Induction of apoptosis by tamoxifen-activation of a p53-estrogen receptor fusion protein expressed in E1A and T24 H-ras transformed p53-/- mouse embryo fibroblasts

Oncogene. 1996 Aug 15;13(4):739-48.

Abstract

A fusion gene consisting of wild-type p53 linked to a modified ligand binding domain of the murine estrogen receptor has been constructed and should be a useful tool for studying controlled activation of wild-type p53 function in a variety of experimental cell systems. The protein product of this gene, p53ERTM, is expressed in cells constitutively but is not functional unless associated with tamoxifen or 4-hydroxytamoxifen. p53ERTM was introduced into p53-deficient mouse embryo fibroblasts (MEFs) expressing the E1A and T24 H-ras oncogenes. Activation of p53 in these transformed cells by the addition of tamoxifen or 4-hydroxytamoxifen resulted in apoptosis. In addition to engaging the apoptotic machinery, the tamoxifen-activated fusion protein exhibited other functions characteristic of wild-type p53, such as induction of WAF1 and MDM2 gene expression and activation of the p53-dependent spindle checkpoint in cells treated with nocodazole. Activation of p53ERTM expressed in p53-positive MEFs coexpressing E1A and ras had, at most, only a small cytotoxic effect. When three cell lines of transformed p53+/+ fibroblasts not expressing p53ERTM were tested for sensitivity to the DNA-damaging drug doxorubicin, the p53+/+ clones displayed either comparable sensitivity, or at most an increase in drug sensitivity of less than fourfold, as compared to several p53-/- cell lines. Our data show that restoration of wild-type p53 activity is sufficient to trigger apoptosis in p53-/- MEFs transformed with E1A and T24 H-ras and suggest that rare propagable clones of p53-normal MEFs expressing the E1A and T24 H-ras oncogenes have suffered compensatory alterations that compromise the ability to undergo p53-dependent apoptosis.

MeSH terms

  • Adenovirus E1A Proteins / genetics*
  • Animals
  • Apoptosis / genetics*
  • Cell Line, Transformed
  • Embryo, Mammalian / cytology
  • Estrogen Antagonists / pharmacology
  • Fibroblasts
  • Gene Expression Regulation
  • Genes, ras*
  • Mice
  • Receptors, Estrogen / genetics
  • Recombinant Fusion Proteins / genetics*
  • Tamoxifen / analogs & derivatives*
  • Tamoxifen / pharmacology
  • Tumor Suppressor Protein p53 / genetics

Substances

  • Adenovirus E1A Proteins
  • Estrogen Antagonists
  • Receptors, Estrogen
  • Recombinant Fusion Proteins
  • Tumor Suppressor Protein p53
  • Tamoxifen
  • afimoxifene