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Identification of target genes for EWS/ATF-1 chimeric transcription factor

Oncogene volume 22, pages 41–49 (2003)Cite this article

Abstract

Chromatin immunoprecipitation is a useful technique to detect in vivo direct interaction between any transcription factor and its binding site on genomic DNA. We applied this skill to identify the direct target gene for EWS/ATF-1 by coupling with a GFP reporter assay. This novel approach isolated 62 of cloned DNA fragments responding upon EWS/ATF-1 expression and 16 of 62 clones included putative ATF-1 binding sites. Further analysis revealed that six of the cloned fragments included possible regulatory regions of ATM, GPP34, ARNT2, NKX6.1, NYD-SP28 and POSH. Most of these clones upregulated reporter activity by overexpression of EWS/ATF-1, suggesting that putative ATF-1 binding sites in these clones are functional elements for ATF-1 in vivo. Consistently, endogenous expression of these genes was upregulated by EWS/ATF-1. Interestingly, the clone containing the promoter region of POSH, which is known to be a strong inducer of apoptosis, repressed reporter activity by overexpression of EWS/ATF-1. Correspondingly, EWS/ATF-1 expression decreased endogenous POSH expression, suggesting that six isolated genes may be involved in direct regulation by EWS/ATF-1. Moreover, induction of POSH brought apoptotic cell death to KAS, the clear cell sarcoma (CCS) cell line, suggesting that repressed expression of POSH in CCS may be relevant to the normal signaling pathway in apoptosis.

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Acknowledgements

We thank Mizuko Osaka for technical assistance, Takeshi Kuwata for critical comments, and Alan Hall for POSH expression constructs. This study was supported in part by a Grant-in-Aid for Scientific Research (B) from the Japan Society for the promotion of Science.

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Authors and Affiliations

  1. Department of Carcinogenesis, The Cancer Institute, Japanese Foundation for Cancer Research, Tokyo, 170-8455, Japan

    Miki Jishage, Takashi Fujino, Yukari Yamazaki & Takuro Nakamura

  2. Department or Orthopedics, Faculty of Medicine, Tokyo Medical and Dental University, Tokyo, 113-0034, Japan

    Hiroshi Kuroda

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  1. Miki Jishage
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  2. Takashi Fujino
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  3. Yukari Yamazaki
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  4. Hiroshi Kuroda
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  5. Takuro Nakamura
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Correspondence to Takuro Nakamura.

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Jishage, M., Fujino, T., Yamazaki, Y. et al. Identification of target genes for EWS/ATF-1 chimeric transcription factor. Oncogene 22, 41–49 (2003). https://doi.org/10.1038/sj.onc.1206074

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