Transmembrane domain of Bcl-2 is required for inhibition of ceramide synthesis, but not cytochrome c release in the pathway of inostamycin-induced apoptosis.
Bcl-2 protein plays important roles in the regulation of apoptosis. However, the exact mechanism by which Bcl-2 blocks apoptosis is still unclear. In the present study, we found that overexpression of Bcl-2 in human small cell lung carcinoma Ms-1 cells inhibited not only the release of cytochrome c from mitochondria into cytosol but also de novo ceramide synthesis induced by inostamycin, a phosphatidylinositol turnover inhibitor. To investigate the correlation between the structure of Bcl-2 and its inhibitory function in inostamycin-induced apoptosis, Ms-1 cells that stably overexpress domain-deletional mutants of Bcl-2 were established. Transmembrane domain-deleted Bcl-2 failed to inhibit inostamycin-induced de novo ceramide synthesis, whereas it inhibited inostamycin-induced cytochrome c release, indicating that anchoring of Bcl-2 to membrane was a requirement for its inhibitory effect on inostamycin-induced ceramide synthesis, but not cytochrome c release. Thus, the deletion mutant of tarnsmembrane domain of Bcl-2 can suppress inostamycin-induced apoptosis by inhibiting cytochrome c release, a downstream event of ceramide synthesis in the pathway of inostamycin-induced apoptosis. We also found that the BH3 and BH4 domains of Bcl-2 were necessary for inhibition of inostamycin-induced apoptosis, and deletion of BH1 or BH2 did not affect the inhibitory effect of Bcl-2 to inostamycin-induced apoptotic events.
Kawatani
Makoto
M
Department of Biosciences and Informatics, Faculty of Science and Technology, Keio University, 3-14-1 Hiyohi, Kohoku-ku, Yokohama 223-8522, Japan.
Uchi
Madoka
M
Simizu
Siro
S
Osada
Hiroyuki
H
Imoto
Masaya
M
D017209:Apoptosis; D018288:Carcinoma, Small Cell; D002470:Cell Survival / Q000502:physiology; D002518:Ceramides / Q000096:biosynthesis*; D003574:Cytochrome c Group / Q000378:metabolism*; D005663:Furans / Q000494:pharmacology*; D006801:Humans; D008175:Lung Neoplasms; D009154:Mutation; D017434:Protein Structure, Tertiary; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D002518:Ceramides; D003574:Cytochrome c Group; D005663:Furans; D019253:Proto-Oncogene Proteins c-bcl-2; C071861:inostamycin