p53 accumulation in apoptotic macrophages is an energy demanding process that precedes cytochrome c release in response to nitric oxide

Oncogene. 1999 Nov 11;18(47):6403-10. doi: 10.1038/sj.onc.1203058.

Abstract

Apoptosis in response to stress signals activates effector caspases known to be regulated by the release of cytochrome c (Cyt c) from mitochondria and the subsequent ATP-dependent activation of the death regulator apoptotic protease-activating factor 1 (Apaf-1). Experiments were carried out to determine whether the release of Cyt c is evoked by NO. in RAW 264.7 macrophages and to position signaling components relative to mitochondria. S-nitrosoglutathione and spermine-NO caused a fast p53 accumulation, followed by Bcl-xL downregulation, Cyt c release, and caspase activation. These alterations were absent in p53 antisense expressing macrophages (R delta p53asn-11). In Bcl-2 overexpressing cells (Rbcl2-14) Cyt c relocation and caspase activation were abrogated although p53 accumulation remained intact. The use of caspase inhibitors revealed Cyt c release and decreased Bcl-xL expression to be caspase independent. ATP-depleted cells showed a shift from apoptosis towards necrosis and no p53 accumulation or caspase activation upon NO. addition. Conclusively, NO.-mediated apoptosis in macrophages is entirely controlled by the mitochondrial pathway with the implication that Cyt c relocation demands p53 accumulation. Moreover, pulse-chase-experiments in combination with the ATP-depletion protocol identified p53 accumulation and stabilization as an energy requiring process. This allowed to dissect two ATP-dependent steps, one is in association with Apaf-1 formation, while the other resides in p53 accumulation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Animals
  • Apoptosis*
  • Cell Line
  • Cytochrome c Group / metabolism*
  • Macrophages / metabolism*
  • Mice
  • Mitochondria / enzymology
  • Nitric Oxide / physiology*
  • Proto-Oncogene Proteins c-bcl-2 / metabolism
  • Tumor Suppressor Protein p53 / metabolism*
  • bcl-X Protein

Substances

  • Bcl2l1 protein, mouse
  • Cytochrome c Group
  • Proto-Oncogene Proteins c-bcl-2
  • Tumor Suppressor Protein p53
  • bcl-X Protein
  • Nitric Oxide
  • Adenosine Triphosphate