Characterizing the role of the polyglutamine neurodegenerative protein ataxin-3 in the cellular response to proteotoxic stress

Polyglutamine diseases are a family of neurodegenerative diseases caused by expansion of a CAG repeat that codes for a polyglutamine tract in the protein. Polyglutamine expansion appears to destabilize the protein leading to increased misfolding, altered interactions, and a toxic gain of function that leads to neurodegeneration. Many studies have shown that the normal functions as well as nuclear localization of polyglutamine disease proteins are critical for pathogenesis. Our lab has focused on understanding the normal functions of ataxin-3 (Atx3), the protein responsible for the polyglutamine disease, spinocerebellar ataxia type 3 (SCA3). Atx3 is transcriptional regulator and deubiquitinating enzyme that functions in several protein quality control (PQC) pathways. My thesis has focused on understanding the function of Atx3 in PQC and identifying cellular processes that regulate nuclear localization of Atx3. Studies presented in this thesis demonstrate that Atx3 interactions with PQC effectors, VCP and hHR23B, are dynamic and modulated by proteotoxic stress. Importantly, select proteotoxic stresses (heat shock and oxidative stress) induced nuclear localization of both WT and pathogenic Atx3. These are the first processes identified to induce Atx3 nuclear localization; this information could provide important insights into SCA3 pathology as the nucleus is likely a key sight for early pathogenesis. Mapping studies determined that two regions of Atx3 were important for heat shock-induced nuclear localization, the Josephin domain and the C-terminus containing the polyglutamine domain; serine-111 was required for nuclear localization of the Josephin domain. Atx3 null fibroblasts were more sensitive to the toxic effects of heat shock suggesting that Atx3 had a protective function in the heat shock response. Data generated while investigating a role for Atx3 in the heat shock response showed that Atx3 null cells and tissue expressed lower levels of basal Hsp70 protein compared to Atx3 WT cells and tissue; low Hsp70 protein levels in Atx3 null fibroblasts were rescued by expressing Atx3. Our data indicated that Hsp70 mRNA levels and expression of an Hsp70-luciferase reporter were decreased in Atx3 null cells which suggested that Atx3 regulated basal Hsp70 by modulating its basal transcription.