Transcriptional control of early neuronal specification in the embryonic midbrain ”

The complex array of neuronal connections in the vertebrate brain derives from a simple, conserved scaffold of axon tracts in the early embryonic brain. The mechanisms governing the formation of these tracts are largely unknown. We are interested in the molecular cues responsible for the formation of the early axon tracts, in particular in linking transcription factors that are expressed in a spatially restricted manner with the specification of neuronal fates. To this aim, we have first characterised the expression patterns of several homeodomain transcription factors like Pax6, Six3, Emx2 and Sax1 in the ventral midbrain-forebrain border. From this area, two prominent early axon tracts are derived: the caudad extending medial longitudinal fascicle (mlf) and the dorsad, contralaterally projecting posterior commissure (pc). We could demonstrate that Sax1 is expressed in the nucleus of the mlf, and subsequently used electroporation in the chick to analyse the role of Sax1 in the development of this tract. Ectopic expression of a Sax1 expression construct leads to an enlargement of the mlf, while conversely VP16-Sax1, in which the eh1-like transrepression domain of Sax1 has been replaced with the transactivation domain of Herpes simplex VP16, results in a reduction of the mlf. The morphological phenotype is mirrored by changes in gene expression, as Sax1 can ectopically activate Emx2 transcription, while VP16-Sax1 represses the expression of Emx2 and Six3. Our results show a critical role for Sax1 in the formation of the mlf, and point towards the positional coding of neuronal fate by combinations of homeobox genes, analogous to recent finding in hindbrain and spinal cord.