Cooperative interception of neuronal apoptosis by BCL-2 and BAG-1 expression: prevention of caspase activation and reduced production of reactive oxygen species.
Neuronally differentiated PC12 cells undergo synchronous apoptosis when deprived of nerve growth factor (NGF). Here we show that NGF withdrawal induces actinomycin D- and cycloheximide-sensitive caspase (ICE-like) activity. The peptide inhibitor of caspase activity, N-acetyl-Asp-Glu-Val-Asp-aldehyde, was more potent than acetyl-Tyr-Val-Ala-Asp-chloromethyl ketone in preventing NGF withdrawal-induced apoptosis, suggesting an important role for caspase-3 (CPP32)-like proteases. We observed a peak of reactive oxygen species (ROS) 6 h after NGF withdrawal. ROS appear to be required for apoptosis, because cell death is prevented by the free radical spin trap, N-tert-butyl-alpha-phenylnitrone, and the antioxidant, N-acetylcysteine. ROS production was blocked by actinomycin D, cycloheximide, and caspase protease inhibitors, suggesting that ROS generation is downstream of new mRNA and protein synthesis and activation of caspases. Forced expression of either BCL-2 or the BCL-2-binding protein BAG-1 blocked NGF withdrawal-induced apoptosis, activation of caspases, and ROS generation, showing that they function upstream of caspases. Coexpression of BCL-2 and BAG-1 was more protective than expression of either protein alone.
Schulz
J B
JB
Department of Neurology, University of Tübingen, Germany.
Bremen
D
D
Reed
J C
JC
Lommatzsch
J
J
Takayama
S
S
Wüllner
U
U
Löschmann
P A
PA
Klockgether
T
T
Weller
M
M
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002352:Carrier Proteins / Q000096:biosynthesis*; D053148:Caspase 3; D020169:Caspases; D016923:Cell Death; D002454:Cell Differentiation; D002470:Cell Survival; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004268:DNA-Binding Proteins; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000378:metabolism; D005609:Free Radicals; D007700:Kinetics; D009414:Nerve Growth Factors / Q000494:pharmacology*; D009474:Neurons / Q000166:cytology* / Q000502:physiology*; D009842:Oligopeptides / Q000494:pharmacology; D016716:PC12 Cells; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis*; D051381:Rats; D017382:Reactive Oxygen Species / Q000502:physiology*; D012402:Rotenone / Q000494:pharmacology; D013113:Spin Labels; D013997:Time Factors; D014157:Transcription Factors
D000975:Antioxidants; C091732:BCL2-associated athanogene 1 protein; D002352:Carrier Proteins; D015853:Cysteine Proteinase Inhibitors; D004268:DNA-Binding Proteins; D004792:Enzyme Precursors; D005609:Free Radicals; D009414:Nerve Growth Factors; D009842:Oligopeptides; D019253:Proto-Oncogene Proteins c-bcl-2; D017382:Reactive Oxygen Species; D013113:Spin Labels; D014157:Transcription Factors; C108637:aspartyl-glutamyl-valyl-aspartal; D012402:Rotenone; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases