Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3.
Animals
Animals, Genetically Modified
Ataxin-3
/ metabolism
Autophagy
/ drug effects
Cell Line, Tumor
Cell Nucleus
/ metabolism
Disease Models, Animal
Flow Cytometry
Green Fluorescent Proteins
/ metabolism
Humans
Machado-Joseph Disease
/ pathology
Neurons
/ metabolism
Peptides
Protein Aggregates
Solubility
Zebrafish
/ physiology
Flow cytometry
Hereditary spinocerebellar ataxias
Insoluble protein species
Machado–Joseph disease
Neurodegenerative disease
Protein aggregates
Proteinopathy
Spinocerebellar ataxia 3
Journal
Disease models & mechanisms
ISSN: 1754-8411
Titre abrégé: Dis Model Mech
Pays: England
ID NLM: 101483332
Informations de publication
Date de publication:
01 10 2021
01 10 2021
Historique:
received:
18
03
2021
accepted:
23
08
2021
pubmed:
3
9
2021
medline:
22
3
2022
entrez:
2
9
2021
Statut:
ppublish
Résumé
Spinocerebellar ataxia 3 (SCA3, also known as Machado-Joseph disease) is a neurodegenerative disease caused by inheritance of a CAG repeat expansion within the ATXN3 gene, resulting in polyglutamine (polyQ) repeat expansion within the ataxin-3 protein. In this study, we have identified protein aggregates in both neuronal-like (SHSY5Y) cells and transgenic zebrafish expressing human ataxin-3 with expanded polyQ. We have adapted a previously reported flow cytometry methodology named flow cytometric analysis of inclusions and trafficking, allowing rapid quantification of detergent insoluble forms of ataxin-3 fused to a GFP in SHSY5Y cells and cells dissociated from the zebrafish larvae. Flow cytometric analysis revealed an increased number of detergent-insoluble ataxin-3 particles per nuclei in cells and in zebrafish expressing polyQ-expanded ataxin-3 compared to those expressing wild-type human ataxin-3. Treatment with compounds known to modulate autophagic activity altered the number of detergent-insoluble ataxin-3 particles in cells and zebrafish expressing mutant human ataxin-3. We conclude that flow cytometry can be harnessed to rapidly count ataxin-3 aggregates, both in vitro and in vivo, and can be used to compare potential therapies targeting protein aggregates. This article has an associated First Person interview with the first author of the paper.
Identifiants
pubmed: 34473252
pii: 272020
doi: 10.1242/dmm.049023
pmc: PMC8524651
pii:
doi:
Substances chimiques
Peptides
0
Protein Aggregates
0
enhanced green fluorescent protein
0
Green Fluorescent Proteins
147336-22-9
polyglutamine
26700-71-0
Ataxin-3
EC 3.4.19.12
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Informations de copyright
© 2021. Published by The Company of Biologists Ltd.
Déclaration de conflit d'intérêts
Competing interests The authors declare no competing or financial interests.
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