Proteomic Study of a Parkinson's Disease Model of Undifferentiated SH-SY5Y Cells Induced by a Proteasome Inhibitor

Abstract

Parkinson's disease (PD) is one of the most common nervous system degenerative diseases. However, the etiology of this disease remains elusive. Here, a proteasome inhibitor (PSI)-induced undifferentiated SH-SY5Y PD model was established to analyze protein alterations through proteomic study. Methods: Cultured undifferentiated SH-SY5Y cells were divided into a control group and a group treated with 2.5 µM PSI (PSI-treated group). An methyl thiazolyl tetrazolium (MTT) assay was applied to detect cell viability. Acridine orange/ethidium bromide (AO/EB), α-synuclein immunofluorescence and hematoxylin and eosin (H&E) staining were applied to evaluate apoptosis and cytoplasmic inclusions, respectively. The protein spots that were significantly changed were separated, analyzed by 2D gel electrophoresis and DIGE De Cyder software, and subsequently identified by MALDI-TOF mass spectrometry and database searching. Results: The results of the MTT assay showed that there was a time and dose dependent change in cell viability following incubation with PSI. After 24 h incubation, PSI resulted in early apoptosis, and cytoplasmic inclusions were found in the PSI-treated group through H&E staining and α-synuclein immunofluorescence. Thus, undifferentiated SH-SY5Y cells could be used as PD model following PSI-induced inhibition of proteasomal function. In total, 18 proteins were differentially expressed between the groups, 7 of which were up-regulated and 11 of which were down-regulated. Among them, 5 protein spots were identified as being involved in the ubiquitin proteasome pathway-induced PD process. Conclusions: Mitochondrial heat shock protein 75 (MTHSP75), phosphoglycerate dehydrogenase (PHGDH), laminin binding protein (LBP), tyrosine 3/tryptophan 5-monooxygenase activation protein (14-3-3ε) and YWHAZ protein (14-3-3ζ) are involved in mitochondrial dysfunction, serine synthesis, amyloid clearance, apoptosis process and neuroprotection. These findings may provide new clues to deepen our understanding of PD pathogenesis.

Cysteine Modification by Ebselen Reduces the Stability and Cellular Levels of 14-3-3 Proteins

“…These cells express multiple 14-3-3 isoforms, including 14-3-3 ε and ζ that were of particular interest in our study. It was recently shown that these 14-3-3 isoforms are subject to ubiquitin regulated proteasomal degradation in SHSY5Y cells (Jiang et al 2019). Our observation that ebselen treatment triggered ubiquitin-mediated proteasomal degradation of 14-3-3s in SHSY5Y cells (Fig.…”

“…It nevertheless possible, that the effect of ebselen on 14-3-3 levels may be indirect, by i.e. increasing the phosphorylation, ubiquitination, or other covalent modification of 14-3-3s, as such modifications are reported to trigger 14-3-3 degradation (Jiang et al 2019;Chen et al 2015).…”

Differential metabolism -associated gene expression of duck pancreatic cells in response to two subtype s of duck hepatitis A virus type 1

“…Finally, 3-phosphoserine was converted into L-serine by PSP [23]. L-serine is an important precursor involved in various processes, such as synthesis of proteins and phospholipids as well as the synthesis of tetrahydrofolate metabolites and specific amino acids, namely glycine, cysteine, D-serine [24][25]. Previous studies have shown that L-serine supplementation can attenuate alcoholic fatty liver formation in mice and rats [26].…”

Differential metabolism -associated gene expression of duck pancreatic cells in response to two subtype s of duck hepatitis A virus type 1

“…Finally, 3-phosphoserine is converted to Lserine by PSP [24]. L-serine is an important precursor involved in various processes, such as synthesis of proteins and phospholipids as well as the synthesis of tetrahydrofolate metabolites and speci c amino acids, namely glycine, cysteine and D-serine [25,26]. Previous studies have shown that L-serine supplementation can attenuate alcoholic fatty liver formation in mice and rats [27].…”