Divergent Ewing's sarcoma EWS/ETS fusions confer a common tumorigenic phenotype on NIH3T3 cells

Thompson, Andrew; Teitell, Michael A.; Arvand, Afsane; Denny, Christopher T.

doi:10.1038/sj.onc.1202928

Cited by 95 publications

(84 citation statements)

References 43 publications

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“…Although soft agar assays are often used as a measure for malignant transformation, the results do not necessarily correspond with the ability of the transfected cells to form tumors in vivo. In Ewing sarcomas, for example, several chimeric transcription factors such as EWS/FLI1 and EWS/ETV1 are thought to modulate aberrant gene expression leading to tumorigenesis (Thompson et al, 1999). Although both fusion products were able to confer a common tumorigenic phenotype onto NIH3T3 cells as measured by tumor formation in SCID mice, only one of them (EWS/FLI1) was able to form colonies in soft agar.…”

Section: Discussionmentioning

confidence: 99%

Transformation capacities of the papillary renal cell carcinoma-associated PRCCTFE3 and TFE3PRCC fusion genes

Weterman

Groningen²,

Hartog

et al. 2001

Oncogene

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A recurrent chromosomal abnormality associated with a subset of papillary renal cell carcinomas is t(X;1)(p11;q21). This translocation leads to the formation of two fusion genes, TFE3PRCC and the reciprocal product PRCCTFE3. Both fusion genes are expressed in t(X;1)-positive renal cell carcinomas and contain major parts of the coding regions of the parental transcription factor PRCC and TFE3 genes, respectively. To ®nd out whether these fusion genes possess transforming capacity, we transfected NIH3T3 and rat-1 cells with the fusion products, either separately or combined. When using soft agar assays, we observed colony formation in all cases. NIH3T3 cells transfected with PRCCTFE3 or PRCCTFE3 together with TFE3PRCC yielded the highest colony forming capacities. Examination of other characteristics associated with malignant transformation, i.e., growth under low-serum conditions and formation of tumors in athymic nude mice, revealed that cells transfected with PRCCTFE3 exhibited all these transformation-associated characteristics. Upon transfection of the fusion products into conditionally immortalized kidney cells, derived from the proximal tubules of an H2Kb-tsA58 transgenic mouse, and consecutive incubation under non-permissive conditions, growth arrest was observed, followed by di erentiation except for those cells transfected with PRCCTFE3. Therefore, we conclude that PRCCTFE3 may be the t(X;1)-associated fusion product that is most critical for the development of papillary renal cell carcinomas.

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Section: Discussionmentioning

confidence: 99%

Transformation capacities of the papillary renal cell carcinoma-associated PRCCTFE3 and TFE3PRCC fusion genes

Weterman

Groningen²,

Hartog

et al. 2001

Oncogene

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“…1,2 During the past decades, EWSR1 has been identified as a translocation partner for many transcription factors including FLI-1, ATF-1, NR4A3, and CHOP in diverse tumors. [3][4][5][6][7] Recently, an increasing number of studies have focused on the characterization of EWSR1 itself. EWSR1 is ubiquitously expressed in most human cell types, except for cardiac muscle cells and melanocytes.…”

Section: Introductionmentioning

confidence: 99%

EWSR1 regulates mitosis by dynamically influencing microtubule acetylation

et al. 2016

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EWSR1, participating in transcription and splicing, has been identified as a translocation partner for various transcription factors, resulting in translocation, which in turn plays crucial roles in tumorigenesis. Recent studies have investigated the role of EWSR1 in mitosis. However, the effect of EWSR1 on mitosis is poorly understood. Here, we observed that depletion of EWSR1 resulted in cell cycle arrest in the mitotic phase, mainly due to an increase in the time from nuclear envelope breakdown to metaphase, resulting in a high percentage of unaligned chromosomes and multipolar spindles. We also demonstrated that EWSR1 is a spindle-associated protein that interacts with a-tubulin during mitosis. EWSR1 depletion increased the cold-sensitivity of spindle microtubules, and decreased the rate of spindle assembly. EWSR1 regulated the level of microtubule acetylation in the mitotic spindle; microtubule acetylation was rescued in EWSR1-depleted mitotic cells following suppression of HDAC6 activity by its specific inhibitor or siRNA treatment. In summary, these results suggest that EWSR1 regulates the acetylation of microtubules in a cell cycledependent manner through its dynamic location on spindle MTs, and may be a novel regulator for mitosis progress independent of its translocation.

