Molecular characterization of an aggregation-prone variant of alpha-synuclein used to model synucleinopathies

Masaracchia, Caterina; König, Anika; Valiente-Gabioud, Ariel A.; Peralta, Pablo Ospina; Favretto, Filippo; Strohäker, Timo; Lázaro, Diana F.; Zweckstetter, Markus; Fernández, Claudio O.; Outeiro, Tiago F.

doi:10.1016/j.bbapap.2019.140298

Cited by 12 publications

(8 citation statements)

References 60 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…4 B). Whereas these values are lower than typical ratios seen in the rigid elements of folded proteins (0.7–0.8), they are significantly higher than those observed in some other IDPs such as α-synuclein (Masaracchia et al 2020 ).…”

Section: Resultsmentioning

confidence: 56%

Partial structure, dampened mobility, and modest impact of a His tag in the SARS-CoV-2 Nsp2 C-terminal region

2021

View full text Add to dashboard Cite

Intrinsically disordered proteins (IDPs) play essential roles in regulating physiological processes in eukaryotic cells. Many viruses use their own IDPs to “hack” these processes to deactivate host defenses and promote viral growth. Thus, viral IDPs are attractive drug targets. While IDPs are hard to study by X-ray crystallography or cryo-EM, atomic level information on their conformational preferences and dynamics can be obtained using NMR spectroscopy. SARS-CoV-2 Nsp2, whose C-terminal region (CtR) is predicted to be disordered, interacts with human proteins that regulate translation initiation and endosome vesicle sorting. Molecules that block these interactions could be valuable leads for drug development. The 13Cβ and backbone 13CO, 1HN, 13Cα, and 15N nuclei of Nsp2’s 45-residue CtR were assigned and used to characterize its structure and dynamics in three contexts; namely: (1) retaining an N-terminal His tag, (2) without the His tag and with an adventitious internal cleavage, and (3) lacking both the His tag and the internal cleavage. Two five-residue segments adopting a minor extended population were identified. Overall, the dynamic behavior is midway between a completely rigid and a fully flexible chain. Whereas the presence of an N-terminal His tag and internal cleavage stiffen and loosen, respectively, neighboring residues, they do not affect the tendency of two regions to populate extended conformations.

show abstract

Section: Resultsmentioning

confidence: 56%

Partial structure, dampened mobility, and modest impact of a His tag in the SARS-CoV-2 Nsp2 C-terminal region

2021

View full text Add to dashboard Cite

show abstract

“…In order to assess whether Hsp27 modulated the MGO‐induced aggregation of aSyn, we used an established cell model using an aggregation‐prone variant of aSyn (known as SynT) 19,64,66‐68 . This model consists of a modified form of aSyn where the C‐terminus is fused to a truncated, nonfluorescent fragment of GFP (known as SynT) 36,69 . Using this model, we have previously shown that MGO treatment of cells expressing SynT increases the percentage of cells with aSyn inclusions (51% to 85%) 19 .…”

Section: Resultsmentioning

confidence: 99%

Hsp27 reduces glycation‐induced toxicity and aggregation of alpha‐synuclein

et al. 2020

View full text Add to dashboard Cite

This is an open access article under the terms of the Creat ive Commo ns Attri butio n-NonCo mmercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.Abstract α-synuclein (aSyn) is a major player in Parkinson's disease and a group of other disorders collectively known as synucleinopathies, but the precise molecular mechanisms involved are still unclear. aSyn, as virtually all proteins, undergoes a series of posttranslational modifications during its lifetime, which can affect its biology and pathobiology. We recently showed that glycation of aSyn by methylglyoxal (MGO) potentiates its oligomerization and toxicity, induces dopaminergic neuronal cell loss in mice, and affects motor performance in flies. Small heat-shock proteins (sHsps) are molecular chaperones that facilitate the folding of proteins or target misfolded proteins for clearance. Importantly, sHsps were shown to prevent aSyn aggregation and cytotoxicity.Upon treating cells with increasing amounts of methylglyoxal, we found that the levels of Hsp27 decreased in a dose-dependent manner. Therefore, we hypothesized that restoring the levels of Hsp27 in glycating environments could alleviate the pathogenicity of aSyn. Consistently, we found that Hsp27 reduced MGO-induced aSyn aggregation in cells, leading to the formation of nontoxic aSyn species. Remarkably, increasing the levels of Hsp27 suppressed the deleterious effects induced by MGO. Our findings suggest that in glycating environments, the levels of Hsp27 are important for modulating the glycation-associated cellular pathologies in synucleinopathies.

show abstract

“…In order to assess the effect of doxycycline treatment on aSyn aggregation in cell cultures, we used the SynT/Synphilin-1 model of inclusion formation (Lázaro et al, 2014; Masaracchia et al, 2020). SynT is a C-terminally modified variant of aSyn that is more prone to aggregation.…”

Section: Resultsmentioning

confidence: 99%

“…SynT is a C-terminally modified variant of aSyn that is more prone to aggregation. H4 cells were transiently transfected with constructs encoding for Synphilin-1 and SynT, as previously described (Lázaro et al, 2014), in order to induce the formation of LB-like inclusions (Supplementary figure S2A), as characterized by our and other groups (Engelender et al, 1999; Masaracchia et al, 2020; McLean et al, 2001). Seventeen hours after transfection, cells were treated with doxycycline at the following concentrations: 0.01 μM, 0.1 μM, 1 μM and 10 μM.…”

Section: Resultsmentioning

confidence: 99%

Doxycycline inhibits α-synuclein-associated pathologiesin vitroandin vivo

Dominguez-Meijide

Parrales

Vasili³

et al. 2020

Preprint

Self Cite

View full text Add to dashboard Cite

Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are neurodegenerative disorders characterized by the misfolding and aggregation of alpha-synuclein (aSyn). Doxycycline, a tetracyclic antibiotic shows neuroprotective effects, initially proposed to be due to its anti-inflammatory properties. More recently, an additional mechanism by which doxycycline may exert its neuroprotective effects has been proposed as it has been shown that it inhibits amyloid aggregation. Here, we studied the effects of doxycycline on aSyn aggregation in vivo, in vitro and in a cell free system using real-time quaking induced conversion (RT-QuiC). Our results show that doxycycline decreases the number and size of aSyn aggregates in cells. In addition, doxycycline inhibits the aggregation and seeding of recombinant aSyn, and attenuates the production of mitochondrial-derived reactive oxygen species. Finally, we found doxycycline induces a cellular redistribution of the aggregates in an animal model of PD that is associated with a recovery of dopaminergic function. In summary, we provide strong evidence that doxycycline treatment may be an effective strategy against synucleinopathies.

show abstract

Molecular characterization of an aggregation-prone variant of alpha-synuclein used to model synucleinopathies

Cited by 12 publications

References 60 publications

Partial structure, dampened mobility, and modest impact of a His tag in the SARS-CoV-2 Nsp2 C-terminal region

Partial structure, dampened mobility, and modest impact of a His tag in the SARS-CoV-2 Nsp2 C-terminal region

Hsp27 reduces glycation‐induced toxicity and aggregation of alpha‐synuclein

Doxycycline inhibits α-synuclein-associated pathologiesin vitroandin vivo

Contact Info

Product

Resources

About