Bax Deletion Further Orders the Cell Death Pathway in Cerebellar Granule Cells and Suggests a Caspase-independent Pathway to Cell Death

Timothy M Miller; Krista L Moulder; C. Michael Knudson; Douglas J Creedon; Mohanish Deshmukh; Stanley J Korsmeyer; Eugene M Johnson

doi:10.1083/jcb.139.1.205

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Bax Deletion Further Orders the Cell Death Pathway in Cerebellar Granule Cells and Suggests a Caspase-independent Pathway to Cell Death

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Bax Deletion Further Orders the Cell Death Pathway in Cerebellar Granule Cells and Suggests a Caspase-independent Pathway to Cell Death

Timothy M Miller, Krista L Moulder, C. Michael Knudson, Douglas J Creedon, Mohanish Deshmukh, Stanley J Korsmeyer and Eugene M Johnson

The Journal of cell biology, Vol.139(1), pp.205-217

10/06/1997

DOI: 10.1083/jcb.139.1.205

PMCID: PMC2139809

PMID: 9314540

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Abstract

Dissociated cerebellar granule cells maintained in medium containing 25 mM potassium undergo an apoptotic death when switched to medium with 5 mM potassium. Granule cells from mice in which Bax, a proapoptotic Bcl-2 family member, had been deleted, did not undergo apoptosis in 5 mM potassium, yet did undergo an excitotoxic cell death in response to stimulation with 30 or 100 μM NMDA. Within 2 h after switching to 5 mM K+, both wild-type and Bax-deficient granule cells decreased glucose uptake to <20% of control. Protein synthesis also decreased rapidly in both wild-type and Bax-deficient granule cells to 50% of control within 12 h after switching to 5 mM potassium. Both wild-type and Bax −/− neurons increased mRNA levels of c-jun, and caspase 3 (CPP32) and increased phosphorylation of the transactivation domain of c-Jun after K+ deprivation. Wild-type granule cells in 5 mM K+ increased cleavage of DEVD–aminomethylcoumarin (DEVD-AMC), a fluorogenic substrate for caspases 2, 3, and 7; in contrast, Bax-deficient granule cells did not cleave DEVD-AMC. These results place BAX downstream of metabolic changes, changes in mRNA levels, and increased phosphorylation of c-Jun, yet upstream of the activation of caspases and indicate that BAX is required for apoptotic, but not excitotoxic, cell death. In wild-type cells, Boc-Asp-FMK and ZVAD-FMK, general inhibitors of caspases, blocked cleavage of DEVD-AMC and blocked the increase in TdT-mediated dUTP nick end labeling (TUNEL) positivity. However, these inhibitors had only a marginal effect on preventing cell death, suggesting a caspase-independent death pathway downstream of BAX in cerebellar granule cells.

Details

Title: Subtitle: Bax Deletion Further Orders the Cell Death Pathway in Cerebellar Granule Cells and Suggests a Caspase-independent Pathway to Cell Death
Creators: Timothy M Miller - Department of Neurology and Department of Molecular Biology and Pharmacology, Department of Medicine and Department of Pathology, Howard Hughes Medical Institute, Washington University School of Medicine, St. Louis, Missouri 63110
Krista L Moulder - Department of Neurology and Department of Molecular Biology and Pharmacology, Department of Medicine and Department of Pathology, Howard Hughes Medical Institute, Washington University School of Medicine, St. Louis, Missouri 63110
C. Michael Knudson - Department of Neurology and Department of Molecular Biology and Pharmacology, Department of Medicine and Department of Pathology, Howard Hughes Medical Institute, Washington University School of Medicine, St. Louis, Missouri 63110
Douglas J Creedon - Department of Neurology and Department of Molecular Biology and Pharmacology, Department of Medicine and Department of Pathology, Howard Hughes Medical Institute, Washington University School of Medicine, St. Louis, Missouri 63110
Mohanish Deshmukh - Department of Neurology and Department of Molecular Biology and Pharmacology, Department of Medicine and Department of Pathology, Howard Hughes Medical Institute, Washington University School of Medicine, St. Louis, Missouri 63110
Stanley J Korsmeyer - Department of Neurology and Department of Molecular Biology and Pharmacology, Department of Medicine and Department of Pathology, Howard Hughes Medical Institute, Washington University School of Medicine, St. Louis, Missouri 63110
Eugene M Johnson - Department of Neurology and Department of Molecular Biology and Pharmacology, Department of Medicine and Department of Pathology, Howard Hughes Medical Institute, Washington University School of Medicine, St. Louis, Missouri 63110
Resource Type: Journal article
Publication Details: The Journal of cell biology, Vol.139(1), pp.205-217
DOI: 10.1083/jcb.139.1.205
PMID: 9314540
PMCID: PMC2139809
ISSN: 0021-9525
eISSN: 1540-8140
Language: English
Date published: 10/06/1997
Academic Unit: Pathology; Radiation Oncology
Record Identifier: 9984047794002771

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