Orientador: Iscia Lopes Cendes / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-04T00:30:33Z (GMT). No. of bitstreams: 1 Costa_FlaviaChagas_M.pdf: 2665617 bytes, checksum: b552b22eff093aa5e800faf30ad82041 (MD5) Previous issue date: 2000 / Resumo: As ataxias espinocerebelares (ABC) formam um grupo heterogêneo de doenças degenerativas que envolvem o sistema nervoso central. Esse grupo se caracteriza clinicamente por apresentar disfunção cerebelar manifestada por ataxia de marcha, incoordenação e disartria. Nos casos familiares, o padrão de herança é variável, podendo ser compatível com herança autossômica dominante (HAD) ou herança autossômica recessiva (HAR). Para as ataxias espinocerebelares com HAR existem três lócus identificados até o momento: ataxia de Friedreich (AP) no cromossomo 9q; ataxia com deficiência de vitamina E (ADVE) no cromossomo 8q,. e uma forma muito rara de início da inf'ancia, IOSCA (Infantile Onset Spinocebellar Ataxia) no cromossomo 10q. A maioria dos casos de AP é causada por uma expansão em homozigose do trinucleotídeo (GAA)n localizado no primeiro ÍI1tron do gene -125. Esse gene codifica uma proteína de 210 aminoácidos denominada de Frataxina, com alvo mitocondrial. Alelos normais variam de sete a 30 unidades (GAA)n enquanto que alelos expandidos' variam de 100 a 1700 (GAA)n. A ADVE é clinicamente semelhante à AP, sendo que a maneira mais confiável de realizar o diagnóstico diferencial entre essas duas formas de ataxia é através da análise molecular. A ADVE é causada por mutações de vários tip?s ,e~ontradas no gene a-TTP, que codifica a proteína transportadora do alfa-tocoferol, com 278 aminoácidos. Pelo menos 13 tipos de mutações foram descritas até o momento. Neste estudo, foram genotipados 97 indivíduos pertencentes a 58 famílias não relacionadas. Uma freqüência de 15% de indivíduos afetados com a mutação AP foi encontrada Este é um estudo pioneiro no Brasil, onde estudamos a freqüência das mutações responsáveis pelas ataxias espinocerebelares com fIAR determinamos as características das mutações encontradas e realizamos estudos de correlação entre o genótipo e características feno típicas desses pacientes / Abstract: The spinocerebellar ataxia form a group of neurodegenerative disorders with clínical heterogeneity. They are characterized by cerebelIar dysfunction with progressive ataxia, incoordenation and dysarthria. Familial cases show variable pattem of inheritance: autossomal dominant (AD) or autossomal recessive (AR). To date, thereare four loci described for the spinocerebelIar ataxia with AR: Friedreich ataxia (F A) on chromosome 9q; ataxia with vitamin E deficiency (ADVE) on chromosome 8q, the lnfantile Onset SpinocerebelIar Ataxia (IOSCA) on chromosome 10q and ataxia telangiectasia on chromosome llq. the %25 gene. This gene encodes for a 210 aminoacids protein named Frataxin, with mitochondrial target. Normal (GAA)n alleles range ITom seven to 30 (GAA)n units and expanded (GAA)n alleles have 200 to 1700 (GAA)n units. ADVE is clinically identical to the FA,-and the best way to di:fferentiate between the two forms is by molecular testing. ADVE is caused by di:fferent types of mutations, found in the a-TTP gene, with codes for alpha tocopherol transfer protein, with 278 aminoacids. To date, there are 13 di:fferent mutations reported in a-TTP. We studied 97 individuais, belonging to 58 unrelated families. The ITequency of patients with FA mutation was 15% among our patients. No mutations were found in a-TTP gene by SSCP ana1ysis. This is the first report on the fTequency of different mutations causing spinocerebellar ataxia with AR inheritance in the Brazilian population. In addition, we analyzed the molecular characteristics of the mutations detected and performed genotype - phenotype correlation / Mestrado / Ciencias Biomedicas / Mestre em Ciências Médicas

