Role of autophagy and proteasome degradation pathways in apoptosis of PC12 cells overexpressing human alpha-synuclein.
Parkinson's disease is a common neurodegenerative disease in the elderly. Its causes and mechanisms are not clearly understood. To explore the specific role of autophagy and the ubiquitin-proteasome pathway in apoptosis, a specific proteasome inhibitor and macroautophagy inhibitor and stimulator were selected to investigate pheochromocytoma (PC12) cell lines transfected with human mutant (A30P) and wild-type (WT) alpha-synuclein. The apoptosis ratio was assessed by flow cytometry. LC3, heat shock protein 70 (hsp70) and caspase-3 expression in cell culture were determined by Western blot. The hallmarks of apoptosis and autophagy were assessed with transmission electron microscopy. Compared to the control group or the rapamycin (autophagy stimulator) group, the apoptosis ratio in A30P and WT cells was significantly higher after treatment with inhibitors of the proteasome and macroautophagy. The results of Western blots for caspase-3 expression were similar to those of flow cytometry; hsp70 protein was significantly higher in the proteasome inhibitor group than in control, but in the autophagy inhibitor and stimulator groups, hsp70 was similar to control. These findings show that inhibition of the proteasome and autophagy promotes apoptosis, and the macroautophagy stimulator rapamycin reduces the apoptosis ratio. And inhibiting or stimulating autophagy has less impact on hsp70 than the proteasome pathway.
Department of Neurology, Second Affiliated Hospital of Soochow University, 1055 Sanxiang Road, Suzhou 215004, China; Department of Neurology, Nanjing Jinling Hospital, Nanjing 210002, China.
Fang
F
Yang
Ya-Ping
YP
Yang
Cheng-Jie
CJ
Mao
Bi-Yin
BY
Cao
Zheng-Lin
ZL
Cai
Ji-Jun
JJ
Shi
Jin-Zhong
JZ
Huang
Ping
P
Zhang
Chun-Feng
CF
Liu
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D001343:Autophagy / Q000502:physiology*; D015153:Blotting, Western; D053148:Caspase 3 / Q000096:biosynthesis; D045744:Cell Line, Tumor; D005434:Flow Cytometry; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D016716:PC12 Cells; D010300:Parkinson Disease / Q000503:physiopathology*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D051381:Rats; D051844:alpha-Synuclein / Q000378:metabolism*
D018840:HSP70 Heat-Shock Proteins; D051844:alpha-Synuclein; D053148:Caspase 3; D046988:Proteasome Endopeptidase Complex