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  • Original Paper
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Raf-1-induced growth arrest in human mammary epithelial cells is p16-independent and is overcome in immortal cells during conversion

Oncogene volume 21, pages 6328–6339 (2002)Cite this article

Abstract

Using an estrogen-inducible retroviral system, we demonstrate that oncogenic Raf-1 induces growth arrest and morphological changes in finite lifespan human mammary epithelial cells (HMEC). This arrest does not rely on expression of the cyclin-dependent kinase inhibitor (CKI) p16INK4a, nor on changes in expression of the CKIs p21Cip1, p14ARF, p27Kip1 or p57Kip2. The Raf-induced arrest is independent of viral oncogene mediated inactivation of p53 and pRB, or c-myc overexpression. Flow cytometric analysis demonstrates that cells arrest in both G1 and G2. The Raf-induced arrest is mitigated or eliminated in some immortally transformed HMEC. Immortal HMEC that have both overcome replicative senescence and undergone the recently described conversion process maintain growth in the presence of transduced oncogenic Raf-1; they also gain EGF-independent growth and a low frequency of anchorage-independent growth. However, HMEC that have overcome replicative senescence but have not undergone conversion and HMEC immortalized by transduction with the catalytic subunit of telomerase, hTERT, remain severely growth arrested. These results indicate that the molecular mechanisms responsible for the Raf-1-induced growth arrest may vary among different finite lifespan cell types, and that in HMEC, this mechanism is altered during the conversion process, rather than as a direct consequence of overcoming senescence or expressing hTERT.

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Acknowledgements

We thank Martin McMahon for providing the EGFPΔRaf-1 : ER and hbER retroviral vectors, as well as helpful discussions. We are grateful to Denise Galloway, Vimla Band, Judy Campisi and Steve Frisch for providing viral oncogene constructs, and to Bruno Amati for the c-myc construct. We also thank Ana Krtolica and Hector Nolla for assistance with flow cytometry. This work was supported by the California Breast Cancer Research Program grant #4JB-0119 (MR Stampfer, CL Olsen, P Yaswen), Association pour la Recherche Contre le Cancer (B Gardie), NIH grant CA-24822 (MR Stampfer, P Yaswen), and the Office of Energy Research, Office of Health and Environmental Research, US Department of Energy under Contract No. DE-AC03-76SF00098 (MR Stampfer, P Yaswen).

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  1. Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, 94720, California, CA, USA

    Catherine L Olsen, Betty Gardie, Paul Yaswen & Martha R Stampfer

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  1. Catherine L Olsen
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  2. Betty Gardie
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  3. Paul Yaswen
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Correspondence to Martha R Stampfer.

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Olsen, C., Gardie, B., Yaswen, P. et al. Raf-1-induced growth arrest in human mammary epithelial cells is p16-independent and is overcome in immortal cells during conversion. Oncogene 21, 6328–6339 (2002). https://doi.org/10.1038/sj.onc.1205780

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