Apaf-1 deficiency confers resistance to ultraviolet-induced apoptosis in mouse embryonic fibroblasts by disrupting reactive oxygen species amplification production and mitochondrial pathway.
Apoptosis requires tightly regulated cell death pathways. The signaling pathways that trigger a cell to undergo apoptosis after UV radiation are cell type specific and are currently being defined. Here, we have used pharmacological and genetic tools to demonstrate the decisive part of the mitochondrial pathway in UVC-induced apoptosis in mouse embryo fibroblasts (MEFs). UVC-induced apoptosis proceeded independent of the activation of death receptor components. In contrast, soon after UV radiation, MAPK activation and generation of reactive oxygen species (ROS) increased, followed by a decline in mitochondrial membrane potential (MMP) and cytochrome c release, as well as activation of caspase-9 and -3 and the upregulation of p47-phox. Deficiency of apaf-1, a critical member of the apoptosome, dramatically abolished all the UV-induced signal deterioration and cell death. In parallel, UVC-induced apoptosis was largely attenuated by either DN-caspase-9 or Bcl-X(L) overexpression. Pretreatment of cells with N-acetylcysteine or catalase but not Tempol decreased UVC-induced MAPK activation and apoptosis. Inhibition of JNK and caspase attenuated p47-phox upregulation. Altogether, we have for the first time demonstrated the critical role of Apaf-1 in the regulation of MAPK, ROS, and MMP in UVC-radiated MEFs and propose that the amplification feedback loop among mitochondrial signal molecules culminates in the demise of the cell.
Department of Pathology, Cancer Institute, University of Pittsburgh, Pittsburgh, PA 15261, USA. ref19@pitt.edu
Rentian
R
Feng
Jie
J
Han
Judith
J
Ziegler
Minying
M
Yang
Vincent
V
Castranova
D000818:Animals; D017209:Apoptosis / Q000528:radiation effects*; D053477:Apoptotic Protease-Activating Factor 1 / Q000172:deficiency*; D053181:Caspase 8 / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism; D002455:Cell Division / Q000528:radiation effects; D053144:Cell Nucleus Shape / Q000528:radiation effects; D002478:Cells, Cultured; D004789:Enzyme Activation; D053200:Fas-Associated Death Domain Protein / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000502:physiology* / Q000528:radiation effects; D020935:MAP Kinase Signaling System; D053078:Membrane Potential, Mitochondrial; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D014466:Ultraviolet Rays
C506593:Apaf1 protein, mouse; D053477:Apoptotic Protease-Activating Factor 1; D018797:Cell Cycle Proteins; C505534:Fadd protein, mouse; D053200:Fas-Associated Death Domain Protein; D017382:Reactive Oxygen Species; C505526:Casp8 protein, mouse; D053181:Caspase 8