Changes in the two-dimensional electrophoresis pattern of the Parkinson's disease related protein DJ-1 in human SH-SY5Y neuroblastoma cells after dopamine treatment.

DJ-1 is a mitochondrial protein linked to Parkinson's disease. DJ-1 has been suggested to have several possible functions, although it has been mainly associated to oxidative stress defence. Changes in the two-dimensional electrophoresis pattern have been thoroughly described as a consequence of oxidative modification of the Cys106 residue. There is accumulating evidence supporting a specific role of DJ-1 in protecting dopaminergic neurons from dopamine itself. By exposing SH-SY5Y human neuroblastoma catecholaminergic cells to dopamine, we observed a specific increase in the most acidic forms in the DJ-1 two-dimensional electrophoresis pattern together with a significant decrease of the most basic spot. Unlike cells exposed to generic oxidative conditions, no additional shift was observed. The results are corroborated by a meta-analysis of the literature showing that in the absence of dopamine treatment the specific acidic form is underrepresented.

Proteomics as a tool to investigate cell models for dopamine toxicity.

Dopaminergic neurons are particularly exposed to oxidative stress because dopamine metabolism gives rise to endogenous toxins. Here, two different models for dopamine toxicity have been investigated using proteomics: stable alpha-synuclein-expressing neuroblastoma (SH-SY5Y) cells to understand how alpha-synuclein modulates dopamine toxicity at the central level, and cultured T-cell leukaemia (Jurkat) cells challenged with dopamine or l-dopa to evaluate protein changes on the surface of peripheral cells. alpha-synuclein-expressing cells were more resistant compared with wild-type cells, showing upregulation of antioxidant proteins including the Parkinson's disease-related DJ-1 protein. In addition, treatment of prototypical T-cells with l-dopa induced significant changes in proteins on the cell surface, indicating that peripheral blood lymphocytes may be good indicators of dopamine toxicity.

alpha-Synuclein protects SH-SY5Y cells from dopamine toxicity.

Dopaminergic human neuroblastoma SH-SY5Y cells were stably transformed to increase expression of alpha-synuclein, a Parkinson's disease-related protein. Transformed cells were more resistant to oxidative insults, showing a cytoprotective role of alpha-synuclein. The expression of redox chaperonins (DJ-1, HSP70, and 14-3-3) was evaluated by Western blotting. Expression of alpha-synuclein reduced HSP70 levels even in the presence of dopamine, with a twofold increase of DJ-1 in the absence of oxidants. DJ-1 is significantly reduced by dopamine, and even more by dopamine and Cu(II). Increased alpha-synuclein expression did not affect 14-3-3, although dopamine increased its level by 60% in wild-type cells. alpha-Synuclein not only upregulated DJ-1, but also shifted all DJ-1 forms to a single spot at pI=5.7 not observed in wild-type cells. Dopamine gradually restored the distribution of DJ-1 forms to a situation similar to wild-type cells, with the form at pI=6.1 progressively enriched under oxidative conditions.