EWS/ATF1 expression induces sarcomas from neural crest–derived cells in mice
Kazunari Yamada, … , Akira Hara, Yasuhiro Yamada
Kazunari Yamada, … , Akira Hara, Yasuhiro Yamada
Published January 2, 2013
Citation Information: J Clin Invest. 2013;123(2):600-610. https://doi.org/10.1172/JCI63572.
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Research Article Oncology

EWS/ATF1 expression induces sarcomas from neural crest–derived cells in mice

Abstract

Clear cell sarcoma (CCS) is an aggressive soft tissue malignant tumor characterized by a unique t(12;22) translocation that leads to the expression of a chimeric EWS/ATF1 fusion gene. However, little is known about the mechanisms underlying the involvement of EWS/ATF1 in CCS development. In addition, the cellular origins of CCS have not been determined. Here, we generated EWS/ATF1-inducible mice and examined the effects of EWS/ATF1 expression in adult somatic cells. We found that forced expression of EWS/ATF1 resulted in the development of EWS/ATF1-dependent sarcomas in mice. The histology of EWS/ATF1-induced sarcomas resembled that of CCS, and EWS/ATF1-induced tumor cells expressed CCS markers, including S100, SOX10, and MITF. Lineage-tracing experiments indicated that neural crest–derived cells were subject to EWS/ATF1-driven transformation. EWS/ATF1 directly induced Fos in an ERK-independent manner. Treatment of human and EWS/ATF1-induced CCS tumor cells with FOS-targeted siRNA attenuated proliferation. These findings demonstrated that FOS mediates the growth of EWS/ATF1-associated sarcomas and suggest that FOS is a potential therapeutic target in human CCS.

Authors

Kazunari Yamada, Takatoshi Ohno, Hitomi Aoki, Katsunori Semi, Akira Watanabe, Hiroshi Moritake, Shunichi Shiozawa, Takahiro Kunisada, Yukiko Kobayashi, Junya Toguchida, Katsuji Shimizu, Akira Hara, Yasuhiro Yamada

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Figure 6

Fos plays a key role in EWS/ATF1-induced cell proliferation.

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Fos plays a key role in EWS/ATF1-induced cell proliferation. (A) Effe...
(A) Effect of Fos knockdown on proliferation of EWS/ATF1-induced cells. G1297 cells were treated with siRNA targeting Fos (si-Fos; 10 nM), a control siRNA (si-Control; 10 nM), or lipofectamine alone (Mock). 48 and 96 hours later, cell viability was determined by WST-8 assay. Results are mean ± SD (n = 4). ***P < 0.001 vs. si-Control and Mock. (B) EWS/ATF1-induced tumor cell lines overexpressing Fos or EGFP (G1297 Fos and G1297 GFP, respectively). pCAG-Fos-IZ vector or pCAG-EGFP-IZ vector were stably transfected in G1297 cells. Western blot analysis revealed that G1297 Fos cells stably expressed Fos protein even in the absence of doxycycline. (C) Cell proliferation assay for G1297, G1297 GFP, and G1297 Fos cells before and after doxycycline withdrawal. Doxycycline treatment (0.2 μg/ml) was withdrawn for 48 hours. Cell viability was determined by WST-8 assay. ***P < 0.001 vs. G1297 and G1297 GFP. (D and E) Effect of FOS knockdown on growth of human CCS cell lines. MP-CCS-SY and KAS cells were treated with siRNA#1 targeting FOS (si-FOS #1; 10 nM and 20 nM), control siRNA (si-Control; 20 nM), or lipofectamine alone (Mock). 48 and 96 hours later, cell viability was determined by WST-8 assay. Data are mean ± SD (n = 4). ***P < 0.001 vs. si-Control and Mock.