PUBLICATION
Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3
- Authors
- Robinson, K.J., Tym, M.C., Hogan, A., Watchon, M., Yuan, K.C., Plenderleith, S.K., Don, E.K., Laird, A.S.
- ID
- ZDB-PUB-210903-6
- Date
- 2021
- Source
- Disease models & mechanisms 14(10): (Journal)
- Registered Authors
- Don, Emily, Laird, Angela, Watchon, Maxinne
- Keywords
- Flow cytometry, Hereditary spinocerebellar ataxias, Insoluble protein species, Machado Joseph disease, Neurodegenerative disease, Protein aggregates, Proteinopathy, Spinocerebellar ataxia-3
- MeSH Terms
-
- 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*
- PubMed
- 34473252 Full text @ Dis. Model. Mech.
Citation
Robinson, K.J., Tym, M.C., Hogan, A., Watchon, M., Yuan, K.C., Plenderleith, S.K., Don, E.K., Laird, A.S. (2021) Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3. Disease models & mechanisms. 14(10):.
Abstract
Spinocerebellar ataxia-3 (SCA3, also 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 (FloIT), allowing rapid quantification of detergent insoluble forms of ataxin-3 fused to a green fluorescent protein in the 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 the cells and zebrafish expressing polyQ expanded ataxin-3 compared to those expressing wildtype human ataxin-3. Treatment with compounds known to modulate autophagy activity was found to alter the number of detergent-insoluble ataxin-3 particles in cells and zebrafish. We conclude that flow cytometry can be harnessed to rapidly count ataxin-3 aggregates, both in vitro and in vivo, and can be utilised to compare potential therapies targeting protein aggregates.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping