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Effects of Q/N-rich, polyQ, and non-polyQ amyloids on the de novo formation of the [PSI+] prion in yeast and aggregation of Sup35 in vitro.

ABSTRACT: Prions are infectious protein conformations that are generally ordered protein aggregates. In the absence of prions, newly synthesized molecules of these same proteins usually maintain a conventional soluble conformation. However, prions occasionally arise even without a homologous prion template. The conformational switch that results in the de novo appearance of yeast prions with glutamine/aspargine (Q/N)-rich prion domains (e.g., [PSI+]), is promoted by heterologous prions with a similar domain (e.g., [RNQ+], also known as [PIN+]), or by overexpression of proteins with prion-like Q-, N-, or Q/N-rich domains. This finding led to the hypothesis that aggregates of heterologous proteins provide an imperfect template on which the new prion is seeded. Indeed, we show that newly forming Sup35 and preexisting Rnq1 aggregates always colocalize when [PSI+] appearance is facilitated by the [RNQ+] prion, and that Rnq1 fibers enhance the in vitro formation of fibers by the prion domain of Sup35 (NM). The proteins do not however form mixed, interdigitated aggregates. We also demonstrate that aggregating variants of the polyQ-containing domain of huntingtin promote the de novo conversion of Sup35 into [PSI+]; whereas nonaggregating variants of huntingtin and aggregates of non-polyQ amyloidogenic proteins, transthyretin, alpha-synuclein, and synphilin do not. Furthermore, transthyretin and alpha-synuclein amyloids do not facilitate NM aggregation in vitro, even though in [PSI+] cells NM and transthyretin aggregates also occasionally colocalize. Our data, especially the in vitro reproduction of the highly specific heterologous seeding effect, provide strong support for the hypothesis of cross-seeding in the spontaneous initiation of prion states.

SUBMITTER: Derkatch IL

PROVIDER: S-EPMC516497 | biostudies-literature | 2004 Aug

REPOSITORIES: biostudies-literature

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Publications

Effects of Q/N-rich, polyQ, and non-polyQ amyloids on the de novo formation of the [PSI+] prion in yeast and aggregation of Sup35 in vitro.

Derkatch Irina L IL Uptain Susan M SM Outeiro Tiago F TF Krishnan Rajaraman R Lindquist Susan L SL Liebman Susan W SW

Proceedings of the National Academy of Sciences of the United States of America 20040823 35

Prions are infectious protein conformations that are generally ordered protein aggregates. In the absence of prions, newly synthesized molecules of these same proteins usually maintain a conventional soluble conformation. However, prions occasionally arise even without a homologous prion template. The conformational switch that results in the de novo appearance of yeast prions with glutamine/aspargine (Q/N)-rich prion domains (e.g., [PSI+]), is promoted by heterologous prions with a similar doma ...[more]

PMID: 15326312

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Project description:Interactions amongst different amyloid proteins have been proposed as a probable mechanism of aggregation and thus an important risk factor for the onset as well as progression of various neurodegenerative disorders including Alzheimer's, Parkinson's, Huntington's, and Amyotrophic Lateral Sclerosis. Evidences suggest that transthyretin (TTR), a plasma protein associated with transthyretin amyloidosis or familial polyneuropathy (FAP) interacts with heterologous amyloid proteins including amyloid beta and islet amyloid polypeptide. In addition, recent clinical studies have revealed the presence of systemic polyneuropathy associated with FAP mutations in patients with spinocerebral ataxia, amyotrophic lateral sclerosis, and new familial systematic prion disease. Hence, it is important to investigate the interactions amongst different amyloid proteins to gain better insight into the pathology of amyloid disorders. Yeast has been an excellent model system to study interaction/ cross-seeding between heterologous amyloid proteins, more because of presence of endogenous yeast prions. Here, we examined interactions of non-glutamine (non-Q)-rich transthyretin, with glutamine (Q)-rich yeast prion protein Sup35. We established aggregation of an engineered double (F87M/L110M) mutant M-TTR-GFP in yeast. This mutant is monomeric and readily formed aggregates compared to WT-TTR-GFP in yeast at acidic pH. Interestingly, aggregation of M-TTR-GFP was significantly enhanced in presence of [PSI+], an endogenous prion form of Sup35. Different variants of [PSI+] seeded M-TTR-GFP with different efficiencies and curing of [PSI+] (losing the prion form) in these strains reduced aggregation. Moreover, overexpression of prion domain of Sup35 fused to RFP (NM-RFP) also increased M-TTR-GFP aggregation. M-TTR-GFP and NM-RFP aggregates co-localized in perivacuolar and juxtranuclear region. Sup35 protein was even immunocaptured in M-TTR-GFP aggregates. However, M-TTR-GFP overexpression did not induce Sup35 aggregation. Thus, it appears to be a unidirectional interaction between these two amyloid proteins. However, no affect on M-TTR-GFP aggregation was observed due to another yeast prion, [PIN+]. Our findings thus show the molecular interaction of transthyretin with yeast prion and support that sequence similarity is not the prime requirement for heterologous amyloid interactions.

Dataset Information

Effects of Q/N-rich, polyQ, and non-polyQ amyloids on the de novo formation of the [PSI+] prion in yeast and aggregation of Sup35 in vitro.

Publications

Effects of Q/N-rich, polyQ, and non-polyQ amyloids on the de novo formation of the [PSI+] prion in yeast and aggregation of Sup35 in vitro.

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