Lack of correlation between p53 protein level and sensitivity of DNA-damaging agents in keratinocytes carrying adenovirus E1a mutants

Oncogene. 1995 Aug 17;11(4):675-82.

Abstract

p53 tumor suppressor protein is required for efficient execution of apoptosis after DNA-damage in many cell systems. Since the oncogene E1a confers susceptibility to DNA-damaging agents and stabilizes p53 protein, we investigate whether the sensitivity to anticancer drugs of E1a-expressing cells was mediated by binding to a specific set of cellular proteins (p60, p105, p107 and p300) and related to the induction of apoptosis and the level of p53 protein. We studied the effect of cisplatin (CDDP), doxorubicin (DOX) and ionizing radiation (RX) on murine keratinocytes (PAM 212) transfected by the wild type E1a oncogene or several E1a mutants which bind to different subsets of cellular proteins. Keratinocytes transfected with the mutant d1787N (which binds to p60, p105, p107 and p300) showed a lethality in response to CDDP (10 micrograms ml-1) fourfold higher than controls and threefold higher in response to DOX and radiation (5 grays). The sensitivity of keratinocytes carrying the mutant NTd1598 (binding to p105, p107 and p60) to DNA-damaging agents was similar to that of control keratinocytes, while mutant d1922/947 (binding only to p300) were resistant to CDDP and RX but sensitive to DOX. Apoptosis (after 24 h) studied by DNA fragmentation and flow cytometry was only observed in cells carrying the wild type E1a or the mutant d1787N. After treatment with DNA-damaging agents, p53 protein expression increased in all the cell lines and no rE1ation to sensitivity to anticancer agents or induction of apoptosis was observed. From these results, we conclude that cell sensitivity to cisplatin and ionizing radiation induced by the E1a oncogene requires binding to p105, p107 and p300 cellular proteins, while sensitivity to Doxorubicin requires binding only to p300. Interestingly, p53 protein levels were related to the binding to the p300 protein. The high levels of p53 after CDDP and DOX in the mutant d1922/947, which are only sensitive to DOX, suggest that E1a oncogene products may induce sensitivity to DNA-damaging agents by p53-related and unrelated pathways.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenovirus E1A Proteins / biosynthesis*
  • Adenovirus E1A Proteins / genetics*
  • Animals
  • Apoptosis / drug effects
  • Apoptosis / radiation effects
  • Blotting, Western
  • Cell Line, Transformed
  • Cisplatin / toxicity*
  • DNA Damage*
  • DNA Replication / drug effects
  • DNA Replication / radiation effects
  • Dose-Response Relationship, Drug
  • Doxorubicin / toxicity*
  • Gamma Rays
  • Keratinocytes / cytology
  • Keratinocytes / drug effects*
  • Keratinocytes / metabolism
  • Mice
  • Mutation
  • Oncogenes*
  • Transfection
  • Tumor Suppressor Protein p53 / analysis
  • Tumor Suppressor Protein p53 / metabolism*

Substances

  • Adenovirus E1A Proteins
  • Tumor Suppressor Protein p53
  • Doxorubicin
  • Cisplatin