Friedreich ataxia (FRDA) is an inherited autosomal recessive neurodegenerative disorder caused by a GAA trinucleotide repeat expansion mutation within the first intron of the FXN gene. Normal individuals have 5 to 30 GAA repeats, whereas affected individuals have from approximately 70 to more than 1,000 GAA triplets. In addition to progressive neurological disability, FRDA is associated with cardiomyopathy and an increased risk of diabetes mellitus. Currently there is no effective therapy for FRDA and this is perhaps due to the lack of an effective system to test potential drugs. Therefore, the main aim of this thesis is to develop a novel cell culture system, to aid in rapid drug screening for FRDA. Firstly, I have demonstrated the establishment of novel cell culture systems, including primary fibroblasts, neural stem cells (NSC) and splenocytes, from FRDA YAC transgenic mouse models (YG8 and YG22). Then, I have shown the differentiation of NSCs into neurons, oligodendrocytes and astrocytes. The presence of these cells was confirmed by using cell specific immunofluorescence assays. I have also shown that both YG8 and YG22 rescue mice have less tolerance to hydrogen peroxide induced oxidative stress than WT mice, as similarly seen in FRDA patient fibroblasts. Recent findings indicate that FRDA is associated with heterochromatin-mediated silencing of the FXN gene accompanied by histone changes, flanking the GAA repeats. This suggested potential therapeutic use of compounds which can reduce the methylation and increase the acetylation of histone proteins. Therefore, using human and mouse primary fibroblast cell lines I have investigated the efficacy and tolerability of various DNA demethylating agents, GAA interacting compounds and class III histone deacetylase (HDAC) inhibitors. Although DNA demethylating agents showed increased FXN expression, no correlation between the level of DNA methylation and FXN expression was identified. Nevertheless, the use of GAA interacting compounds, particularly DB221, and the HDAC inhibitor, nicotinamide, have shown encouraging results, provoking us to use such compounds in future long-term in vivo studies. In addition, I have also investigated the long-term efficacy of two benzamide-type HDAC inhibitors, RGFA 136 and RGFP 109, on the FRDA YAC transgenic mice. No overt toxicity was identified with either drug, indicating a safe administration of these compounds. Both compounds produced improved functional analysis together with significantly reduced DRG neurodegeneration. However, neither of these compounds was shown to significantly increase the FXN mRNA expression. Nevertheless, elevated levels of frataxin protein in the brain tissues were obtained with RGFP 109, suggesting that RGFP 109 is capable of crossing the blood-brain barrier. I have also found increased levels of global acetylated H3 and H4 histone proteins in brain tissues, along with significant increase in aconitase enzyme activity, particularly with RGFP 109 treatments. Overall, these results support future clinical trial development with such compounds.

Spinocerebellar ataxia type 1 (SCA1) is an adult onset, autosomal dominant neurodegenerative disease caused by a CAG repeat expansion in ataxin-1, which encodes the ataxin-1 protein. SCA1 is one of nine polyQ-expansion gain-of-function diseases which includes Huntington's disease, spinal-bulbar muscular atrophy, dentatorubral-pallidoluysian atrophy and other ataxias. Clinical symptoms of SCA1 include ataxia, dysarthria, ophthalmoparesis, muscle wasting, and extrapyramidal and bulbar dysfunction. Cerebellar Purkinje cells (PCs), neurons in the inferior olive and nuclei of the brainstem are affected. No disease-modifying therapy exists for SCA1. The goals of my thesis were to assess the safety and efficacy of AAV-delivered artificial miRNAs targeting ataxin-1 to alleviate neuropathological and behavioral phenotypes in the knock-in and transgenic SCA1 mouse models. In the knock-in SCA1 mouse model AAVs expressing an artificial miRNA (miSCA1) targeting sequences conserved in mouse and human ataxin-1 were injected directly to the deep cerebellar nuclei. This achieved long term silencing of ataxin-1 mRNA and significantly improved rotarod performance, gait deficiencies, and neuropathology of the cerebellum. In the transgenic SCA1 mouse model the same method of delivery was executed with an artificial microRNA (miR) (miS1) designed to optimize potency, efficacy and safety to suppress Atxn1 expression. Additionally the therapeutic potential of continuous overexpression of ataxin-1-like was examined. Delivery of either ataxin-1-like or miS1 viral vectors to SCA1 mouse cerebellum resulted in widespread cerebellar Purkinje cell transduction. There was significant improvement to rotarod performance, gait deficiencies, coordination and balance, as well as the neuropathology of cerebellar Purkinje cells. In summary, these data indicate the utility of these approaches as possible therapies for SCA1 patients.

Thesis (Ph. D.)--University of Alabama at Birmingham, 2007. / Title from first page of PDF file (viewed Feb. 6, 2008). Includes bibliographical references.

