Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3
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 aggre...
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| Format: | Article |
| Language: | English |
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The Company of Biologists
2021-10-01
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| Series: | Disease Models & Mechanisms |
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| Online Access: | http://dmm.biologists.org/content/14/10/dmm049023 |
| _version_ | 1819172639386632192 |
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| author | Katherine J. Robinson Madelaine C. Tym Alison Hogan Maxinne Watchon Kristy C. Yuan Stuart K. Plenderleith Emily K. Don Angela S. Laird |
| author_facet | Katherine J. Robinson Madelaine C. Tym Alison Hogan Maxinne Watchon Kristy C. Yuan Stuart K. Plenderleith Emily K. Don Angela S. Laird |
| author_sort | Katherine J. Robinson |
| collection | DOAJ |
| description | 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. |
| first_indexed | 2024-12-22T20:10:23Z |
| format | Article |
| id | doaj.art-58a28b3fec1546bcae968ee506d3ac17 |
| institution | Directory Open Access Journal |
| issn | 1754-8403 1754-8411 |
| language | English |
| last_indexed | 2024-12-22T20:10:23Z |
| publishDate | 2021-10-01 |
| publisher | The Company of Biologists |
| record_format | Article |
| series | Disease Models & Mechanisms |
| spelling | doaj.art-58a28b3fec1546bcae968ee506d3ac172022-12-21T18:14:03ZengThe Company of BiologistsDisease Models & Mechanisms1754-84031754-84112021-10-01141010.1242/dmm.049023049023Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3Katherine J. Robinson0Madelaine C. Tym1Alison Hogan2Maxinne Watchon3Kristy C. Yuan4Stuart K. Plenderleith5Emily K. Don6Angela S. Laird7 Centre for Motor Neuron Disease Research, Department of Biomedical Sciences, Faculty of Medicine, Health and Human Sciences, Macquarie University, Sydney, NSW 2109, Australia Centre for Motor Neuron Disease Research, Department of Biomedical Sciences, Faculty of Medicine, Health and Human Sciences, Macquarie University, Sydney, NSW 2109, Australia Centre for Motor Neuron Disease Research, Department of Biomedical Sciences, Faculty of Medicine, Health and Human Sciences, Macquarie University, Sydney, NSW 2109, Australia Centre for Motor Neuron Disease Research, Department of Biomedical Sciences, Faculty of Medicine, Health and Human Sciences, Macquarie University, Sydney, NSW 2109, Australia Centre for Motor Neuron Disease Research, Department of Biomedical Sciences, Faculty of Medicine, Health and Human Sciences, Macquarie University, Sydney, NSW 2109, Australia Centre for Motor Neuron Disease Research, Department of Biomedical Sciences, Faculty of Medicine, Health and Human Sciences, Macquarie University, Sydney, NSW 2109, Australia Centre for Motor Neuron Disease Research, Department of Biomedical Sciences, Faculty of Medicine, Health and Human Sciences, Macquarie University, Sydney, NSW 2109, Australia Centre for Motor Neuron Disease Research, Department of Biomedical Sciences, Faculty of Medicine, Health and Human Sciences, Macquarie University, Sydney, NSW 2109, Australia 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.http://dmm.biologists.org/content/14/10/dmm049023spinocerebellar ataxia 3machado–joseph diseasehereditary spinocerebellar ataxiasneurodegenerative diseaseflow cytometryproteinopathyinsoluble protein speciesprotein aggregates |
| spellingShingle | Katherine J. Robinson Madelaine C. Tym Alison Hogan Maxinne Watchon Kristy C. Yuan Stuart K. Plenderleith Emily K. Don Angela S. Laird Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3 Disease Models & Mechanisms spinocerebellar ataxia 3 machado–joseph disease hereditary spinocerebellar ataxias neurodegenerative disease flow cytometry proteinopathy insoluble protein species protein aggregates |
| title | Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3 |
| title_full | Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3 |
| title_fullStr | Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3 |
| title_full_unstemmed | Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3 |
| title_short | Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3 |
| title_sort | flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3 |
| topic | spinocerebellar ataxia 3 machado–joseph disease hereditary spinocerebellar ataxias neurodegenerative disease flow cytometry proteinopathy insoluble protein species protein aggregates |
| url | http://dmm.biologists.org/content/14/10/dmm049023 |
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