Abstract 4459: EWS-FLI1 orchestrates beta-catenin regulation by a mechanism involving post-translational KDM1A stabilization
Authors: Ban Jozef , Aryee Dave N. , Katschnig Anna M. , Schwentner Raphaela , Kauer Maximilian O. , Kovar Heinrich 
Published: August 2018

Beta-catenin (CTNNB) fulfills different functions depending on its subcellular localization. At the membrane it is involved in homotypic cell adhesion. In the nucleus it acts as a transcriptional co-activator, while cytoplasmic CTNNB sequesters and stabilizes the transcriptional co-activator TAZ and prohibits its nuclear translocation. The most extensively characterized CTNNB function is that of a nuclear effector of canonical Wnt signaling, binding and activating TCF/LEF family transcription factors and leading to upregulation of master regulators of epithelial-to-mesenchymal transition. In Ewing sarcoma (EwS), the presence of the driver fusion oncogene EWS-FLI1 renders canonical Wnt signaling unresponsive to Wnts. The exact mechanism of EWS-FLI1 mediated suppression of this signaling pathway is still elusive, but may involve downregulation of the beta-catenin partner transcription factor TCF7L2. In the absence of Wnt/CTNNB/TCF signaling we still find nuclear localization of CTNNB in EwS cells, and demonstrate sensitivity of the tumor cells to the CTNNB targeting cell-permeable oxazole compound iCRT3, suggesting an essential TCF independent role of beta-catenin for EwS survival. The regulation of CTNNB stability is complex and involves phosphorylation and the activity of a multi-protein cytoplasmic destruction complex containing Axin, APC , GSK3B and other proteins. Here, we identify involvement of lysine specific demethylase KDM1A (LSD1) in EWS-FLI1 dependent nuclear CTNNB stabilization. We describe a mechanism, by which EWS-FLI1 directly activates expression of the ubiquitin specific protease USP1 leading to KDM1A post-translational stabilization and ultimately CTNNB nuclear accumulation. Consequently, knockdown of EWS-FLI1 resulted in modulation of USP1, depletion of KDM1A protein, activation of the LSD1 target and negative Wnt signaling regulator DKK1, and cytoplasmic sequestration of CTNNB. Strikingly, in presence of EWS-FLI1, genetic or pharmacologic inhibition of either USP1 or LSD1, or forced DKK1 expression, led to CTNNB degradation. As we find upregulation of TAZ upon EWS-FLI1 knockdown and loss of CTNNB upon modulation of TAZ by siRNA, we speculate that TAZ may be involved in CTNNB cytoplasmic stabilization in absence of EWS-FLI1, while LSD1 is required for nuclear stabilization in presence of the fusion oncogene. These findings indicate complex CTNNB regulation in EwS, orchestrated by EWS-FLI1, and suggest USP1, KDM1A and CTNNB as potential therapeutic targets in EwS.

Citation Format: Jozef Ban, Dave N. Aryee, Anna M. Katschnig, Raphaela Schwentner, Maximilian O. Kauer, Heinrich Kovar. EWS-FLI1 orchestrates beta-catenin regulation by a mechanism involving post-translational KDM1A stabilization [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4459.