EWS/FLI1 up regulates mE2-C, a cyclin-selective ubiquitin conjugating enzyme involved in cyclin B destruction

Arvand, Afsane; Bastians, Holger; Welford, Scott M.; Thompson, Andrew; Ruderman, Joan V.; Denny, Christopher T.

doi:10.1038/sj.onc.1202129

Cited by 86 publications

(63 citation statements)

References 17 publications

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“…Based upon its action as a transcription factor, it is reasonable to think that EWS-FLI1 may deregulate expression or alter selection of FLI1 target genes. Several putative EWS-FLI1 regulated genes have been identified mostly in heterologous systems, only few of which might have transforming potential by themselves (Kovar et al, 1996;Thompson et al, 1996;May et al, 1997;Arvand et al, 1998;Hahm et al, 1999;Dauphinot et al, 2001;Matsumoto et al, 2001;Zwerner and May, 2001;Lessnick et al, 2002;Fukuma et al, 2003). Results obtained with an EWS-FLI1 mutant lacking a functional ets DNA-binding domain suggest that EWS-FLI1 may have additional, transcriptionindependent functions in the cellular transformation process (Jaishankar et al, 1999;Welford et al, 2001).…”

Section: Introductionmentioning

confidence: 93%

Homotypic and heterotypic interactions of EWS, FLI1 and their oncogenic fusion protein

et al. 2003

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In Ewing's sarcoma family tumors, the ets transcription factor gene FLI1 is rearranged with one EWS allele resulting in coexpression of germline EWS and chimeric EWS-FLI1 proteins. Here, we investigated the potential of germline EWS, FLI1 and EWS-FLI1 to oligomerize. In two functional in vivo tests, fluorescence resonance energy transfer (FRET) and the mammalian two-hybrid (MTH) assay, self-association of EWS and EWS-FLI1, but not of FLI1 was detected. In addition, interaction of EWS-FLI1 with EWS and FLI1 was observed. GST pulldown assays and immunoprecipitation experiments largely confirmed these results. The EWS N-terminal domain present in both EWS and EWS-FLI1 was found to contribute to homotypic and heterotypic interactions of these proteins. However, in the context of germline EWS, the presence of the whole or part of the C-terminal RNAbinding domain greatly supported the self-association potential of the protein. Involvement of an RNA component in EWS oligomerization was confirmed by sensitivity of the corresponding GST pull-down assay to RNaseA treatment. In contrast, EWS-FLI1 was able to self-associate and also bind to FLI1 via its C-terminal domain, which comprises the FLI1 DNA-binding motif. Accordingly, the EWS-FLI1 interaction was not disrupted by RNaseA treatment. Despite its potential to oligomerize, EWS-FLI1 bound to a tandem ets-binding site of the TGFb type II receptor promoter as a monomer. Therefore, the functional consequences of homo-and heterooligomerization of EWS and EWS-FLI1 proteins remain to be elucidated.

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Section: Introductionmentioning

confidence: 93%

Homotypic and heterotypic interactions of EWS, FLI1 and their oncogenic fusion protein

et al. 2003

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“…These observations have suggested that EWS-FLI-1 transforms cells through the aberrant activation of speci®c target genes. Indeed, genes upregulated by EWS-FLI-1 have been identi®ed through di erential expression screening between EWS-FLI-1-transformed and normal NIH3T3 cells May et al, 1997;Arvand et al, 1998). However, a direct role of EWS-FLI-1 on transcriptional activation of these genes has not been unambiguously demonstrated.…”

Section: Introductionmentioning

confidence: 99%

Analysis of the expression of cell cycle regulators in Ewing cell lines: EWS-FLI-1 modulates p57KIP2 and c-Myc expression

