Abstract
The molecular hallmark of the Ewing's family of tumors is the presence of balanced chromosomal translocations, leading to the formation of chimerical transcription factors (that is, EWS/FLI1) that play a pivotal role in the pathogenesis of Ewing's tumors by deregulating gene expression. We have recently demonstrated that DAX1 (NR0B1), an orphan nuclear receptor that was not previously implicated in cancer, is induced by the EWS/FLI1 oncoprotein and is highly expressed in Ewing's tumors, suggesting that DAX1 is a biologically relevant target of EWS/FLI1-mediated oncogenesis. In this study we demonstrate that DAX1 is a direct transcriptional target of the EWS/FLI1 oncoprotein through its binding to a GGAA-rich region in the DAX1 promoter and show that DAX1 is a key player of EWS/FLI1-mediated oncogenesis. DAX1 silencing using an inducible model of RNA interference induces growth arrest in the A673 Ewing's cell line and severely impairs its capability to grow in semisolid medium and form tumors in immunodeficient mice. Gene expression profile analysis demonstrated that about 10% of the genes regulated by EWS/FLI1 in Ewing's cells are DAX1 targets, confirming the importance of DAX1 in Ewing's oncogenesis. Functional genomic analysis, validated by quantitative RT–PCR, showed that genes implicated in cell-cycle progression, such as CDK2, CDC6, MCM10 or SKP2 were similarly regulated by EWS/FLI1 and DAX1. These findings indicate that DAX1 is important in the pathogenesis of the Ewing's family of tumors, identify new functions for DAX1 as a cell-cycle progression regulator and open the possibility to new therapeutic approaches based on DAX1 function interference.
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Acknowledgements
We are grateful to B Bardoni for gift of the pDAX-1 PROM.CAT plasmid and CT Denny for gift of the pSRα EWS/FLI1 and the EWS/FLI1 mutants pSRα EWS/FLI1 Δ65 and pSRα EWS/FLI1 triple mutant used in this study. This study was funded by Ministerio de Educación y Ciencia grants SAF2006-07586 and SAF2007-62101; Fundación Inocente Inocente, Fundación Enriqueta Villavecchia and Fundación Científica Asociación Española Contra el Cáncer. E Lalli is supported by Fondation pour la Recherche Médicale, Association Recherche sur le Cancer and a CNRS-ATIP grant; E García-Aragoncillo and J Carrillo are predoctoral fellows from the Ministerio de Educación y Ciencia. N Agra is a predoctoral fellow from the Fundación General de la U.A.M; G Gómez-López is supported by a contract from de Fondo de Investigaciones Sanitarias; J Alonso has been supported by a Ramón y Cajal contract from the Ministerio de Educación y Ciencia.
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Supplementary Information accompanies the paper on the Oncogene website (http://www.nature.com/onc)
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García-Aragoncillo, E., Carrillo, J., Lalli, E. et al. DAX1, a direct target of EWS/FLI1 oncoprotein, is a principal regulator of cell-cycle progression in Ewing's tumor cells. Oncogene 27, 6034–6043 (2008). https://doi.org/10.1038/onc.2008.203
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DOI: https://doi.org/10.1038/onc.2008.203
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