Next Article in Journal
Changes in the Oxidation-Reduction State of Human Dermal Fibroblasts as an Effect of Lomefloxacin Phototoxic Action
Next Article in Special Issue
Microglial Depletion Has No Impact on Disease Progression in a Mouse Model of Machado–Joseph Disease
Previous Article in Journal
LTα, TNF, and ILC3 in Peyer’s Patch Organogenesis
Previous Article in Special Issue
Drosophila as a Model of Unconventional Translation in Spinocerebellar Ataxia Type 3
 
 
Article

A Robust Assay to Monitor Ataxin-3 Amyloid Fibril Assembly

1
Instituto de Investigação e Inovação em Saúde (i3S), Universidade do Porto, 4200-135 Porto, Portugal
2
Instituto de Biologia Molecular e Celular (IBMC), Universidade do Porto, 4200-135 Porto, Portugal
3
International Iberian Nanotechnology Laboratory (INL), 4715-330 Braga, Portugal
4
Instituto de Ciências Biomédicas Abel Salazar (ICBAS), Universidade do Porto, 4050-313 Porto, Portugal
5
Department of Biology, Faculty of Sciences, University of Porto, 4169-007 Porto, Portugal
6
Life and Health Sciences Research Institute (ICVS), School of Medicine, University of Minho, 4710-057 Braga, Portugal
7
ICVS/3B’s—PT Government Associate Laboratory, 4710-057 Braga, Portugal
8
Dynamic Biosensors GmbH, 82152 Martinsried, Germany
*
Authors to whom correspondence should be addressed.
Academic Editor: Takahiro Seki
Cells 2022, 11(12), 1969; https://doi.org/10.3390/cells11121969
Received: 10 May 2022 / Revised: 14 June 2022 / Accepted: 16 June 2022 / Published: 19 June 2022
Spinocerebellar ataxia type 3 (SCA3) is caused by the expansion of a glutamine repeat in the protein ataxin-3, which is deposited as intracellular aggregates in affected brain regions. Despite the controversial role of ataxin-3 amyloid structures in SCA3 pathology, the identification of molecules with the capacity to prevent aberrant self-assembly and stabilize functional conformation(s) of ataxin-3 is a key to the development of therapeutic solutions. Amyloid-specific kinetic assays are routinely used to measure rates of protein self-assembly in vitro and are employed during screening for fibrillation inhibitors. The high tendency of ataxin-3 to assemble into oligomeric structures implies that minor changes in experimental conditions can modify ataxin-3 amyloid assembly kinetics. Here, we determine the self-association rates of ataxin-3 and present a detailed study of the aggregation of normal and pathogenic ataxin-3, highlighting the experimental conditions that should be considered when implementing and validating ataxin-3 amyloid progress curves in different settings and in the presence of ataxin-3 interactors. This assay provides a unique and robust platform to screen for modulators of the first steps of ataxin-3 aggregation—a starting point for further studies with cell and animal models of SCA3. View Full-Text
Keywords: thioflavin-T; polyglutamine expansion; reproducibility; ubiquitin; self-association rates; equilibrium dissociation constant; switchSENSE thioflavin-T; polyglutamine expansion; reproducibility; ubiquitin; self-association rates; equilibrium dissociation constant; switchSENSE
Show Figures

Graphical abstract

MDPI and ACS Style

Figueiredo, F.; Lopes-Marques, M.; Almeida, B.; Matscheko, N.; Martins, P.M.; Silva, A.; Macedo-Ribeiro, S. A Robust Assay to Monitor Ataxin-3 Amyloid Fibril Assembly. Cells 2022, 11, 1969. https://doi.org/10.3390/cells11121969

AMA Style

Figueiredo F, Lopes-Marques M, Almeida B, Matscheko N, Martins PM, Silva A, Macedo-Ribeiro S. A Robust Assay to Monitor Ataxin-3 Amyloid Fibril Assembly. Cells. 2022; 11(12):1969. https://doi.org/10.3390/cells11121969

Chicago/Turabian Style

Figueiredo, Francisco, Mónica Lopes-Marques, Bruno Almeida, Nena Matscheko, Pedro M. Martins, Alexandra Silva, and Sandra Macedo-Ribeiro. 2022. "A Robust Assay to Monitor Ataxin-3 Amyloid Fibril Assembly" Cells 11, no. 12: 1969. https://doi.org/10.3390/cells11121969

Find Other Styles
Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.

Article Access Map by Country/Region

1
Back to TopTop