Mendiola Marta, Carrillo Jaime, García Eva, Lalli Enzo, Hernández Teresa, de Alava Enrique, Tirode Franck, Delattre Olivier, García-Miguel Purificación, López-Barea Fernando, Pestaña Angel, Alonso Javier
Departamento de Biología Molecular y Celular del Cáncer, Instituto de Investigaciones Biomédicas A. Sols CSIC-UAM, 28029 Madrid, Spain.
Int J Cancer. 2006 Mar 15;118(6):1381-9. doi: 10.1002/ijc.21578.
The Ewing family of tumors harbors chromosomal translocations that join the N-terminal region of the EWS gene with the C-terminal region of several transcription factors of the ETS family, mainly FLI1, resulting in chimeric transcription factors that play a pivotal role in the pathogenesis of Ewing tumors. To identify downstream targets of the EWS/FLI1 fusion protein, we established 293 cells expressing constitutively either the chimeric EWS/FLI1 or wild type FLI1 proteins and used cDNA arrays to identify genes differentially regulated by EWS/FLI1. DAX1 (NR0B1), an unusual orphan nuclear receptor involved in gonadal development, sex determination and steroidogenesis, showed a consistent up-regulation by EWS/FLI1 oncoprotein, but not by wild type FLI1. Specific induction of DAX1 by EWS/FLI1 was confirmed in two independent cell systems with inducible expression of EWS/FLI1. We also analyzed the expression of DAX1 in Ewing tumors and derived cell lines, as well as in other nonrelated small round cell tumors. DAX1 was expressed in all Ewing tumor specimens analyzed, and in seven out of eight Ewing tumor cell lines, but not in any neuroblastoma or embryonal rhabdomyosarcoma. Furthermore, silencing of EWS/FLI1 by RNA interference in a Ewing tumor cell line markedly reduced the levels of DAX1 mRNA and protein, confirming that DAX1 up-regulation is dependent upon EWS/FLI1 expression. The high levels of DAX1 found in Ewing tumors and its potent transcriptional repressor activity suggest that the oncogenic effect of EWS/FLI1 may be mediated, at least in part, by the up-regulation of DAX1 expression.
尤因肿瘤家族存在染色体易位，该易位将EWS基因的N端区域与ETS家族的几种转录因子的C端区域连接起来，主要是FLI1，从而产生嵌合转录因子，这些因子在尤因肿瘤的发病机制中起关键作用。为了鉴定EWS/FLI1融合蛋白的下游靶点，我们建立了组成性表达嵌合EWS/FLI1或野生型FLI1蛋白的293细胞，并使用cDNA阵列来鉴定受EWS/FLI1差异调节的基因。DAX1（NR0B1）是一种参与性腺发育、性别决定和类固醇生成的不寻常孤儿核受体，显示出被EWS/FLI1癌蛋白持续上调，但不被野生型FLI1上调。在两个具有诱导性表达EWS/FLI1的独立细胞系统中证实了EWS/FLI1对DAX1的特异性诱导。我们还分析了DAX1在尤因肿瘤及其衍生细胞系以及其他不相关的小圆细胞肿瘤中的表达。DAX1在所分析的所有尤因肿瘤标本中以及在八个尤因肿瘤细胞系中的七个中表达，但在任何神经母细胞瘤或胚胎性横纹肌肉瘤中均不表达。此外，在一个尤因肿瘤细胞系中通过RNA干扰使EWS/FLI1沉默显著降低了DAX1 mRNA和蛋白水平，证实DAX1的上调依赖于EWS/FLI1的表达。在尤因肿瘤中发现的高水平DAX1及其强大的转录抑制活性表明，EWS/FLI1的致癌作用可能至少部分是由DAX1表达的上调介导的。
