Lrrk2 Mutants Induced By Dopamine Neurons Proteomics : A Preliminary Study On Apoptosis

Background and purpose:PD(parkinson's disease) is considered as the second neurodegenerative disease, affecting estimated 0.3% at age of 50, and 4.3% by 85. This syndrome is characterized clinically with resting tremor, rigidity, bradykinesia and postural instability, while pathologically progressive loss of dopaminergic neurons in the substantia nigra pars compacta and the presence of intracellular Lewy bodies in surviving neurons of the brainstem. So far the etiology of PD remains unclear, genetic studies on familiar PD resulted in discovery of sevaral pathogenic genes, including Parkin, PINK1, DJ-1,α-Synuclein, UCHL1 and LRRK2. Among them, LRRK2 has recently captured a lot of attention for its association with indistinguishable symptoms from typical late-onset PD. Anatomy studies on PD autopsies with LRRK2 mutants displayed all the pathological phenotypes that have been observed in parkinsonism including:ubiquitin intranuclear inclusions (Marinesco bodies), a-synucleinopathy and tauopathy, except for nigral loss alone. Therefore, studies on LRRK2 will provide a great opportunity to further understand the etiology of PD.Methords:we transfected SH-SY5Y cells with either wildtype or G2019S-mutant LRRK2 plasmid to examine the toxicity of LRRK2 mutant. Using 2-D electrophoresis combind with MS-MS, we explored the proteomic variation resulted from the toxicity.Results:Using 2-D electrophoresis and MS-MS, we identified six proteins with different expression between wildtype-LRRK2 and G2019S-LRRK2 overexpressed cells including:GID alpha, GSTP1, EEF1G, protein disulfide isomerase, Lactate dehydrogenase B and YWHAZ protein. Further study revealed that total GSTP1 activity was diminished and overexpression of GSTP1 was found protective.Conclusion:These results demonstrate that either Wt or G2019S LRRK2 overexpression is toxic to cell, while the mutant LRRK2 displayed more severe toxicity. Also, the proteomics results demonstrate six differentially expressed proteins in G2019S LRRK2 overexpressed cells including: GID alpha, GSTP1, EEF1G, protein disulfide isomerase, Lactate dehydrogenase B and YWHAZ protein. Futhermore, we found that overexpression of GSTP1 protected against LRRK2-mutant induced apoptosis, indicating that oxidative stress may be implicated in G2019S-LRRK2 toxicity.