Mario Amzel

Huntingtin spheroids and protofibrils as precursors in polyglutamine fibrilization

The pathology of Huntingtons disease is characterized by neuronal degeneration and inclusions containing N-terminal fragments of mutant huntingtin (htt). To study htt aggregation, we examined purified htt fragments in vitro, finding globular and protofibrillar intermediates participating in the genesis of mature fibrils. These intermediates were high in β-structure. Furthermore, Congo Red, a dye that stains amyloid fibrils, prevented the assembly of mutant htt into mature fibrils, but not the formation of protofibrils. Other proteins capable of forming ordered aggregates, such as amyloid β and α-synuclein, form similar intermediates, suggesting that the mechanisms of mutant htt aggregation and possibly htt toxicity may overlap with other neurodegenerative disorders.

Polyglutamine fibrillogenesis: The pathway unfolds

Nine neurodegenerative diseases are caused by expanding CAG repeats coding for polyglutamine (polyGln) (1–4). These include Huntingtons disease, dentatorubral and pallidoluysian atrophy, several forms of spino-cerebellar ataxia, and spinal and bulbar muscular atrophy. Within the central nervous system, each disease has a distinctive pattern of degeneration, with considerable overlap among the diseases (5, 6). The genes containing CAG repeats show no homology to each other outside of the glutamine repeats, and most are genes of unknown function. Thus, speculation concerning pathogenesis has focused on the polyGln expansion itself.

A Compact β Model of huntingtin Toxicity

Huntington disease results from an expanded polyglutamine region in the N terminus of the huntingtin protein. HD pathology is characterized by neuronal degeneration and protein inclusions containing N-terminal fragments of mutant huntingtin. Structural information is minimal, though it is believed that mutant huntingtin polyglutamine adopts β structure upon conversion to a toxic form. To this end, we designed mammalian cell expression constructs encoding compact β variants of Htt exon 1 N-terminal fragment and tested their ability to aggregate and induce toxicity in cultured neuronal cells. In parallel, we performed molecular dynamics simulations, which indicate that constructs with expanded polyglutamine β-strands are stabilized by main-chain hydrogen bonding. Finally, we found a correlation between the reactivity to 3B5H10, an expanded polyglutamine antibody that recognizes a compact β rich hairpin structure, and the ability to induce cell toxicity. These data are consistent with an important role for a compact β structure in mutant huntingtin-induced cell toxicity.