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PMID: 8522579 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Thiol agents and Bcl-2 identify an alphavirus-induced apoptotic pathway that requires activation of the transcription factor NF-kappa B.

The Journal of cell biology ·Vol. 131 ·No. 5 ·1995-12-00 ·Pages 1149-61

Lin KI, Lee SH, Narayanan R, Baraban JM, Hardwick JM, Ratan RR

Abstract

Oxidative stress has been proposed as a common mediator of apoptotic death. To investigate further the role of oxidants in this process we have studied the effects of antioxidants on Sindbis virus (SV)-induced apoptosis in two cell lines, AT-3 (a prostate carcinoma line) and N18 (a neuroblastoma line). The thiol antioxidant, N-acetylcysteine (NAC), at concentrations above 30 mM, completely abrogates SV-induced apoptosis in AT-3 and N18 cells. The effects of NAC cannot be attributed to inhibition of viral entry or viral replication, changes in extracellular osmolarity or to increases in cellular glutathione levels, nor can they be mimicked by chelators of trace metals, inhibitors of lipid peroxidation or peroxide scavengers. In contrast, other thiol agents including pyrrolidine dithiocarbamate (PDTC, 75 microM) are protective. Because NAC and PDTC are among the most effective inhibitors of the transcription factor NF-kappa B, we examined SV's ability to activate NF-kappa B before the onset of morphologic or biochemical evidence of apoptosis. Within hours of infection, SV induced a robust increase in nuclear NF-kappa B activity in AT-3 and N18 cells; this activation was suppressible by NAC and PDTC. Over-expression of bcl-2 in AT-3 cells, which has been shown to inhibit SV-induced apoptosis, also inhibits SV-induced NF-kappa B activation. To determine if NF-kappa B activation is necessary for SV-induced apoptosis in these cells, we used double stranded oligonucleotides with consensus NF-kappa B sequences as transcription factor decoys (TFDs) to inhibit NF-kappa B binding to native DNA sites. Wild-type, but not mutant, TFDs inhibit SV-induced apoptosis in AT-3 cells. In contrast, TFD inhibition of NF-kappa B nuclear activity in N18 cells did not prevent SV-induced apoptosis. Taken together, these observations define a cell type-specific, transcription factor signaling pathway necessary for SV-induced apoptosis. Understanding the precise mechanism by which Bcl-2 and thiol agents inhibit SV-induced nuclear NF-kappa B activity in AT-3 cells may provide insights into the pluripotent antiapoptotic actions of these agents.

MeSH Terms
Acetylcysteine/pharmacology Animals Apoptosis Base Sequence Cell Nucleus/metabolism Chelating Agents/pharmacology DNA Dithiothreitol/pharmacology Iron Lipid Peroxidation/drug effects Male Mercaptoethanol/pharmacology Mice Molecular Sequence Data NF-kappa B/antagonists & inhibitors,metabolism Neuroblastoma/pathology Prostatic Neoplasms/pathology Proto-Oncogene Proteins/metabolism Proto-Oncogene Proteins c-bcl-2 Pyrrolidines/pharmacology Rats Sindbis Virus/physiology Sulfhydryl Reagents/pharmacology Thiocarbamates/pharmacology Tumor Cells, Cultured
Chemicals
Chelating Agents NF-kappa B Proto-Oncogene Proteins Proto-Oncogene Proteins c-bcl-2 Pyrrolidines Sulfhydryl Reagents Thiocarbamates pyrrolidine dithiocarbamic acid Mercaptoethanol DNA Iron Dithiothreitol Acetylcysteine
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Lin K I
Department of Molecular Microbiology and Immunology, Johns Hopkins School of Hygiene and Public Health, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Lee S H
Narayanan R
Baraban J M
Hardwick J M
Ratan R R
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.131.5.1149
Published
1995-12-00
Pages
1149-61
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2120641
Subset
IM
Grants
NICHD NIH HHS · 5T32HD07428 · United States
NIDA NIH HHS · DA-00266 · United States
NIMH NIH HHS · MH-00926 · United States
External Links
PubMed source DOI: 10.1083/jcb.131.5.1149 PMC Full Text View journal details
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