The ordered aggregation of proteins into amyloid fibrils is associated with a variety of human diseases, including Alzheimer’s (AD), Parkinson’s (PD), and Huntington’s disease (HD). Amyloid formation occurs via a series of oligomeric intermediates. The complex diversity of aggregates complicates the assignment of toxic mechanisms to specific species. With regard to HD, aggregation is triggered by an expanded polyglutamine tract (polyQ) in the huntingtin protein (HTT). The polyQ expansion associated with HD is typically greater than 40 repeat glutamine residues. Here, a simple centrifugal separation method was implemented to isolate distinct, relatively homogeneous aggregate populations of a mutant huntingtin protein (mHTT) with a polyQ length of 46. These mHTT aggregates were characterized via atomic force microscopy (AFM) and dynamic light scattering. Different aggregate species were then exogenously introduced to Caenorhabditis elegans ( C. elegans ) to determine their impact on viability and motility. The mHTT oligomers were significantly toxic to wild-type C. elegans , but fresh preparations of mHTT or fibrils did not impact the worms. However, exposing C. elegans that express a nontoxic form of HTT to mHTT fibrils results in reduced viability. That is, fibrils themselves were not appreciably harmful to C. elegans , but if there was a pool of HTT monomers that could be seeded by the presence of HTT, exposure to exogenous fibrils was toxic. To further explore oligomer toxicity, several strategies were employed to manipulate oligomers, including chemical cross-linking, peptide-based aggregation inhibitors, and a variety of small molecules that alter mHTT aggregation. Overall, this approach offers a straightforward method to study and modulate the toxicity of specific mHTT aggregate species in a controlled setting. In addition, AFM was used to probe the impact of expressing nonpathogenic HTT or mHTT on age-related cuticle roughening and decreased mechanical integrity in C. elegans .
Justin Legleiter (Sun,) studied this question.