The role of protein aggregation in Huntington's disease
thesisposted on 19.05.2010, 11:54 by Claire Martina Didszun
Huntington’s disease (HD) is a hereditary progressive neurodegenerative disorder characterised by chorea, general motor impairment, psychiatric disturbances and dementia, leading to death within 10 to 20 years of onset. It is caused by a (CAG)n trinucleotide repeat expansion in the gene IT15 that is translated into a prolonged polyglutamine tract in the protein huntingtin. This mutation leads to the self-association of huntingtin to form aggregates in the brains of affected people. Work in Professor Nicotera’s group at the MRC Toxicology Unit identified expression changes in Rab11, a protein involved in endosomal recycling, in a cell culture model of HD. The aim of this thesis was to investigate whether endosomal recycling is affected in this model. Monitoring the trafficking of labelled transferrin by Western blotting and live-cell imaging showed rapid uptake of transferrin into all cells, but a significantly reduced rate of clearance in cells containing huntingtin aggregates, which was associated with the accumulation of transferrin in the endosomal recycling compartment (ERC). This finding demonstrates an aggregate-specific lesion in the exit of cargo from the ERC in the absence of cell death. As endosomal recycling is essential for correct neuronal function, this process provides a mechanism whereby protein inclusions may contribute to the cognitive and motor deficits seen in HD. A second project focused on purification of aggregates to determine if they could sequester material important for cellular function. Many attempts to purify the aggregates in their intact globular form revealed their highly unspecific affinity towards other proteins, which renders purification methods very susceptible to artefacts. The large globular aggregates could, however, be dispersed by SDS-treatment into fibrils of approximately 10 nm diameter, which were subsequently purified and visualised by electron microscopy.