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TGF-β-induced nuclear localization of Smad2 and Smad3 in Smad4 null cancer cell lines

Oncogene volume 22, pages 1317–1323 (2003)Cite this article

Abstract

Smad4 is a tumor suppressor gene that is commonly lost or mutated in colorectal and pancreatic cancers. The activated transforming growth factor-β (TGF-β) receptor phosphorylates Smad2 and Smad3, which then complex with Smad4 and translocate to the nucleus. Smad4 mutations when detected as present in some human cancers have been considered sufficient to inactivate TGF-β signaling. In this work, we describe a colon cancer cell line, VACO-9M, that is Smad4 null when analysed by multiple assays. To study the role of Smad4 in TGF-β-induced translocation of the receptor-activated Smads to the nucleus, we analysed by immunofluorescence the cellular localization of endogenous Smad2 and Smad3 after TGF-β treatment of VACO-9M, plus four additional Smad4 null cell lines of breast (MDA-MB-468), or pancreatic (BxPC3, Hs766T, CFPAC-1) origin. In each cell line, TGF-β treatment resulted in both Smad2 and Smad3 moving to the nucleus in a Smad4-independent fashion. Nuclear translocation of Smad2 and Smad3 was, however, not sufficient to activate reporters for TGF-β-induced transcriptional responses, which were however restored by transient transfection of wild-type Smad4. We conclude that Smad4 is not required for nuclear translocation of Smad2 and Smad3, but is needed activation of at least certain transcriptional responses.

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Acknowledgements

We thank Dr Dawn Dawson for expert assistance in acquiring the immunofluorescence pictures for this study. This work was supported by PHS Grants RO1 CA67409, RO1 CA72160, and P30 CA43703, and by the National Colon Cancer Research Alliance. SM is an associate investigator of the Howard Hughes Medical Institute.

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Authors and Affiliations

  1. Howard Hughes Medical Institute, Cleveland, 44106, OH, USA

    Stephen P Fink & Sanford Markowitz

  2. Department of Medicine, Cleveland, 44106, OH, USA

    Debra Mikkola, James K V Willson & Sanford Markowitz

  3. Ireland Cancer Center, Case Western Reserve University and Research Institute of University Hospitals of Cleveland, Cleveland, 44106, OH, USA

    James K V Willson & Sanford Markowitz

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  1. Stephen P Fink
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  2. Debra Mikkola
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Correspondence to Sanford Markowitz.

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Fink, S., Mikkola, D., Willson, J. et al. TGF-β-induced nuclear localization of Smad2 and Smad3 in Smad4 null cancer cell lines. Oncogene 22, 1317–1323 (2003). https://doi.org/10.1038/sj.onc.1206128

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