The Interplay between PolyQ and Protein Context Delays Aggregation by Forming a Reservoir of Protofibrils

Pastore A (2006) The Interplay between PolyQ and Protein Context Delays Aggregation by Forming a Reservoir of Protofibrils. PLoS ONE 1(1): e111. doi:10.1371/journal.pone.0000111 The Interplay between PolyQ and Protein Context Delays Aggregation by Forming a Reservoir of Protofibrils Donatella Bulone 0 1 Laura Masino 0 1 David J. Thomas 0 1 Pier Luigi San Biagio 0 1 Annalisa Pastore 0 1 0 Academic Editor: Suzannah Rutherford, Fred Hutchinson Cancer Research Center , United States of America 1 1 National Research Council (CNR), Istituto di Biofisica di Palermo , Palermo , Italy , 2 National Institute for Medical Research , London , United Kingdom , 3 Scientific Software Solutions, Paisley , United Kingdom Polyglutamine (polyQ) diseases are inherited neurodegenerative disorders caused by the expansion of CAG codon repeats, which code for polyQ in the corresponding gene products. These diseases are associated with the presence of amyloid-like protein aggregates, induced by polyQ expansion. It has been suggested that the soluble aggregates rather than the mature fibrillar aggregates are the toxic species, and that the aggregation properties of polyQ can be strongly modulated by the surrounding protein context. To assess the importance of the protein carrier in polyQ aggregation, we have studied the misfolding pathway and the kinetics of aggregation of polyQ of lengths above (Q41) and below (Q22) the pathological threshold fused to the well-characterized protein carrier glutathione S-transferase (GST). This protein, chosen as a model system, is per se able to misfold and aggregate irreversibly, thus mimicking the behaviour of domains of naturally occurring polyQ proteins. We prove that, while it is generally accepted that the aggregation kinetics of polyQ depend on its length and are faster for longer polyQ tracts, the presence of GST alters the polyQ aggregation pathway and reverses this trend. Aggregation occurs through formation of a reservoir of soluble intermediates whose populations and kinetic stabilities increase with polyQ length. Our results provide a new model that explains the toxicity of expanded polyQ proteins, in which the interplay between polyQ regions and other aggregation-prone domains plays a key role in determining the aggregation pathway. - INTRODUCTION Polyglutamine (polyQ) diseases are caused by the expansion of CAG codon repeats resulting in extended polyQ tracts in the expressed proteins [1]. This family of inherited neurodegenerative disorders includes Huntingtons chorea, spinobulbar muscular atrophy, dentatorubral-pallidoluysian atrophy, and spinocerebellar ataxias (SCAs) 1, 2, 3, 6, 7, and 17. The polyQ region is the only common feature of the proteins associated to these diseases, that are otherwise totally unrelated [2]. In affected individuals, the polyQ tract is expanded above a threshold of ca. 35 consecutive glutamines, resulting in the aggregation of the mutant protein and the consequent formation of intranuclear inclusions [3]. Although the role of aggregation and fibre formation of expanded polyQ proteins has not yet been established clearly, protein misfolding and aggregation are accepted to be central issues for understanding the molecular mechanisms of these pathologies [4]. In vitro studies have shown that polyQ aggregation depends on protein concentration, repeat-length, and time and that it occurs with a nucleation-dependent mechanism [57]. A conformational transition from random coil to b-sheet, which share most of the features typical of amyloids, takes place during the process of fibre formation [810]. However, detailed structural information on polyQ aggregates is still unavailable and the steps leading to the assembly of mature fibres are not yet fully understood. Kinetic studies of polyQ protein aggregation in vitro have shown that formation of amyloid or amyloid-like fibres generally occurs via fibrous intermediates that can have distinct morphologies [10 13]. Cell biology studies have suggested that these early aggregates or proto-fibres rather than the insoluble aggregates are the main cytotoxic species, with mature fibres having a beneficial role for neuronal cells [1417]. This hypothesis has been formulated also for other neurodegenerative diseases related to protein misfolding and aggregation, such as Parkinsons and Alzheimers diseases and the transmissible spongiform encephalopathies. Increasing evidence suggests that soluble aggregates-mediated toxicity might be a common pathogenesis mechanism for these disorders [1823]. The characterisation of the early phases of fibrillation is therefore critical for understanding the molecular causes of pathogenesis. Another central issue is the relationship between the polyQ tracts and other regions of the proteins that host them. Although polyQ expansion is certainly the main factor responsible for protein aggregation, various studies have demonstrated that the protein context plays an important role in determi (...truncated)