Differential autophagic cell death under stress with ectopic cytoplasmic and mitochondrial-specific PPP2R2B in human neuroblastoma cells
| Autor: | Ding Chieh Song, Chang Heng Hsieh, Wan Ting Cheng, Dan Yu Li, Hui Fang Li, Zhi Xuan Guo, Kang Fang |
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| Rok vydání: | 2013 |
| Předmět: |
Cancer Research
Programmed cell death Clinical Biochemistry Pharmaceutical Science Nerve Tissue Proteins Biology Transfection chemistry.chemical_compound Cytosol Stress Physiological Cell Line Tumor Autophagy Humans Protein Phosphatase 2 Pharmacology Membrane Potential Mitochondrial Neurons Endoplasmic reticulum Adenine Tunicamycin Biochemistry (medical) Cell Biology Endoplasmic Reticulum Stress Molecular biology Cell biology Clone Cells Mitochondria Isoenzymes chemistry Gene Expression Regulation Cell culture Apoptosis Unfolded protein response Macrolides Reactive Oxygen Species Signal Transduction |
| Zdroj: | Apoptosis : an international journal on programmed cell death. 18(5) |
| ISSN: | 1573-675X |
| Popis: | Protein phosphatase 2A is one of four major classes of serine/threonine phosphatases. Overexpression of brain-specific regulatory subunit PPP2R2 in neuron cells is implicated in pathogenesis. The alternative splicing of PPP2R2B encodes two isoforms. They are subunit of cytoplasmic specific Bβ1 and mitochondria-targeted Bβ2. The two constructs were transfected into human neuroblastoma cells, SK-N-SH, respectively, and the stable clones overexpressing either Bβ1 or Bβ2 established. We have reported that Bβ2 clones are sensitive to reactive oxygen species (ROS) treatment by inducing autophagic cell death. To study more on the onset of neuropathogenesis under strain, both clones were exposed to different environmental stress, e.g. starvation and endoplasmic reticulum (ER) stress. To learn how PPP2R2B overexpression responds to starvation, cells were incubated in Hank’s buffered salt solution of deprived nutrient. Cell death was induced in Bβ1 clones after 6 h starvation, but not in Bβ2 clones. The pharmacological inhibitor, Bafilomycin A1, rescued the cell death while suppressing autophagy. On the other hand, to assess how cells respond to ER stress, the cells were treated with 0.1 μM of N-glycosylation inhibitor, tunicamycin (TM). In contrast with Bβ1, the apoptotic cell death appeared in Bβ2 after 48 h treatment. The formation of autophagolysosome was detected in Bβ2 following 12 h treatment with TM as evidenced by lysotracker and GFP-LC3 staining for fluorescence microscopy analysis. The autophagy inhibitor, 3-methyladenine, salvaged the final apoptosis. The stable cell lines with ectopically transfected PPP2R2B genes encoding isoforms of brain-specific regulatory subunit exhibit distinct apoptosis under different stressors. The induced autophagic apoptotic cell death is related to mitochondrial membrane potential drop and ROS generation. Disturbance of autophagy alleviates the induced cell death. The results promised a good model for understanding the onset in pathogenesis under stress in neuron cells with aberrant PPP2R2B expression. |
| Databáze: | OpenAIRE |
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