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Correlation Between Structure of Bcl‐2 and Its Inhibitory Function of JNK and Caspase Activity in Dopaminergic Neuronal Apoptosis
Author(s) -
Choi WonSeok,
Yoon SoYoung,
Chang In Ick,
Choi EuiJu,
Rhim Hyewhon,
Jin Byung K.,
Oh Tae H.,
Krajewski Stanislaw,
Reed John C.,
Oh Young J.
Publication year - 2000
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.2000.0741621.x
Subject(s) - staurosporine , programmed cell death , microbiology and biotechnology , chemistry , caspase , apoptosis , cell culture , biology , kinase , protein kinase a , biochemistry , genetics
To examine the correlation between the structure of Bcl‐2and its inhibitory function of c‐Jun N‐terminal kinase (JNK) and caspaseactivity, we established a dopaminergic neuronal cell line, MN9Doverexpressing Bcl‐2 (MN9D/Bcl‐2) or its structural mutants. The mutantscomprised 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 (terminaldeoxynucleotidyltransferase nick end‐labeling) and MTT[3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide] reductionassay, apoptotic death of MN9D/Neo cells reached 80‐90% within 24 h inresponse to 1 μ M 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.