et al. 2001

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Ewing tumour is characterized by speci®c chromosome translocations which fuse EWS to a subset of genes encoding ETS transcription factors, most frequently FLI-1. We report the analysis of the expression of various cell cycle regulators both in Ewing tumour derived cell lines and in di erent cellular models with either inducible or constitutive EWS-FLI-1 cDNA expression. In Ewing cell lines, cyclin D1, CDK4, Rb, p27 KIP1 and c-Myc were consistently highly expressed whereas p57 KIP2 , p15 INK4B and p14 ARF demonstrated undetectable or low expression levels. The amount of p16 INK4A , p21 CIP1 , p18 INKAC and CDK6 was variable from one cell line to the other. The inducible expression of EWS-FLI-1 led to a strong upregulation of c-Myc and a considerable downregulation of p57 KIP2 . Other proteins did not show evident modi®cation. High c-Myc and very low p57 KIP2 expression levels were also observed in neuroblastoma NGP cells constitutively expressing EWS-FLI-1 as compared to parental cells. Analysis of the p57 KIP2 promoter indicated that EWS-FLI-1 downregulates, possibly through an indirect mechanism, the transcription of this gene. Finally, we show that ectopic expression of p57 KIP2 in Ewing cells blocks proliferation through a complete G1 arrest. These results suggest that the modulation of p57 KIP2 expression by EWS-FLI-1 is a fundamental step in Ewing tumorigenesis. Oncogene (2001) 20, 3258 ± 3265.

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“…Using a variety of techniques and systems, several EWS/FLI1 target genes have been identified. Among them, c-myc, 13,14 cyclin D1, 14 CD99, 11 Id2, 14,15 Tenascin-C, 16 mE2-C, 17 PDGF-C, 18 EAT-2, 19 Manic Fringe, 20 uridine phosphorylase, 21 COL11A2 22 or PIM3 23 have been shown to be up-regulated by EWS/FLI1 or EWS/ERG oncoproteins, while the TGF-b type II receptor, 24,25 p57 KIP2 , 13 p21 WAF1/CIP1 26 and IGFBP3 9 are down-regulated. However, the exact mechanism of tumorigenesis mediated by EWS/ETS remains unclear.…”

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confidence: 99%

The orphan nuclear receptor DAX1 is up-regulated by the EWS/FLI1 oncoprotein and is highly expressed in Ewing tumors

Mendiola¹,

Carrillo²,

García³

et al. 2005

Int. J. Cancer

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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. ' 2005 Wiley-Liss, Inc.Key words: Ewing tumors; EWS/FLI1 oncoprotein; orphan nuclear receptor DAX1; cDNA arrays Ewing sarcoma and peripheral primitive neuroectodermal tumors (PNETs) are referred to as the Ewing family of tumors, a group of highly malignant tumors arising in children, adolescents and young adults. 1 Ewing tumors are characterized by specific balanced chromosomal translocations, leading to the formation of chimeric proteins that join the N-terminal region from the EWS gene product in frame with the C-terminal portion of out of five different members of the ETS family of transcription factors: FLI1, ERG or FEV (ERG subfamily) and E1AF or ETV1 (PEA3 subfamily). The EWS/FLI1 combination is most frequent and is present in nearly 85% of all cases. 2 A large body of evidence has shown the oncogenic potential of EWS/FLI1 protein. Thus, the ectopic expression of EWS/FLI1 in murine fibroblasts promotes anchorage-independent growth in vitro 3 and accelerates tumorigenesis in vivo. 4 Moreover, EWS/FLI1 knock-down using a variety of techniques inhibits the growth of Ewing tumor-derived cell lines both in vitro and in vivo. 5-9 These data, together with the observation that EWS/ETS chimeric proteins are present in virtually all Ewing tumors, have provided strong evidence that these fusion genes are necessary for the development of these neoplasms. However, EWS/FLI1 appears to be toxic for mouse embryo fibroblasts 10 and p...

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EWS/FLI1 up regulates mE2-C, a cyclin-selective ubiquitin conjugating enzyme involved in cyclin B destruction

Cited by 86 publications

References 17 publications

Homotypic and heterotypic interactions of EWS, FLI1 and their oncogenic fusion protein

Homotypic and heterotypic interactions of EWS, FLI1 and their oncogenic fusion protein

Analysis of the expression of cell cycle regulators in Ewing cell lines: EWS-FLI-1 modulates p57KIP2 and c-Myc expression

The orphan nuclear receptor DAX1 is up-regulated by the EWS/FLI1 oncoprotein and is highly expressed in Ewing tumors

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