Exploding the Repeat Length Paradigm while Exploring Amyloid Toxicity in Huntington's Disease

Area of Science:

• Neuroscience
• Molecular Biology
• Biochemistry

Background:

• Huntington's disease (HD) is a neurodegenerative disorder caused by expanded polyglutamine (polyQ) tracts in the huntingtin protein (htt).
• Identifying the specific toxic htt species has been challenging due to numerous self-associated states and overlapping formation kinetics.
• Previous research focused on polyQ length as the primary driver of toxicity.

Purpose of the Study:

• To investigate the role of htt aggregation states in Huntington's disease toxicity.
• To determine if amyloid aggregates, rather than polyQ repeat length, are the primary toxic species.
• To develop novel htt variants to dissect aggregation pathways and toxicity.

Main Methods:

• Engineered pro-β-hairpin (βHP) and β-breaker polyQ and htt-exon1 (htt-ex1) variants with specific mutations.
• Utilized fluorescence correlation spectroscopy (FCS) and a novel thioflavin-T (ThT) assay to characterize htt-ex1 assembly intermediates.
• Expressed engineered htt variants in mammalian cells (PC12), rat neuronal models, and *Drosophila* models of HD.

Main Results:

• Expanded polyQ htt-ex1 versions showed tetramers, oligomers, and fibrils, but not monomers, correlating with early toxicity.
• Engineered htt-ex1-βHP variants with short polyQ repeats but strong aggregation were toxic, challenging the repeat length paradigm.
• Non-toxic β-breaker variants favored non-amyloid oligomers, while toxic βHP variants rapidly formed amyloid structures.

Conclusions:

• Amyloid aggregates of huntingtin protein, rather than polyQ length or non-amyloid oligomers, are strongly implicated as the major toxic species in Huntington's disease.
• The formation of the fundamental amyloid folding motif is critical for htt-induced toxicity.
• Engineered htt variants provide powerful tools to study HD pathogenesis and break the traditional repeat length paradigm.