Dephosphorylation-induced decrease of anti-apoptotic function of Bcl-2 in neuronally differentiated P19 cells following ischemic insults.
It is known that Bcl-2 has a protective effect against neuronal ischemia. Some reports speculate anti-apoptotic function of Bcl-2 depends not on the expression level but on the phosphorylation state. We found induction of apoptosis and CPP32 activation by energy impairment (3-nitropropionic acid (3-NP)-treatment or glucose-deprivation) in the neuronally differentiated P19 cells. Time course study of cell viability following ischemic insults showed that the number of viable cells decreased along with the increase in the amount of dephosphorylated Bcl-2 without obvious quantitative alteration of the protein. Then, we generated differentiated P19 cells overexpressing wild-type Bcl-2 (P19/wt. Bcl-2) or phosphorylation-negative Bcl-2 mutant (P19/mut.Bcl-2), in which alanine was substituted for serine 70. When the cell viability was examined within 24 h, P19/mut.Bcl-2 was more vulnerable to energy impairment as compared with P19/wt.Bcl-2. In addition, overexpression of wild-type Bcl-2 inhibited DNA laddering and CPP32 activation induced by the insults, while that of mutant Bcl-2 did not. These findings suggest that the phosphorylation state, as well as the expression level, of Bcl-2 plays an important role to modulate its protective effect against ischemic insults.
Yokote
H
H
Department of neurosurgery, Wakayama Medical College, Kimiidera 811-1, Wakayama, Japan. hyokote@wakayama-med.ac.jp
Terada
T
T
Matsumoto
H
H
Kakishita
K
K
Kinoshita
Y
Y
Nakao
N
N
Nakai
K
K
Itakura
T
T
D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology*; D053148:Caspase 3; D020169:Caspases / Q000187:drug effects / Q000378:metabolism; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000187:drug effects* / Q000502:physiology*; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D003292:Convulsants / Q000494:pharmacology; D004734:Energy Metabolism / Q000187:drug effects; D020925:Hypoxia-Ischemia, Brain / Q000188:drug therapy* / Q000503:physiopathology*; D051379:Mice; D009154:Mutation / Q000187:drug effects / Q000235:genetics; D014411:Neoplastic Stem Cells / Q000187:drug effects* / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism*; D009574:Nitro Compounds; D010766:Phosphorylation; D011422:Propionates / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*
D003292:Convulsants; D009574:Nitro Compounds; D011422:Propionates; D019253:Proto-Oncogene Proteins c-bcl-2; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases; C015392:3-nitropropionic acid