Title: Transcriptional regulation of Olig2 in motor neuron development
Description: A fundamental question in the field of neurobiology is how to generate specific types of neurons in the central nervous system (CNS). Understanding the mechanism underlying the generation of neuronal diversity would be of potential use in the design of new strategies to regenerate CNS tissue. Specifically, if we could understand how to induce neural stem cells to develop in vitro into specific subtypes of neurons and glia, they may have therapeutic potential for patients with neurological diseases to promote functional recovery. One such gene that promotes motoneuron differentiation from neural progenitors is Olig2 . Olig2 is a basic helix-loop-helix (bHLH) transcription factor that is specifically expressed in motoneuron precursors, and is both necessary and sufficient for the differentiation of motoneurons. The underlying mechanism activating Olig2 expression is unknown. To address this question, we first identified a regulatory element that controlled Olig2 expression in motoneuron progenitors. Using Cre recombinase to fate map the lineage of cells derived from the Olig2 regulatory element, I showed that it directed reporter expression in motoneurons, but not oligodendrocytes, indicating motoneuron-lineage specific regulation of Olig2 . This showed that the regulation of Olig2 transcription is complex, and governed by distinct neural and glial-specific regulatory elements. Further, it suggests that motoneurons are derived from a transit-amplifying motoneuron progenitor.

The Olig2/K23 regulatory element provided a unique opportunity for identifying the signals that regulate Olig2 expression. Specifically, I used the Olig2/K23 regulatory element to identify homeodomain transcription factors that act upstream of Olig2 in the specification of motoneuron progenitors. Analyzing the Olig2/K23 regulatory element for transcription factor binding sites revealed conserved Nkx6, Hox , and Meis binding sites. Here, I demonstrate that Nkx6.3 , a member of the Nkx6 class of transcription factors is sufficient for the induction of ectopic V2 interneurons with repression of Olig2 , and motoneuron development. As motoneuron and V2 interneuron progenitors both initially express Nkx6.1 , these data suggest a novel mechanism by which Nkx6.3 expression induces hindbrain V2 interneuron development while antagonizing the premature expression of motoneuron genes. In the next study I investigated a general function for HoxA and Meis in activating Olig2 expression. Expression analysis confirmed that multiple members of the HoxA and Meis subfamilies were appropriately expressed in the pMN ventricular zone at E9.5 in an Olig2 -independent manner, consistent with a role upstream of Olig2 . Misexpression of either HoxA3 or Meis1 did not result in ectopic expression of endogenous Olig2 of the chick spinal cord. However, co-expression of HoxA3 and Meis1 resulted in ectopic expression of Olig2 , and motoneuron development in the dorsal spinal cord. Together, these data suggest that interactions of HoxA proteins together with Meis1 may account for activation of Olig2 , and specification of motoneuron progenitors from neural stem cells.