Investigation of the Involvement of Parkin in Parkinson's Disease and Cancer by Monitoring the Changes in SH-SY5Y Cells at the Nuclear Proteome Level.

Abstract

During the last two decades, Parkinson's disease (PD)-associated genes have been associated with cancer; however, a shared pathogenic mechanism has yet to be discovered. Parkin, an E3 ubiquitin ligase that is involved in early-onset Parkinson's disease, has also been reported to exert tumor suppressor activity. However, the details about the role of Parkin in cancer remain unknown. The present study aimed at identifying differentially regulated nuclear proteins and nuclear phosphoproteins whose levels were affected by Parkin expression. SHS-SY5Y cells expressing either wild-type Parkin or its mutant under tetracycline control were used in this study; cells not expressing Parkin served as control. Nuclear proteins were enriched from Parkin-expressing and control cells to perform a comparative proteomics study using two-dimensional gel electrophoresis (2D) coupled to matrix assisted laser desorption/ionisation-time of flight (MALDI-TOF/TOF) mass spectrometry analysis. Changes in phosphoproteome and nuclear phosphoproteome were also studied by staining the 2D gels with ProQ diamond phosphoprotein stain. The identified proteins were subjected to bioinformatics analysis to elucidate the reactomes and relevant pathways. Six nuclear proteins, namely NCL, DDIT3, PARP1, HMGB1, TCTP and TPI were shown to be differentially regulated in cells expressing Parkin protein. Regulations in phosphorylation levels of ENPL, PRDX4, ECHM, ALDOA SET, DHSA, RCC1 and DULRD were also detected. Bioinformatics analysis of differentially regulated proteins highlighted the involvement of Parkin in DNA repair. Several nuclear protein candidates whose expression or phosphorylation levels were altered in cells expressing Parkin. Bioinformatics analysis of these proteins indicated that the nuclear form of Parkin may play a significant role in DNA repair and contribute to prevention of tumorogenesis via maintaining DNA integrity.