Orientador: Marcondes Cavalcante França Júnior / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-26T03:00:02Z (GMT). No. of bitstreams: 1 Silva_CynthiaBonilhada_D.pdf: 5479490 bytes, checksum: 024402553720f1d64b73e9db5fae2ab4 (MD5) Previous issue date: 2014 / Resumo: Ataxia de Friedreich é a ataxia autossômiva recessiva mais frequente, causada por uma expansão de tripletos GAA em homozigoze no primeiro íntron do gene FXN, localizado no cromossomo 9. Trata-se de uma doença neurodegenerativa de início precoce, com curso progressivo. A patologia é caracterizada por perda neuronal nos gânglios das raízes dorsais, seguida por degeneração da coluna dorsal da medula espinhal e dos tractos espinocerebelares, atrofia da medula espinhal e do núcleo denteado de cerebelo. Os estudos clínicos prévios realizados na Ataxia de Friedreich focaram na incapacidade motora e muito pouco é conhecido sobre sintomas não-motores. Da mesma forma, estudos de neuroimagem prévios avaliaram coortes reduzidas e não incluíram avaliação longitudinal. Dessa forma, nosso estudo foi desenhado para caracterizar melhor a história natural da Ataxia de Friedreich. Avaliamos a presença de sintomas neuropsiquiátricos e se há substrato anatômico para tais manifestações. Realizamos, ainda, estudo de neuroimagem multimodal e longitudinal, a fim de identificar possíveis marcadores evolutivos da doença. Os resultados mostraram que fadiga e depressão são freqüentes nos pacientes com Ataxia de Friedreich, enquanto que sintomas relativos ao sono são raros. Evidenciamos que a depressão apresenta correlação com alterações neuroanatômicas, especialmante com a atrofia do giro cingulado anterior. O tempo de relaxação T2 estava significativa e especificamente reduzido nos núcleos denteados dos pacientes. Este parâmetro apresentou progressão ao longo do tempo e de forma paralela à deterioração motora, sugerindo assim que o tempo de relaxação T2 nesta estrutura pode ser um marcador de evolução da doença. O estudo volumétrico evidenciou uma extensa atrofia de substância cinzenta e branca, incluindo regiões profundas do cerebelo, tronco encefálico e também estruturas supratentoriais, como os giros precentrais. Estas alterações se correlacionaram com parâmetros de gravidade da doença. Na avaliação longitudinal, houve piora da atrofia em regiões dos lobos temporais e frontais (incluindo os giros precentrais), além do lobo posterior do cerebelo. Nos pacientes, evidenciamos também extenso dano microestrutural à substância branca na FRDA. Além dos pedúnculos cerebelares superiores, constatamos alterações em parâmetros de difusão nos tratos piramidais e no corpo caloso. Embora estas alterações tenham se correlacionado com a incapacidade motora, não detectamos progressão ao longo do tempo / Abstract: Friedreich¿s ataxia is the most common autosomal recessive ataxia. It is caused by a homozygous triplet GAA expansion in the first intron of the FXN gene on chromossome 9. It is an early onset disease, with slowly progressive evolution. Pathology is characterized by neuronal loss in the dorsal root ganglia, followed by degeneration of the dorsal columms of the spinal cord, spinocerebelar tracts and atrophy of dentate nuclei of cerebellum. On clinical grounds, most studies have focused on motor disability and little is known about non-motor symptoms. Also, previous neuroimaging studies evaluated small samples of patients and did not include longitudinal analysis. Therefore, our study was designed to better characterize the course of the disease. In particular, we looked at neuropsychiatric manifestations and their structural substrate. We also performed multimodal MRI scans in a longitudinal setting, in an attempt to find reliable neuroimaging markers. Our results showed that fatigue and major depression are indeed frequent in patients with Friedreich¿s ataxia, but sleep complaints are rare. In these patients, depression was associated with neuroanatomical abnormalities, especially anterior cingulate cortex atrophy. The dentate nuclei T2 relaxometry was significantly shorter in patients, presented a progressive worsening over time and correlated with clinical parameters. These data suggest that dentate nuclei T2 might be a useful marker in this disease. The volumetric analyses showed widespread gray and white matter atrophy, including deep cerebellar nuclei, brainstem and also supratentorial structures, such as the precentral gyri. Such abnormalities correlated with disease severity. After 2 years of follow-up, we identified progressive volumetric reduction in parts of the temporal and frontal lobes (including the precentral gyri), as well as the posterior cerebellar lobe. We also found widespread microstructural damage to the white matter in Friedreich's ataxia. Such damage involved not only the superior cerebellar peduncles, but also the pyramidal tracts and the corpus callosum. Although these abnormalities did correlate with motor disability, we did not find progression over time / Doutorado / Neurologia / Doutora em Ciências Médicas

