Abstract 2105: An EWS-FLI1/MRTF gene regulatory network in Ewing sarcoma

Abstract

Abstract The pediatric bone cancer Ewing sarcoma (ES) is characterized by the expression of the chimeric transcription factor EWS-FLI1, derived from a chromosomal translocation t(11;22)(q24;q12). The early onset of metastasis in ES leads to poor overall survival and limited treatment options. Remodeling of the actin cytoskeleton is a prerequisite of a tumor cell in order to become metastatic. Previous studies suggest that EWS-FLI1 alters cell morphology and deregulates various cytoskeletal genes. The major point of convergence of extracellular signals influencing cytoskeletal regulation is the Rho pathway. Activation of Rho GTPases promotes polymerization of monomeric G-actin into stress fibers (F-actin) thereby releasing the myocardin-related transcription factor family (MRTF-A, MRTF-B) of transcriptional coactivators. MRTFs translocate to the nucleus and interact with the transcription factor serum response factor (SRF) bound on CArG promoter elements of genes. However, the ternary complex factor (TCF) family, a class of transcriptional coactivators activated downstream of Ras signaling, competes with MRTFs for SRF binding in presence of ets binding sites adjacent to the CArG consensus sequence. The ets-transcription factor EWS-FLI1 has been shown to substitute for TCFs and interact with SRF. We hypothesized that EWS-FLI1 competes with MRTFs for SRF binding and uncouples target gene expression from Rho signaling. In combinatorial knockdown experiments of MRTFA/B and EWS-FLI1 we demonstrate that global changes in gene expression elicited by EWS-FLI1 silencing largely depend on the presence of MRTFs, suggesting that the fusion oncogene prohibits MRTF activity. Using gene expression analysis we identified six different patterns of MRTF/EWS-FLI1 interaction, with predominance of EWS-FLI1 inversely regulating MRTF target genes. Furthermore, we found MRTF transcriptional effects are already present in the absence of serum in A673 ES cells. MRTFB, however, is cytoplasmic under serum starved conditions and translocates to the nucleus only upon serum stimulation unless latrunculin B (LatB), an actin polymerization inhibitor, is added. In contrast, MRTFA is constitutively nuclear even under serum starved conditions or upon LatB treatment indicating that MRTFA might be uncoupled from actin regulation. These findings reveal a previously unknown role of MRTFs and the “CArGome” in EWS-FLI1 driven transcriptional dysregulation. Supported by the Liddy Shriver Sarcoma Initiative. Citation Format: Anna M. Katschnig, Maximilian Kauer, Raphaela Schwentner, Dave N.T Aryee, Elizabeth Lawlor, Heinrich Kovar. An EWS-FLI1/MRTF gene regulatory network in Ewing sarcoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2105. doi:10.1158/1538-7445.AM2015-2105