A network of protein interactions determines polyglutamine toxicity

Abstract

Several neurodegenerative diseases are associated with the toxicity of misfolded proteins. This toxicity must arise from a combination of the amino acid sequences of the misfolded proteins and their interactions with other factors in their environment. A particularly compelling example of how profoundly these intramolecular and intermolecular factors can modulate the toxicity of a misfolded protein is provided by the polyglutamine (polyQ) diseases. All of these disorders are caused by glutamine expansions in proteins that are broadly expressed, yet the nature of the proteins that harbor the glutamine expansions and the particular pathologies they produce are very different. We find, using a yeast model, that amino acid sequences that modulate polyQ toxicity in cis can also do so in trans. Furthermore, the prion conformation of the yeast protein Rnq1 and the level of expression of a suite of other glutamine-rich proteins profoundly affect polyQ toxicity. They can convert polyQ expansion proteins from toxic to benign and vice versa. Our work presents a paradigm for how a complex, dynamic interplay between intramolecular features of polyQ proteins and intermolecular factors in the cellular environment might determine the unique pathobiologies of polyQ expansion proteins.huntingtin ͉ Huntington's disease ͉ protein misfolding ͉ yeast A variety of human diseases are characterized by the accumulation of aggregated, misfolded proteins (1, 2). In some cases, such as cystic fibrosis, disease is caused by the loss of a vital protein function due simply to this misfolding and aggregation. In other cases, however, protein misfolding creates novel, toxic properties. For example, in Alzheimer's disease, Parkinson's disease, and the polyglutamine (polyQ) disorders, misfolded proteins disrupt proper cellular function and cause cytotoxicity (3,4). In all of these diseases the amino acid sequences of the particular misfolded protein and their interactions determined by its specific environment must govern toxicity. Yet, despite intense research, we know little about these intramolecular and intermolecular factors. Without identifying and understanding these factors at a molecular level it will be difficult to devise the most effective therapeutic strategies to treat protein misfolding diseases.The polyQ diseases present a promising starting point to study this very general problem. In all of these diseases polyQ expansions constitute the molecular basis of disease (5). However, the disease pathologies (including the cell types most strongly affected) and the individual proteins that bear the disease-causing polyQ expansions are very distinct (5). Consequently, the amino acids flanking the polyQ region and the sets of cellular factors that specifically interact with each polyQ expansion protein are different. Combinations of these factors must account for their different pathobiologies. Recent studies have started to identify protein-protein and genetic interaction networks for polyQ proteins (6-8), but the current data do not yet ...

Chaperone-dependent amyloid assembly protects cells from prion toxicity

“…1a). Cell growth defects observed were more extreme than any we have observed with other misfolded proteins in yeast (21,22). At this modest level of Rnq1 overexpression, Ϸ25% of [RNQ ϩ ] cells were dead within 4 h, as determined by the percentage of colony-forming units and dye exclusion (data not shown).…”

“…Case in point, when GPI-anchorless PrP was expressed together with WT PrP, it accelerated scrapie disease and resulted in increased deposits of both amyloid and nonamyloid proteinase K-resistant PrP (32). Similarly, in yeast, the toxicity of Huntington exon 1 depends on the [RNQ ϩ ] prion state (16,22). This concept may even extend to the heterokaryon incompatibility mediated by the [Het-s] prion in Podospora anserina (34).…”

Biochemical, Cell Biological, and Genetic Assays to Analyze Amyloid and Prion Aggregation in Yeast

“…ThT has an emission spectrum that is red-shifted upon amyloid-binding, therefore allowing co-localization with aggregation-prone Interestingly, toxic and non-toxic aggregates of glutamine-expanded huntingtin have distinct subcellular aggregation patterns. Toxic huntingtin forms multiple punctuate foci, whereas the non-toxic structural variant is present in a single cytosolic focus (Duennwald et al 2006a;Duennwald et al 2006b). …”

“…Diverse yeast expression vectors are now available for the tagging of aggregation-prone proteins with FPs, most of which are suitable for determining the aggregation propensities of a protein (Krobitsch and Lindquist 2000;Duennwald et al 2006a;Alberti et al 2007;Alberti et al 2009). The choice of the promoter and the copy number of the yeast plasmid are also important parameters that need to be considered carefully.…”

“…Rnq1, however, not only induces benign conformational transitions of other prion proteins, it can also induce toxic conformational changes of glutamine-expansion (polyQ) proteins that cause Huntington's disease and several other late-onset neurodegenerative disorders. When polyQ fragments are expressed in yeast, they create a polyQ length-dependent toxicity that is accompanied by the formation of visible amyloid-containing aggregates in a [RNQ+]-dependent manner (Krobitsch and Lindquist 2000;Duennwald et al 2006a). Yeast prions have been widely used as model systems for the study of protein aggregation in vivo.…”