and four classes of interneuron (V0–V3). These are specified in five dorsoventrally arranged progenitor domains (pMN and p0–p3), which are delineated by the expression of class I and class II homeodomain (HD) genes, including members of the Pax, Nkx, Dbx and Irx families. Their expression patterns are established by graded responses to sonic hedgehog ( Shh) signalling from the notochord and floorplate, and the interdomain boundaries are sharpened and maintained by mutual repression between pairs of dorsally expressed class I and ventrally expressed class II proteins. The HD proteins also control neuronal specification by regulating the expression of subtype determinants, and in a new study reported in Cell, Muhr et al. present evidence that this role is an extension of their repressor activity.

Eight of the ten known progenitor HD genes possess a conserved eh1 domain that recruits co-repressors of the Groucho-TLE (Gro/TLE) family. In an in vitro reporter assay, the authors showed that the repressor functions of class II Nkx proteins depend on the presence of the eh1 domain. They went on to test whether the eh1 domain is also essential in vivo, both for the repression of class I genes and for the promotion of neuronal subtype determinant expression. By electroporating full-length or eh1-deleted Nkx genes into the neural tube of a chick embryo, they showed that misexpression of Nkx2.2 or Nkx6.1 caused downregulation of class I HD genes ( Pax6 and Dbx2 respectively), but only if the eh1 domain was present. Also, misexpression of Nkx6.1 normally induces ectopic expression of the MN determinant MNR2, but this effect was abolished if eh1 was deleted. The spatiotemporal expression of Gro/TLE co-repressors in the neural tube is consistent with a role in neuronal specification, and the authors showed that ectopic expression of Grg5, an endogenous dominant-negative Gro/TLE inhibitor, prevented mutual repression between the class I/class II pairs. Also, although Grg5 expression caused dorsal spreading of the Nkx6.1 expression domain, this did not lead to ectopic specification of MNs, confirming that this too requires Gro/TLE co-repressor activity.

On the basis of these findings, Muhr et al. propose a derepression model for specification of neuronal fate in the neural tube. The pMN domain illustrates how this might work; here, Nkx6.1 directly represses expression of the V1 interneuron determinant and also represses Dbx2 expression, preventing its repression of the gene encoding MNR2. Simultaneously, Pax6, presumably acting through an intermediary repressor because it does not possess an eh1 domain, inhibits expression of the V3 interneuron determinant. In this way, MNs are specified by derepression of MNR2, while alternative neuronal phenotypes are suppressed. Similar repressive interactions are thought to contribute to neuronal specification in the other progenitor domains.

This model undoubtedly enhances our understanding of neuronal specification in the neural tube, but the question of how the neuronal subtype determinant genes become activated in the first place remains to be answered. Therefore, there is still some way to go before we identify the root of the pathway.