Comprehensive proteome and phosproteome analysis of human LRRK2 Drosophila model of Parkinson's disease

摘要

Gene mutations in the leucine-rich repeat kinase 2 (LRRK2) are the most common cause of autosomal dominant Parkinson`s Disease (PD) and elevated levels of hLRRK2 mutant variants in Drosophila induces PD. Here, we introduced the human LRRK2 (R1441C) variant in dopaminergic neurons of flies and observed a reduced locomotor activity, an age dependent degeneration of dopaminergic neurons, and shorter lifetime. To better understand the hLRRK2 (R1441C) induced pathobiology, we performed stable isotope labeling in fly to accurately quantify the proteome and phosphoproteome dynamics. We quantified almost 3000 proteins and found several regulated cytoskeletal, mitochondrial, and synaptic vesicle (SV) proteins in our PD fly model. To explore the hLRRK2 (R1441C) function more precisely, we compared our model to three different alpha-Synuclein overexpressing fly strains (WT,A30P, A53T), which show a similar PD phenotype. For example, synaptotagmin, syntaxin and rab3 were only affected in hLRRK2 (R1441C) flies compared to all other tested fly strains. Moreover, our global phosphoproteome analysis revealed several synaptic vesicle proteins with enhanced phosphorylation, including synaptojanin (pT1131) and the microtubule-associated protein futsch (pS4106). Consistently, a protein-protein interaction screen confirmed that hLRRK2 is tightly associated with synaptic vesicle proteins. Thus, our results provide a systemic view on the pathobiology mechanism caused by hLRRK and S overexpression and suggest that the increased kinase activity of the hLRRK2 (R1441C) mutant results in enhanced phosphorylation of synaptojanin. These findings may contribute to develop new therapeutic strategies to prevent hLRRK2-induced Parkinson disease.