582. Role of MicroRNA Deregulation in Machado-Joseph Disease

Abstract

Machado-Joseph disease (MJD) or spinocerebellar ataxia type 3 (SCA3) is an autosomal dominant neurodegenerative disorder caused by an abnormal CAG expansion in the coding region of ATXN3/MJD1 gene. This confers a toxic gain-of-function to the ataxin-3 protein leading to neurodegeneration and severe motor impairments. There is no therapy to delay disease progression.MicroRNAs (miRNAs) are small non-coding RNA molecules that regulate gene expression at post-transcriptional level. miRNAs have been reported to play a significant role in a variety of diseases, including neurodegenerative disorders. However, the profiles and regulation of miRNAs in MJD are not well understood. Therefore, the aims of this study were 1) to screen the microRNA levels in a MJD mouse model and 2) to investigate functional effects of reestablishing normal levels of the de-regulated miRNAs using the same MJD mouse model.For this purpose, we used a MJD striatal lentiviral mouse model that we previously described (Simoes et al., 2012), by injecting mutant ataxin-3 in one hemisphere and wild type ataxin-3 (as an internal control) in the other hemisphere. Two weeks post-injection, miRNA expression was profiled by miRNA microarray analysis.Six miRNAs were found differentially expressed in MJD striatal samples, a result that was validated by qRT-PCR. We then used bioinformatic tools to predict the targets for the validated miRNAs and analyzed the respective levels. One candidate miR was up-regulated and the predicted target was down-regulated. To investigate whether reinstating the normal levels of the candidate miR would alleviate neuropathology, we then co-injected lentiviral vectors encoding an inhibitor of the candidate miR and mutant ataxin-3 in one hemisphere, and mutant ataxin-3 with miR negative inhibitor (as an internal control) in the other hemisphere. Five weeks post-injection we evaluated the neuropathological features and observed that in the hemisphere injected with the miR inhibitor there was a significant reduction of the loss of DARPP-32 neuronal marker and also a reduction of the mutant ataxin-3 aggregates as verified by western blot.Altogether our results provide new insights in the pathogenesis of MJD and identify a candidate miR that may be a promising target for MJD.