Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3

Katherine Jane Robinson, Madelaine C. Tym, Alison Hogan, Maxinne Watchon, Kristy C. Yuan, Stuart K. Plenderleith, Emily K. Don, Angela S. Laird*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

1 Citation (Scopus)

Abstract

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.

Original languageEnglish
Article numberdmm049023
Pages (from-to)1-12
Number of pages12
JournalDMM Disease Models and Mechanisms
Volume14
Issue number10
DOIs
Publication statusPublished - 1 Oct 2021

Bibliographical note

Copyright 2021. Version archived for private and non-commercial use with the permission of the author/s and according to publisher conditions. For further rights please contact the publisher.

Keywords

  • Spinocerebellar ataxia-3
  • Machado Joseph disease
  • hereditary spinocerebellar ataxias
  • neurodegenerative disease
  • flow cytometry
  • proteinopathy
  • insoluble protein species
  • protein aggregates

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