Apaf-1 overexpression partially overcomes apoptotic resistance in a cisplatin-selected HeLa cell line.

P Kamarajan, NK Sun, CL Sun, Chuck C.-K. Chao

Research output: Contribution to journalJournal Article peer-review

Abstract

Inhibition of caspase-3-mediated apoptosis has been hypothesized to be associated with chemoresistance. Investigations of apoptosis revealed that cytosolic cytochrome c is associated with a complex of apoptotic protease activating factor-1 (Apaf-1), an adapter molecule, and caspase-9 to activate caspase-3. However, whether these apoptotic molecules are involved in acquired cisplatin resistance is not understood. The present work shows reduced activation of caspase-3 and apoptosis in a cisplatin-selected HeLa cell line. Ac-DEVD-CHO, a caspase-3 inhibitor, inhibited cisplatin-induced apoptosis about 60-70% in both cell lines. Ac-LEHD-CHO, a caspase-9 inhibitor or Ac-IETD-CHO, a caspase-8 inhibitor, inhibited cisplatin-induced caspase-3 activation and apoptosis similarly in both cell lines. In addition, cisplatin induced the activation of caspase-9, the upstream activator of caspase-3, in a dose-dependent manner, and the activation of caspase-9 was less induced in resistant cells. The accumulation of cytosolic cytochrome c, an activator of caspase-9, and the induction of the mitochondrial membrane-associated voltage-dependent anion channel were also reduced in cisplatin-resistant cells. However, the concentration of Bcl-2 family proteins in cisplatin-resistant cells was normal. The concentration of Apaf-1 was unaltered in both cell lines. Increasing the cellular concentration of Apaf-1 through the transient expression of the gene increased the induction of apoptosis in resistant cells, associated with enhanced activation of caspase-9, caspase-3 and DNA fragmentation factor. Regression analysis reveals that the modification factor, the ratio of the slope in the linear range of the dose-response curve with Apaf-1 to the slope without Apaf-1, is 1.5 and 4.75 in the HeLa and cisplatin-resistant HeLa cells, respectively. These results indicate that apoptosis and caspases are less induced in cisplatin-selected HeLa cells. They also suggest that ectopic overexpression of Apaf-1 may partially reverse the acquired cisplatin resistance.
Original languageAmerican English
Pages (from-to)206-212
JournalFEBS Letters
Volume505
Issue number2
DOIs
StatePublished - 2001

Keywords

  • Adenoviridae/metabolism
  • Antineoplastic Agents/pharmacology
  • Apoptosis
  • Apoptotic Protease-Activating Factor 1
  • Blotting, Western
  • Caspase 8
  • Caspase 9
  • Caspases/antagonists & inhibitors
  • Cisplatin/pharmacology
  • Cytochrome c Group/metabolism
  • Cytochrome c Group/pharmacology
  • Cytosol/metabolism
  • DNA Fragmentation
  • Dose-Response Relationship, Drug
  • Drug Resistance, Neoplasm
  • Enzyme Activation/drug effects
  • Enzyme Inhibitors/pharmacology
  • HeLa Cells
  • Humans
  • Mitochondria/metabolism
  • Oligopeptides/pharmacology
  • Proteins/metabolism
  • Proto-Oncogene Proteins c-bcl-2/metabolism
  • Regression Analysis
  • Tumor Cells, Cultured

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