STUDIES OF FACTORS AFFECTING INTRACELLULAR TOXICITY OF THE SCA7 DISEASE PROTEIN ATAXIN - 7 : FOCUS ON ATAXIN-7 DEGRADATION AND OXIDATIVE STRESS

Abstract: Spinocerebellar ataxia type 7 (SCA7) is one of nine neurodegenerative disorders caused by expansion of CAG/polyglutamine repeats. Proteins carrying expanded polyglutamine (polyQ) domains are suggested to be resistant to degradation and aggregate. Furthermore, a negative correlation between aggregation and toxicity has been shown. So far, little is known about the turn-over rate and degradation of the SCA7 disease protein ataxin-7 (ATXN7) and how this protein induces cellular toxicity. For the studies in this thesis work, we constructed stable inducible PC12 cell lines expressing GFP-tagged ATXN7 with 10 or 65 glutamines (Qs). Using these cell lines, we studied the turn-over of ATXN7 and the relationship between mutant ATXN7 and oxidative stress.We showed that ATXN7 with a normal glutamine repeat (ATXN7Q10-GFP) has a short half-life and is mainly degraded by the UPS. In cells expressing expanded ATXN7 (ATXN7Q65-GFP), aggregation and reduced viability was observed. The aggregation increased the half-life of mutant ATXN7. For expanded full-length ATXN7, UPS was still the main degradation pathway; however autophagy also played a role in clearance of soluble ATXN7 fragments and possibly in aggregated ATXN7 material. Moreover, activation of autophagy reduced the level of aggregation and ameliorated the toxicity in cells expressing mutant ATXN7. From this study, we could get the conclusion that although expansion of the polyQ repeat increases the stability of expanded ATXN7, the protein can still be degraded via both UPS and autophagy. Furthermore, stimulation of autophagy could ameliorate the expanded ATXN7 toxicity and could therefore be a potential therapeutic approach for SCA7.Regarding the role of oxidative stress we showed that expression of mutant ATXN7 leads to increased ROS levels and oxidative stress. Treatment with an antioxidant or blockage of NADPH oxidase complexes (NOX) decreased ATXN7 aggregation, the levels of ROS and ameliorated ATXN7 induced toxicity. Based on these results, we suggest that mutant ATXN7 cause increased ROS production from NOX and antioxidants treatment and or inhibition of NADPH-oxidase might potentially be used as a therapeutic strategy in SCA7.