Abstract
Oxidative stress induces apoptosis in many cellular systems including glioblastoma cells, with caspase-8 activation was regarded as a major contribution to H2O2-induced cell death. This study focused on the role of the autophagic protein p62 in H2O2-induced apoptosis in U87MG cells. Oxidative stress was applied with H2O2, and cell apoptosis and viability were measured with use of caspase inhibitors or autophagic mediators or siRNA p62, GFP-p62 and GFP-p62-UBA (del) transfection. We found that H2O2 -induced U87MG cell death was correlated with caspase-8. To understand the role of p62 in MG132-induced cell death, the levels of p62/SQSTM1 or autophagy in U87MG cells were modulated with biochemical or genetic methods. The results showed that the over-expression of wild type p62/SQSTM1 significantly reduced H2O2 induced cell death, but knockdown of p62 aggravated the process. In addition, inhibition of autophagy promoted p62 and active caspase-8 increasing H2O2 -induced apoptosis while induction of autophagy manifested the opposite effect. We further demonstrated that the function of p62/SQSTM1 required its C-terminus UBA domain to attenuate H2O2 cytotoxity by inhibition of caspase-8 activity. Our results indicated that p62/SQSTM1 was a potential contributor to mediate caspase-8 activation by autophagy in oxidative stress process.
Highlights
Reactive oxygen species (ROS), which leads to oxidative stress (OS) at excess amount, positively contributes to carcinogenesis and to malignant progression of tumor cells (Gerald et al, 2004)
This study focused on the role of the autophagic protein p62 in H2O2-induced apoptosis in U87MG cells
Oxidative stress was applied with H2O2, and cell apoptosis and viability were measured with use of caspase inhibitors or autophagic mediators or siRNA p62, GFP-p62 and GFP-p62-UBA transfection
Summary
Reactive oxygen species (ROS), which leads to oxidative stress (OS) at excess amount, positively contributes to carcinogenesis and to malignant progression of tumor cells (Gerald et al, 2004). H2O2-induced apoptosis is directly through caspase-8 and is not through the mitochondria-dependent caspase-9 activation (Wu et al, 2011). It induced autophagy in H2O2treated cells, accompanied with a decreased level of p62 and the formation of autophagic vacuoles (Seo et al, 2011). P62 is a central regulator of tumorigenesis due to its abilities to modulate autophagy, to control the levels of ROS and misfolded proteins, and to ensure a timely transit of cell through mitosis; all crucial factors in cancer (Moscat and Diaz-Meco, 2012). We attempted to explored the role of caspase-8 in H2O2-induced associated apoptosis in U87MG cells, to evaluated the function of p62 in caspase-8 autophagic degradation response to oxidative insult by using the UBA domain deletion and knockdown of p62
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