Orientadores: Marcondes Cavalcante França Junior, Anamarli Nucci / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-21T18:19:14Z (GMT). No. of bitstreams: 1 Takazaki_KarenAntoniaGirotto_M.pdf: 1348260 bytes, checksum: 48e434fbe077053824d92b0221e439fc (MD5) Previous issue date: 2012 / Resumo: A SCA3/MJD é a ataxia hereditária mais freqüente em nosso meio e apresenta sintomatologia bastante variada, incluindo aspectos motores e não - motores. Dentre estes, ressaltamos a disfunção autonômica, achado pouco estudado e muitas vezes subestimado nestes pacientes. Neste estudo tivemos por objetivos: 1. determinar a frequência e a intensidade das manifestações autonômicas na SCA3/MJD; 2. avaliar se a disfunção autonômica envolve o sistema simpático e/ou o parassimpático; 3.determinar se existe correlação entre as manifestações autonômicas, parâmetros genéticos e clínicos; e 4. verificar se existe correlação entre as manifestações autonômicas e o envolvimento do sistema nervoso periférico. Para isso realizamos uma avaliação clínica e eletrofisiológica em 40 pacientes com confirmação molecular da doença e 38 controles saudáveis pareados por idade e sexo. Ambos os grupos foram submetidos a exame clínico, incluindo o uso da escala SARA (Scale for the Assessment and Rating of Ataxia), o questionário SCOPA-AUT (Scales for Outcomes in Parkinson's Disease - Autonomic Questionnaire), além da aferição da pressão arterial nas posições supina e ortostática. Também realizamos o estudo da variabilidade da frequência cardíaca no repouso, no desafio ortostático (razão 30:15), durante a manobra de Valsalva (índice de Valsalva) e na respiração profunda (razão E:I). Realizamos ainda a análise espectral dos intervalos RR no repouso e a resposta simpática cutânea. As queixas mais frequentes dos pacientes relacionam-se aos sintomas do controle urinário, cardiovascular e sudomotor. Observamos diferença significativa na média dos intervalos RR em repouso dos pacientes em relação aos controles (811,8 x 933,4 ms; p=0,001). A análise da razão 30:15 também mostrou diferença significativa entre os dois grupos (1,10 x 1,15; p=0,038). Já o índice de Valsalva e a razão E:I não mostraram diferença significativa (p=0,373 e p=0,08). A análise espectral mostrou diferença significativa entre os grupos, em relação ao LFPA (poder de baixa frequência), com média de 23,6 x 43,3 ms² (p<0,001), e ao HFPA (poder de alta frequência), com média de 28,8 x 54,4ms² (p<0,001). Obtivemos a frequência de 30% de disautonomia cardiovascular e 45% de disautonomia simpática sudomotora nos pacientes com SCA3/MJD. Verificamos envolvimento tanto do simpático quanto do parassimpático. Não observamos correlação entre as manifestações autonômicas e parâmetros genéticos (tamanho da expansão CAG) ou clínicos, mas encontramos correlação entre disfunção autonômica simpática sudomotora e o envolvimento do sistema nervoso periférico / Abstract: SCA3/MJD is the most frequent autosomal dominant ataxia worldwide and characterized by a variety of symptoms, including motor and non-motor manifestations. Autonomic dysfunction has been described in SCA3/MJD, but there are only small studies and several important questions remain unanswered. In this study we had the following objectives: 1. To determine the frequency and the intensity of the autonomic manifestations in SCA3/MJD. 2. To evaluate whether the autonomic dysfunction compromises the sympathetic and/or the parasympathetic system. 3. To identify possible correlations between the autonomic manifestations, genetic and clinical parameters. 4. To verify whether there is correlation between the autonomic manifestations and peripheral nervous system damage. We have thus performed clinical and electrophysiological evaluation of 40 patients with molecular confirmation of SCA3/MJD and 38 healthy controls matched by age and gender. In the clinical study we used the SARA scale (Scale for the Assessment and Rating of Ataxia), the SCOPA-AUT questionnaire (Scales for Outcomes in Parkinson's Disease - Autonomic Questionnaire), and we measured blood pressure in supine and orthostatic positions. We also studied the heart rate variability at rest, during the orthostatic challenge (30:15 ratio), during the Valsalva maneuver (Valsalva index) and deep breathing (E:I ratio). We included in our study the spectral analysis of the RR intervals at rest and the cutaneous sympathetic response. The most