The Role Of Ataxin-3on Parkin (C289G) And Relationship With The Enzyme Activity Of Ubiquitination

Objective:Observation of the role of wild-type and mutant ataxin-3PolyQ protein on the Parkin (C289G) protein’s aggregation tendency of in the cytoplasmic and on the mitochondrial distribution and its relationship with the ubiquitin’s enzyme activitymethods:1. Use the Immunofluorescence technique to observe the role of wild-type and mutant ataxin-3PolyQ protein on the Parkin (C289G) protein’s aggregation tendency of in the cytoplasmic and its relationship with the ubiquitin’s enzyme activity2. Use the Immunofluorescence technique to observe the role of wild-type and mutant ataxin-3PolyQ protein on the Parkin (C289G) protein’s mitochondrial distribution and its relationship with the ubiquitin’s enzyme activityresults:1. In this study, immunofluorescence-confocal results show that in the co-transfected with ataxin-3-Q22and Parkin (C289G) group, or in the co-transfected with ataxin-3-Q68and Parkin (C289G) group or in the co-transfected with ataxin-3-Q22-C14A and Parkin (C289G) group, or in the co-transfected with ataxin-3-Q80-C14A and Parkin (C289G) group, all of the ataxin-3can be co-localization with Parkin (C289G). our finding suggest that ataxin-3is direct interaction with Parkin (C289G)2. In this study, Immunofluorescence shows that Parkin (C289G) protein appear significantly clustered in the cytoplasm.however,in the co-transfected with ataxin-3-Q24and Parkin (C289G) group, or in the co-transfected with ataxin-3-Q68and Parkin (C289G) group, the number of Parkin (C289G) protein aggregates were significantly reduced,(P<0.05), and the ataxin-3-Q24was more effective (P<0.05), suggest that both the ataxin-3-Q24or the ataxin-3-Q68significantly inhibited the aggregation of Parkin (C289G), and ataxin-3-Q24’s role is more apparent3. When the ataxin-3occurred the C14A point mutant, the in the co-transfected with ataxin-3-Q22-C14A and Parkin (C289G) group, or in the co-transfected with ataxin-3-Q80-C14A and Parkin (C289G) group,the number of Parkin (C289G) protein’s aggregates were also significantly increased (P <0.05), suggest that when the ataxin-3occurred the C14A point mutant, the inhibition role of the ataxin-3-Q22-C14A and the ataxin-3-Q80-C14A on protein aggregates of the Parkin mutation (C289G) were reduced significantly4. treating293T cell with CCCP12hours later,Immunofluorescence confocal laser shows that Parkin (C289G) had no significant effect on the tendency of the mitochondria’s distribution and in the co-transfected with ataxin-3-Q24and Parkin (C289G) group and in the co-transfected with ataxin-3-Q68and Parkin (C289G) group, Parkin (C289G) appear the distribution of mitochondria, indicating that ataxin-3-Q24and ataxin-3-Q68can promote the mitochondrial distribution of Parkin (C289G)5. When the ataxin-3occurred C14A point mutant, in the co-transfected with ataxin-3-Q22-C14A and Parkin (C289G) and mitochondrial plasmid groups, or in the co-transfected with ataxin-3-Q80-C14A and Parkin (C289G) and mitochondrial plasmid group, there was no significant distribution change mitochondria in Parkin (C289G), indicating that ataxin-3C14A point mutation of ataxin-3-Q22or ataxin-3-Q80had no promotion effect on the mitochondrial distribution of Parkin (C289G) proteinConclusions:1. Parkin (C289G) protein can occur significant accumulation in the cytoplasm, and whether ataxin-3wild-type or polyQ mutant significantly inhibited the aggregation, especially the wild-type ataxin-3was more effective, but the role of ataxin-3needs to rely on its ubiquitination enzymes activity.2. when the function of mitochondrial was damaged, Parkin (C289G) protein lose their ability to transfer to the mitochondria, and both wild-type ataxin-3and its mutant polyQ can significantly promote the Parkin (C289G) protein transfer to the mitochondria, but the role of ataxin-3that does not need to rely on its ubiquitin enzyme activity...