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Evidence for the transforming activity of a truncated Int6 gene, in vitro

Oncogene volume 20, pages 5291–5301 (2001)Cite this article

Abstract

Int6/eIF3-p48 was first identified as a common integration site for MMTV in mouse mammary tumors. In all cases, the MMTV integration event resulted in an interruption of the normal Int6 transcript from one allele leaving the second allele intact and operative. We hypothesize that insertion of MMTV into Int6 results in a mutated allele that encodes a shortened Int6 mRNA and protein (Int6sh), which either modifies normal Int6 function or possesses a new independent function. To confirm the transforming potential of the mutation and its dominant function, we transfected two mammary epithelial cell lines, MCF10A (human), and HC11 (mouse), with Int6sh under the control of the elongation factor-1α (eEF1A) promoter. Expression of Int6sh in MCF10A and HC11 mammary epithelial cells leads to anchorage-independent growth in soft agar indicative of a transformed phenotype. Colonies selected from agar exhibited high levels of mutated Int6sh and wild type Int6 RNA transcripts by RT–PCR and Northern blot analysis. In addition, Int6sh transformed MCF10A and HC11 cells formed nodular growths, in vivo, in immune compromised hosts. NIH3T3 cells, mouse embryo fibroblasts, were also transformed to anchorage-independent growth in vitro by Int6sh expression. These observations provide direct evidence that the Int6 mutations observed in MMTV-induced tumors and hyperplasia contribute to the malignant transformation of the mammary epithelial cells.

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Acknowledgements

We are indebted to Dr Omar Coso for the generous gift of the pCEFLHA vector and Dr Chris Baumann for his tireless work in visualizing the Int6 wild type and Int6sh-GFP fusion proteins. Brenda Jones in Dr D Salomon's laboratory for providing the transformed cell lines for anchorage-independent growth studies. We would also like to thank Dr Mary Hilburger-Ryan and Dr David Salomon for critical reading of this manuscript.

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Authors and Affiliations

  1. National Institutes of Health, National Cancer Institute, Laboratory of Tumor Immunology and Biology, 10 Center Drive Room 8B07, Bethesda, MD 20892-1750, Maryland, USA

    Susan B Rasmussen, Robert Callahan & Gilbert H Smith

  2. Academia Nacional de Medicina, Buenos Aires, Argentina

    Edith Kordon

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  1. Susan B Rasmussen
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  2. Edith Kordon
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Correspondence to Gilbert H Smith.

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Rasmussen, S., Kordon, E., Callahan, R. et al. Evidence for the transforming activity of a truncated Int6 gene, in vitro. Oncogene 20, 5291–5301 (2001). https://doi.org/10.1038/sj.onc.1204624

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