frequent complaints in our patients are related to urinary, cardiovacular and sweat control. We found significant difference of the mean RR intervals at rest between patients and controls (811.8 x 933.4 ms; p=0.001 respectively). The 30:15 ratio was also different between the two groups (1.10 x 1.15; p=0.038 respectively). The Valsalva index and the E:I ratio were similar between the groups (p=0.373 and p=0.08). Spectral analysis presented distinct results in patients and controls, related to LFPA - low frequency power (p<0.001) and to HFPA - high frequency power (p<0.001). We found cardiovascular and sympathetic sweat disautonomia in 30% and 45% of the patients with SCA3/MJD, respectively. We found evidence of both sympathetic and parasympathetic dysfunction. We did not find correlation between autonomic manifestations and genetic (CAG repeat length) or clinical parameters, but we found correlation between sympathetic sweat autonomic dysfunction and peripheral nervous system damage / Mestrado / Neurologia / Mestra em Ciências Médicas

Ataxia-telangiectasia mutated gene product (ATM) is a 350 kDa Serine/Threonine kinase belonging to the family of Phosphatidylinositol-3 kinase like kinases. ATM functions as a key element in DNA Damage Response (DDR), a mechanism that maintains genomic integrity within the cells. ATM is activated after double stranded DNA damage and initiates signalling cascades that determine the process of decision-making of cell fate and involves the participation of multiple proteins. This vital protein acts first by sensing double stranded DNA breaks and second by transducing the signal and activating other downstream proteins of the repair pathway via its kinase function. This provides an important link between signals generated after DNA damage, the cell cycle pathway and apoptotic machinery. This function is crucial for mammalian cells which are constantly challenged by genotoxic agents from a variety of sources and therefore require a robust sensing and repair mechanism to maintain cell vitality. Cells lacking ATM are hypersensitive to cytotoxic insults, particularly genotoxic stress, induced through radiation or radiomimetic drugs. This thesis describes the discovery and characterisation of novel autoregulatory feedback loops of ATM kinase in human cells. Firstly, I have discovered that inhibition of ATM kinase activity causes induction of ATM protein expression followed by time dependent oscillations. This novel autoregulatory mechanism was demonstrated in cell cycle independent manner and both in the absence and presence of DNA damage. ATM promoter assay revealed that this autoregulation was governed at the transcriptional level. Furthermore, this autoregulatory induction of ATM was also accompanied by a transient upregulation of P53, pATR and E2F1 levels. Elucidation of the underlying trafficking mechanism of ATM during such autoregulation and in DDR also revealed a novel ATM sub-cellular trafficking mechanism which was dependent on its own kinase activity. This trafficking mechanism involved DNA damage induced Golgi to nuclear transport of phosphorylated ATM S-1981 to elicit DDR. This was found to be a conserved pathway required during the initiation of DDR and was demonstrated in multiple cell lines. Further studies into the sub-cellular transport machinery revealed the involvement of β-COPI coatomer protein in this mechanism of ATM trafficking, which was found to be autoregulated by ATM kinase, and required 387-388 ATM di-Lysine motif. The discovery of these functionally important autoregulatory mechanisms of ATM were further utilised to develop Luciferase reporter based biosensor of DNA damage and single cell fluorescence based ATM inhibition assay to screen for ATM inhibitors. Finally, following the discovery and characterisation of these functional spatio-temporal autoregulations of ATM, quantitative estimations of the kinetics of signalling cascades initiated by it during DDR and its overall outcome on cellular fate were determined to study ATM pathway systematically for employing a quantitative systems biology approach. These novel findings have immensely increased our understanding of ATM regulation and function. Elucidation of the mechanisms of novel autoregulations of ATM provide new dimensions through which DDR pathway could be manipulated, and as such could be utilised for achieving targeted cellular sensitivity in therapeutic intervention of cancer.

El presente estudio de investigación, tiene como objetivo determinar la incidencia de artrosis de cadera y la incidencia de dismetría de miembro inferior en la población adulta menor de 65 años, y conocer la distribución que tiene la artrosis de cadera por grupo etario y sexo.

