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Correlation between structure of Bcl-2 and its inhibitory function of JNK and caspase activity in dopaminergic neuronal apoptosis

 Won-Seok Choi  ;  So-Young Yoon  ;  In Ick Chang  ;  Eui-Ju Choi  ;  Hyewhon Rhim  ;  Byung K. Jin  ;  Tae H. Oh  ;  Stanislaw Krajewski  ;  John C. Reed  ;  Young J. Oh 
 Journal of Neurochemistry, Vol.74(4) : 1621-1626, 2000 
Journal Title
Issue Date
Apoptosis/drug effects ; Apoptosis/physiology* ; Caspases/metabolism* ; Cells, Cultured ; Dopamine/physiology ; Enzyme Inhibitors/pharmacology ; Humans ; In Situ Nick-End Labeling ; JNK Mitogen-Activated Protein Kinases ; Mitogen-Activated Protein Kinases/metabolism* ; Mutation/physiology ; Neurons/cytology* ; Neurons/enzymology* ; Parkinson Disease/metabolism ; Protein Structure, Tertiary ; Proto-Oncogene Proteins c-bcl-2/chemistry ; Proto-Oncogene Proteins c-bcl-2/genetics ; Proto-Oncogene Proteins c-bcl-2/metabolism* ; Staurosporine/pharmacology ; Structure-Activity Relationship ; Transfection
Apoptosis ; Bcl‐2 homology domain ; MN9D ; Parkinson's disease
To examine the correlation between the structure of Bcl-2 and its inhibitory function of c-Jun N-terminal kinase (JNK) and caspase activity, we established a dopaminergic neuronal cell line, MN9D overexpressing Bcl-2 (MN9D/Bcl-2) or its structural mutants. The mutants comprised a point mutation in the BH1 (G145A; MN9D/BH1) or BH2 (W188A; MN9D/BH2) domain and a deletion mutation in the C-terminal (MN9D/C22), BH3 (MN9D/BH3), or BH4 (MN9D/BH4) domain. As determined by the TUNEL (terminal deoxynucleotidyltransferase nick end-labeling) and MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] reduction assay, apoptotic death of MN9D/Neo cells reached 80-90% within 24 h in response to 1 microM staurosporine. Upon staurosporine treatment, JNK activity increased six- to sevenfold over the basal level within 2-4 h. Treatment of MN9D/Neo with both staurosporine and a caspase inhibitor, Z-VAD, attenuated cell death without suppressing JNK activation. Both staurosporine-induced cell death and JNK activation were attenuated in MN9D/Bcl-2. As determined by cleavage of poly(ADP-ribose) polymerase into 85 kDa, Bcl-2 blocked caspase activity as well. When cells overexpressing one of the Bcl-2 mutants were treated with staurosporine, death was attenuated in MN9D/BH1, MN9D/BH2, and MN9D/C22 but not in MN9D/BH3 and MN9D/BH4. Similarly, both JNK and caspase activation were blocked in MN9D/BH1, MN9D/BH2, and MN9D/C22, whereas they were not suppressed in MN9D/BH3 and MN9D/BH4. Taken together, our data indicate that there exists a close structural and functional correlation of Bcl-2 to JNK and caspase activity in staurosporine-induced dopaminergic neuronal cell death.
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2. College of Dentistry (치과대학) > Dept. of Oral Biology (구강생물학교실) > 1. Journal Papers
Yonsei Authors
Chang, In Ik(장인익)
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