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“…Most mutations or deletions in the DBD result in almost complete loss of transforming potential (9,10). More recently, however, alternative in vivo-based tumor assays have been used to assay the oncogenic activity of EWS/ETS fusions proteins (11)(12)(13). In some of these experiments, it has become apparent that although often in agreement, these two different transformation assays do not always concur (13,14).…”

mentioning

confidence: 99%

“…More recently, however, alternative in vivo-based tumor assays have been used to assay the oncogenic activity of EWS/ETS fusions proteins (11)(12)(13). In some of these experiments, it has become apparent that although often in agreement, these two different transformation assays do not always concur (13,14). Because the ets DBD is one of two necessary FLI1 domains to promote anchorageindependent growth (10, 11), we investigated whether EWS/ FLI1 is dependent on this domain for the acceleration of tumor formation in SCID mice as well.…”

mentioning

confidence: 99%

DNA Binding Domain-independent Pathways Are Involved in EWS/FLI1-mediated Oncogenesis

Welford¹,

Hebert²,

Deneen³

et al. 2001

Journal of Biological Chemistry

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Specific chromosomal translocations involving the ews gene and one of five members of the ets family of transcription factors create ews/ets fusion genes that are found in ϳ85% of Ewing's family of tumors. ews/ets fusion genes consistently maintain an intact and functional ets DNA binding domain (DBD) in all of these cases. We demonstrate here, however, that EWS/FLI1, the most prevalent EWS/ETS fusion, activates oncogenic pathways independent of its DBD. In in vivo tumor assays, EWS/FLI1 molecules with either point mutations or a large deletion in the ets DBD retain the ability to accelerate tumors in NIH 3T3 cells, whereas they lose the ability to bind DNA in vitro. Additionally, whereas inhibition of DBD functions of EWS/FLI1 with a dominant negative form of FLI1 is sufficient to inhibit anchorage-independent growth in NIH 3T3 cells, it is ineffective in inhibiting tumor growth in SCID mice. Usage of this dominant negative construct in a Ewing's tumor cell line, however, does reduce the rate of tumor formation, supporting the need for a functional DBD in this context. Together, these results suggest that EWS/FLI1 induces both DBD-dependent and DBD-independent oncogenic pathways.The production of chimeric fusion genes through chromosomal translocation is a common theme in the development of neoplasia. The resultant gene products often utilize domainassociated functions derived from the native participants of the fusions but in novel, atypical contexts. Such is the case for the Ewing's family of tumors, in which the ews gene on chromosome 22 is fused to one of five members of the ets family of transcription factors (FLI1, ERG, ETV1, E1AF, or FEV) (1). The consequent ews/ets fusion genes are thought to utilize amino-terminal EWS transactivation domains and intact carboxyl-terminal ets DNA binding domains (DBDs) 1 to form aberrant transcription factors that induce neoplastic phenotypes through altered gene regulation (2).EWS is a member of the TET family of RNA-binding proteins, which also includes TLS and TAF II 68 (3). These genes are commonly involved in chromosomal translocations that lead to the development of cancer, particularly sarcomas. In addition to Ewing's sarcoma, TET translocations are associated with at least five other sarcomas including desmoplastic small round cell tumors, clear cell sarcoma, and myxoid liposarcoma (1). A pervasive feature of all of the TET translocations is that a TET family member is fused to an intact DBD of a transcription factor. This observation supports the hypothesis that TET translocation products, including EWS/ETS fusions, utilize their DBDs to bind DNA and directly alter transcriptional activity.Although the physiological functions of wild type EWS are not well defined, EWS may have a role in coupling transcription and RNA processing. The amino-terminal transcriptional activation domain of EWS includes protein-protein interaction domains that direct binding to RNA polymerase II subunits (4, 5). In addition, an interaction with splicing factor U1C has been mapped to ...

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Divergent Ewing's sarcoma EWS/ETS fusions confer a common tumorigenic phenotype on NIH3T3 cells

Cited by 95 publications

References 43 publications

Transformation capacities of the papillary renal cell carcinoma-associated PRCCTFE3 and TFE3PRCC fusion genes

Transformation capacities of the papillary renal cell carcinoma-associated PRCCTFE3 and TFE3PRCC fusion genes

EWSR1 regulates mitosis by dynamically influencing microtubule acetylation

DNA Binding Domain-independent Pathways Are Involved in EWS/FLI1-mediated Oncogenesis

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