DYNAMICS AND NUCLEAR EFFECTS OF NON-POLYQ AGGREGATES

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Protein aggregation and the formation of inclusion bodies are two hallmarks of the cytopathology in neurodegenerative diseases. In diseases such as Parkinson's and Alzheimer's, the deposits are cytoplasmic, while in Huntington's disease and the ataxias, the deposits form predominantly within the nuclei. The inclusions recruit key cellular components, such as chaperones and proteasomes, nuclear matrix components, and specific transcriptional regulators. The aim of this proposal is to explore the hypothesis that the pathology associated with aggregation is related to the sequestration of cellular components involved in critical cellular processes such as protein folding and degradation, and gene expression. To this end, we plan to use a novel model system, based on an artificial protein we call GFP170*. GFP170* lacks polyQ stretches, yet induces cellular responses analogous to those caused by the deposition of polyQ aggregates. Like polyQ proteins, GFP170* forms cytoplasmic and nuclear aggregates, recruits cytoplasmic and nuclear components, alters transcriptional regulation, and kills cells. We propose twin lines of investigation. First, we will examine the physical nature of the sequestered cellular components. GFP-based real-time imaging of live cells will be used to explore the mobility of GFP170* and of cellular components recruited to the aggregates. Such data will inform on the physical stability of the aggregates, and will influence possible models of cytopathology. Second, we will follow the functional consequences of nuclear aggregation on gene expression. In vivo read-out systems and genomic approaches will be used to explore the modulation of transcriptional regulators by aggregating GFP170*. Completion of the proposed studies will establish a firm basis for defining the role of nuclear aggregates in cytopathology.