Dans de nombreuses maladies héréditaires monogéniques, les bases moléculaires du mode de transmission (dominant vs récessif) et l'origine de nombreux phénotypes associés restent obscurs. C'est le cas pour les ataxies épisodiques (EA) et progressives liées au canal calcique Cav2.1. La plupart des mutations présentes dans ces ataxies génèrent une protéine mal repliée retenue et dégradée dans le réticulum endoplasmique. J'ai exploré plusieurs approches afin d'associer les phénotypes dues à la présence des mutants et ceux dues à une « pure » perte de fonction. A l'aide de vecteurs viraux codants pour des ARNs interférents de Cav2.1, j'ai montré que la réduction chez la souris adulte du canal récapitulait de nombreux symptômes de l'EA2.J'étudie également un nouveau modèle murin knock-In pour l'EA2 produisant une forme tronquée mal repliée. L'étude de ces modèles permettra une meilleure compréhension des mécanismes physiopathologiques et l'exploration de nouvelles voies thérapeutiques. / Voltage-gated calcium channels (VGCC) regulate an array of physiological process. The P/Q-type VGCC is principally expressed in the cerebellum and at the neuromuscular junction, where it plays an essential role at the presynaptic neuronal nerve. Interestingly, mutations in a1 subunit (Cav2.1) of P/Q-type VGCC gene have been found to be linked for three autosomal dominant human disorders, familial hemiplegic migraine type 1 (FHM1), spinocerebellar ataxia 6 (SCA6) and episodic ataxia type 2 (EA2). Mutations causing EA2 lead to loss-of-function of Cav2.1 channels and are principally non-sense. The origin of dominant transmission and the heterogeneity of the symptoms are not known. Recent data from different groups have shown a dominant-negative effect of Cav2.1 mutants in cultured cell lines. Indeed, the molecular mechanism of this dominant-negative effect appears to be due to the interference of EA2 mutants with the folding of the wildtype subunit, thereby abolishing channel activity. This destructive interaction mechanism promoted by the EA2 mutants is likely to occur in the disease.The first part of my thesis was to monitor the effect of a “pure” silincing of P/Q-type channel in adult mice. Suppression of Cav2.1 channel by RNAi lentiviral based-vector approaches has produced episodic as well as permanent ataxia without signs of progressive ataxia. As a direct approach to understanding the physiologicalcontributions of misfolded truncated mutants in EA2 phenotypes, a Cav2.1 knock-inmutant, CACNA1AR1479x has been generated. This is a fundamental issue to understand the pathogenic mechanisms of EA2 and more generally to the other neuronal and neuromuscular diseases.

Polyglutamine (polyQ) diseases are progressive fatal neurodegenerative movement disorders. Although many cellular processes are perturbed in polyQ disease, recent studies highlight the importance of protein misfolding as a central event in polyQ toxicity. Spinocerebellar ataxia type 3 (SCA3), also known as Machado-Joseph disease, is a particularly interesting polyQ disease because of the special qualities of the disease protein ataxin-3, which normally participates in cellular protein quality control. Here I use multiple mouse models of disease to explore toxic protein species and the role of protein homeostasis in SCA3 pathogenesis. In Chapter 1, I review the key features of polyQ disease, and outline the background and rationale behind our strategy for identifying toxic protein species in SCA3. In Chapter 2, I examine the role of the protein quality control ubiquitin ligase, CHIP (C-terminus of Hsp70 interacting protein), in regulating the toxicity of expanded ataxin-3 in vivo. Genetic reduction or removal of CHIP increases formation of detergent-resistant ataxin-3 microaggregates specifically in the brain. Concomitant with this, reduction or removal of CHIP exacerbates the phenotype of SCA3 mice, revealing a correlation between high levels of microaggregates and phenotypic severity. Additional cell-based studies confirm that CHIP may not directly mediate ataxin-3 degradation, suggesting that CHIP reduces expanded ataxin-3 toxicity in the brain primarily by enhancing ataxin-3 solubility. In Chapter 3, I use various biochemical techniques to reveal the presence of brain-specific ataxin-3 microaggregates in two genetically distinct mouse models of SCA3. Selective neuropathological evaluation of SCA3 mice reveals that major neuronal loss and reactive glial proliferation are not shared features of phenotypically-manifesting SCA3 mice. Additional studies fail to provide evidence for loss-of-function of endogenous ataxin-3 in SCA3 mice. Our results suggest that neuronal dysfunction in SCA3 is mediated through a toxic gain-of-function mechanism by ataxin-3 microaggregates in the CNS. In Chapter 4, I discuss important areas for future research in polyQ disease. I describe studies that would help elucidate the structural nature of toxic soluble microaggregates, and their effects on other cellular proteins. I also consider how the results described in this thesis inform potential treatment strategies.