Restoration of transforming growth factor-beta signaling enhances radiosensitivity by altering the Bcl-2/Bax ratio in the p53 mutant pancreatic cancer cell line MIA PaCa-2

J Biol Chem. 2002 Jan 18;277(3):2234-46. doi: 10.1074/jbc.M110168200. Epub 2001 Nov 1.

Abstract

In this study, we investigated whether lack of transforming growth factor beta (TGF-beta) type II receptor (RII) expression and loss of TGF-beta signaling played a role in radiation resistance of pancreatic cancer cells MIA PaCa-2 that possess a mutated p53 gene. Transfection of this cell line with a RII cDNA led to a stimulation of the transcriptional activity of p3TP-Lux, a TGF-beta-responsive reporter construct. The RII transfectants (MIA PaCa-2/RII) showed a significant increase in sensitivity to radiation when compared with MIA PaCa-2/vector cells. The increase in sensitivity to radiation was reversed by neutralizing antibodies to TGF-beta, indicating that these changes were dependent on TGF-beta signaling. Compared with MIA PaCa-2/vector cells, MIA PaCa-2/RII cells showed a greater than 3-fold increase in apoptosis after radiation. Enhanced radiation sensitivity of MIA PaCa-2/RII cells was associated with an induction of Bax mRNA and protein that was followed by a release of cytochrome c and activation of caspase-3 and poly(ADP-ribose) polymerase cleavage after radiation exposure. Overexpression of Bcl-x(L) or treatment with antisense oligodeoxynucleotides targeted against Bax significantly inhibited radiation-induced apoptosis in MIA PaCa-2/RII but not in MIA PaCa-2/Vector cells, suggesting that Bax induction is necessary for radiation-induced TGF-beta signaling-mediated apoptosis. Thus, restoration of TGF-beta signaling sensitized these cells to ionizing radiation, although these cells possess a mutated p53 gene. In addition, disruption of RII function by dominant negative mutant of RII inhibited the radiation-induced TGF-beta signaling and apoptosis in primary cultures of mouse embryonic fibroblasts. Together, these observations imply that RII is an important component of radiation-induced TGF-beta signaling, and loss of function of RII may enhance resistance to radiation-induced apoptosis.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Apoptosis
  • Caspase 3
  • Caspases / metabolism
  • Cyclin-Dependent Kinase Inhibitor p21
  • Cyclins / genetics
  • Cytochrome c Group / metabolism
  • Humans
  • Mutation*
  • Pancreatic Neoplasms / metabolism*
  • Pancreatic Neoplasms / pathology
  • Proto-Oncogene Proteins / genetics
  • Proto-Oncogene Proteins / metabolism*
  • Proto-Oncogene Proteins c-bcl-2 / genetics
  • Proto-Oncogene Proteins c-bcl-2 / metabolism*
  • RNA, Messenger / genetics
  • Radiation Tolerance*
  • Receptors, Transforming Growth Factor beta / physiology
  • Signal Transduction*
  • Transfection
  • Transforming Growth Factor beta / metabolism*
  • Tumor Cells, Cultured
  • Tumor Suppressor Protein p53 / metabolism
  • bcl-2-Associated X Protein

Substances

  • BAX protein, human
  • CDKN1A protein, human
  • Cyclin-Dependent Kinase Inhibitor p21
  • Cyclins
  • Cytochrome c Group
  • Proto-Oncogene Proteins
  • Proto-Oncogene Proteins c-bcl-2
  • RNA, Messenger
  • Receptors, Transforming Growth Factor beta
  • Transforming Growth Factor beta
  • Tumor Suppressor Protein p53
  • bcl-2-Associated X Protein
  • CASP3 protein, human
  • Casp3 protein, mouse
  • Caspase 3
  • Caspases