GENETIC AND PHARMACOLOGICAL INHIBITORS OF NECROPTOSIS IN STUB1-DEPENDENT ATAXIAS

The inability to maintain cellular protein quality control (PQC) is often linked to neurodegenerative conditions. The carboxyl terminus of heat shock 70-interacting protein (CHIP) is a well-established regulator of PQC due to its ability to function as both a co-chaperone and ubiquitin ligase. Coding mutations in STUB1, the gene that encodes CHIP, are causal for two forms of ataxias known as Spinocerebellar Ataxia Autosomal Recessive 16 (SCAR16) and Spinocerebellar Ataxia 48 (SCA48). Previous studies by our lab and others suggest that a pro-inflammatory form of cell death known as necroptosis may promote the cerebellar degeneration observed in SCAR16 and SCA48 patients. The receptor-interacting protein kinases 1 and 3 (RIPK1/3), critical kinases involved in necroptotic activation, are known CHIP substrates. In this dissertation, we set out to investigate the therapeutic impact that RIPK1/3 inhibition has on disease progression and severity in preclinical mouse models of STUB1-dependent ataxias. In chapter two, we subjected preclinical mouse models of STUB1-dependent ataxias to a year-long behavioral battery assay to investigate how CHIP dysfunction impacts neurodegeneration and ataxic severity. Here, we inhibited the necroptotic signaling pathway by genetically or pharmacologically targeting RIPK1/3 to determine the therapeutic effects of long-term necroptotic inhibition on motor coordination and cognitive function. We demonstrate that STUB1 disease mutations cause an increase in motor and cognitive dysfunction resulting in an accelerated aging process. However, mice with inhibited RIPK1/3 demonstrated improved motor coordination, cognitive function, and viability throughout aging as compared to those that were untreated. In chapter three, we successfully demonstrated the ability to culture organotypic cerebellum slices from aged mice. We observed that specific alterations to the culture media allowed for the prolonged viability of adult cerebellar brain slices and the preservation of aging-related structural, biological, and functional changes. The development of this model addresses one of the most significant rate-limiting steps preventing us from discovering more about necroptosis and its role in disease. Altogether, the findings presented in this dissertation demonstrate the potential that druggable targets within the CHIP-regulated necroptotic signaling pathway could have on mitigating cell death, ultimately providing hope for ataxic therapeutic candidates.