Ionizing Radiation and Teniposide Increase p21waf1/cip1 and Promote Rb Dephosphorylation but Fail to Suppress E2F Activity in MCF-7 Breast Tumor Cells

  1. Michael S. Orr1,
  2. Nicole C. Watson,
  3. Sujatha Sundaram,
  4. Joyce K. Randolph,
  5. Pramod T. Jain and
  6. David A. Gewirtz
  1. Department of Pharmacology/Toxicology and Medicine, Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia 23298

    Abstract

    Ionizing radiation and the topoisomerase II inhibitor, teniposide (VM-26) both increase levels of the cyclin dependent kinase inhibitor, p21waf1/cip1 and promote dephosphorylation of the retinoblastoma tumor suppressor protein, Rb, in MCF-7 breast tumor cells, perturbations associated with suppression of the activity of the transcription factor, E2F. However, studies using an E2F binding site-luciferase reporter plasmid transfected into MCF-7 cells failed to demonstrate a reduction in E2F activity in response to VM-26 or to ionizing radiation. In contrast, E2F activity (both basal and E1A stimulated) could be suppressed by transfection with a plasmid expressing Rb, indicating that the capacity of E2F to bind to Rb and to be inactivated by Rb is functionally intact in MCF-7 cells. These findings in MCF-7 breast tumor cells suggest that E2F activity may not be directly susceptible to modulation by endogenous p21waf1/cip1 and Rb.

    Footnotes

    • Send reprint requests to: Dr. David A. Gewirtz, Department of Medicine, Medical College of Virginia, P.O. Box 980230, Richmond, VA 23298. E-mail: gewirtz{at}gems.vcu.edu

    • ↵1 Current affiliation: National Cancer Institute, National Institutes of Health, Bethesda, MD 20892.

    • ↵2 In separate studies, in which cell number was assessed by trypan blue exclusion over a period of 3 days, we independently verified that both VM-26 and ionizing radiation interfere with the growth of MCF-7 breast tumor cells.

    • ↵3 In a separate study using four sample replicates, we found no decrease in E2F activity by VM-26 even after 24 hr.

    • This research was supported in part by Grant NRC0493080 from the Nuclear Regulatory Commission, Grant CA55815 from the National Cancer Institute, and United States Army Medical Research and Material Command Award DAMD 17-96-1-6167.

    • Abbreviations:
      PBS
      phosphate-buffered saline
      TBS
      Tris-buffered saline
      • Received March 14, 1997.
      • Accepted May 19, 1997.
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    1. Molecular Pharmacology September 1, 1997 vol. 52 no. 3 373-379
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