Oncogenic Targets of Hedgehog-GL/1 Activation in Ewing Sarcoma
Ewing Tumors are addicted to aberrant oncogenic transcription factors of the EWS/FLI1 class. Means to target these proteins face substantial technical and economic obstacles. However, my lab and others have shown the importance of Hedgehog-GLI (HH-GLI) signaling in Ewing tumor biology. More recently, data from my lab has shown that a majority of targets transcriptionally deregulated by EWS/FLI1 are co-regulated by GLI1, demonstrating the central importance of HH-GLI in Ewings. We now have evidence that the reason for this tremendous target overlap involves a novel mechanism in which GLI1 interacts with EWS/FLI1 to synergistically upregulate jointly-targeted promoters. This synergism represents a novel aspect of EWS/FLI1 and GLI1 biology that can have tremendous translational implications for Ewing tumors since it will add key details to the only proven driver mutation in this disease. We propose to detail the transcriptional mechanisms by which this interaction occurs. We will analyze specific target promoters and mutagenize specific GLI1 and EWS/FLI1 binding sites to measure their effects on the observed transcriptional synergy. Identified sites will be confirmed by CHIP analysis to demonstrate that this process also occurs in vivo. More extensive analysis with CHIP-Seq will be undertaken, time permitting. We will use the knowledge generated from these studies to devise a reporter system which will permit large scale screening for compounds to interfere with the EWS/FL1-GLI1 interaction. Finally, we will explore the nature of this interaction by confirming our preliminary data that this interaction involves direct physical association between GLI1 and EWS/FLI. Structural mutants will be used to further detail the important moieties involved.