Assessing Bad Sub-cellular Localization Under Conditions Associated with Prevention or Promotion of Mitochondrial Permeability Transition-Dependent Toxicity - Protein Misfolding and Cellular Stress in Disease and Aging - page 278

Chapter 20
Assessing Bad Sub-cellular Localization Under Conditions
Associated with Prevention or Promotion of Mitochondrial
Permeability Transition-Dependent Toxicity
Liana Cerioni and Orazio Cantoni
Abstract
Cells belonging to the monocyte/macrophage lineage are in general highly resistant to peroxynitrite, a
reactive nitrogen species extensively produced by these and other cell types under inflammatory condi-
tions. Resistance is not dependent on the scavenging of peroxynitrite but is rather associated with the
prompt activation of a survival signaling in response to various molecules largely available at the inflam-
matory sites, as arachidonic acid and products of the 5-lipoxygenase or cyclooxygenase pathways. We
detected significant levels of Bad in the mitochondria of monocytes/macrophages and found that these
signaling pathways converge in Bad phosphorylation, and thus in its cytosolic accumulation.
Phosphorylation inhibits binding of Bad to Bcl-2, or BclX L , and promotes its translocation to the cyto-
sol, thereby enabling Bcl-2 and BclX L to exert effects leading to prevention of mitochondrial permeability
transition (MPT). Upstream inhibition of the survival signaling indeed promotes the mitochondrial
accumulation of Bad and the rapid onset of MPT-dependent toxicity. The above results contribute to
the definition of the mechanism(s) whereby monocytes/macrophages survive to peroxynitrite in
inflamed tissues.
Key words: Bad, Peroxynitrite, Mitochondria, Mitochondrial permeability transition, Inflammation,
Sub-cellular localization, Western blot analysis
1. Introduction
The Bcl-2 family of proteins, a critical death regulator that resides
immediately upstream of mitochondria, comprises anti-apoptotic/
pro-survival members (Bcl-2, BclX L , Bcl-w, Mcl-1 and A1) and two
groups of pro-apoptotic/pro-death members, the “multidomain”