Suppression of extensive neurofilament phosphorylation rescues α-Internexin/peripherin-overexpressing PC12 cells from neuronal cell death

• Published in: PloS one 2012-08, Vol.7 (8), p.e43883-e43883
• Main Authors: Lee, Wen-Ching, Kan, Daphne, Chen, Yun-Yu, Han, Shan-Kuo, Lu, Kuo-Shyan, Chien, Chung-Liang
• Format: Article
• Language: English
• Subjects: Aberration; Accumulation; Amyotrophic lateral sclerosis; Animals; Apoptosis; Biology; Cell death; Cell Death - drug effects; Cell Death - genetics; Cell Survival - drug effects; Cell Survival - genetics; Confocal microscopy; Cyclin-dependent kinase 5; Cyclin-Dependent Kinase 5 - metabolism; Cyclin-dependent kinases; Cytoplasm; Damage accumulation; Dementia; Disaggregation; Fluorescence; Gene Expression; Glycogen Synthase Kinase 3 - antagonists & inhibitors; Glycogen Synthase Kinase 3 - metabolism; Glycogen Synthase Kinase 3 beta; Growth factors; Indoles - pharmacology; Inhibitors; Intermediate Filament Proteins - genetics; Kinases; Maleimides - pharmacology; Medical treatment; Medicine; Membrane Glycoproteins - genetics; Microscopy; Mitochondria - drug effects; Mitochondria - metabolism; Mortality; Nerve Tissue Proteins - genetics; Nervous system; Neurodegeneration; Neurodegenerative diseases; Neurofilament Proteins - metabolism; Neurological diseases; Neurons; Neurons - cytology; Neurons - drug effects; Neurons - metabolism; Neuropathology; Organelles; Pathogenesis; PC12 Cells; Peripherin; Peripherins; Pharmacology; Pheochromocytoma cells; Phosphorylation; Phosphorylation - drug effects; Protein Transport - drug effects; Protein Transport - genetics; Proteins; Purines - pharmacology; Rats; Transgenic animals; Viability; Western blotting
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0043883

Intermediate filament (IF) overproduction induces abnormal accumulation of neuronal IF, which is a pathological indicator of some neurodegenerative disorders. In our study, α-Internexin- and peripherin-overexpressing PC12 cells (pINT-EGFP and pEGFP-peripherin) were used as models to study neuropathological pathways responsible for neurodegenerative diseases. Microarray data revealed that Cdk5-related genes were downregulated and Cdk5 regulatory subunit-associated protein 3 (GSK-3α and GSK-3β) were upregulated in pINT-EGFP cells. Increased expression of phosphorylated neurofilament and aberrant activation of Cdk5 and GSK-3β were detected in both pEGFP-peripherin and pINT-EGFP cells by Western blotting. In addition, pharmacological approaches to retaining viability of pINT-EGFP and pEGFP-peripherin cells were examined. Treatment with Cdk5 inhibitor and GSK-3β inhibitor significantly suppressed neuronal death. Dynamic changes of disaggregation of EGFP-peripherin and decrease in green fluorescence intensity were observed in pEGFP-peripherin and pINT-EGFP cells by confocal microscopy after GSK-3β inhibitor treatment. We conclude that inhibition of Cdk5 and GSK-3β suppresses neurofilament phosphorylation, slows down the accumulation of neuronal IF in the cytoplasm, and subsequently avoids damages to cell organelles. The results suggest that suppression of extensive neurofilament phosphorylation may be a potential strategy for ameliorating neuron death. The suppression of hyperphosphorylation of neuronal cytoskeletons with kinase inhibitors could be one of potential therapeutic treatments for neurodegenerative diseases.