Calpain inhibition is protective in Machado–Joseph disease zebrafish due to induction of autophagy

Maxinne Watchon, Kristy C. Yuan, Nick Mackovski, Adam J. Svahn, Nicholas J. Cole, Claire Goldsbury, Silke Rinkwitz, Thomas S. Becker, Garth A. Nicholson, Angela S. Laird

Research output: Contribution to journalArticleResearchpeer-review

Abstract

The neurodegenerative disease Machado–Joseph disease (MJD), also known as spinocerebellar ataxin-3, affects neurons of the brain and spinal cord, disrupting control of the movement of muscles. We have successfully established the first transgenic zebrafish (Danio rerio) model of MJD by expressing human ataxin-3 protein containing either 23 glutamines (23Q, wild-type) or 84Q (MJD-causing) within neurons. Phenotypic characterization of the zebrafish (male and female) revealed that the ataxin-3-84Q zebrafish have decreased survival compared with ataxin-3-23Q and develop ataxin-3 neuropathology, ataxin-3 cleavage fragments and motor impairment. Ataxin-3-84Q zebrafish swim shorter distances than ataxin-3-23Q zebrafish as early as 6 days old, even if expression of the human ataxin-3 protein is limited to motor neurons. This swimming phenotype provides a valuable readout for drug treatment studies. Treating the EGFP-ataxin-3-84Q zebrafish with the calpain inhibitor compound calpeptin decreased levels of ataxin-3 cleavage fragments, but also removed all human ataxin-3 protein (confirmed by ELISA) and prevented the early MJD zebrafish motor phenotype. We identified that this clearance of ataxin-3 protein by calpeptin treatment resulted from an increase in autophagic flux (indicated by decreased p62 levels and increased LC3II). Cotreatment with the autophagy inhibitor chloroquine blocked the decrease in human ataxin-3 levels and the improved movement produced by calpeptin treatment. This study demonstrates that this first transgenic zebrafish model of MJD is a valuable tool for testing potential treatments for MJD. Calpeptin treatment is protective in this model of MJD and removal of human ataxin-3 through macro-autophagy plays an important role in this beneficial effect.

LanguageEnglish
Pages7782-7794
Number of pages13
JournalJournal of Neuroscience
Volume37
Issue number32
DOIs
Publication statusPublished - 2017

Fingerprint

Machado-Joseph Disease
Calpain
Autophagy
Zebrafish
Ataxin-3
Phenotype
Therapeutics
Neurons
Chloroquine
Motor Neurons
Glutamine
Neurodegenerative Diseases

Keywords

  • autophagy
  • calpain
  • Joseph disease
  • machado
  • neurodegeneration
  • spinocerebellar ataxia-3
  • zebrafish

Cite this

Watchon, Maxinne ; Yuan, Kristy C. ; Mackovski, Nick ; Svahn, Adam J. ; Cole, Nicholas J. ; Goldsbury, Claire ; Rinkwitz, Silke ; Becker, Thomas S. ; Nicholson, Garth A. ; Laird, Angela S. / Calpain inhibition is protective in Machado–Joseph disease zebrafish due to induction of autophagy. In: Journal of Neuroscience. 2017 ; Vol. 37, No. 32. pp. 7782-7794.
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abstract = "The neurodegenerative disease Machado–Joseph disease (MJD), also known as spinocerebellar ataxin-3, affects neurons of the brain and spinal cord, disrupting control of the movement of muscles. We have successfully established the first transgenic zebrafish (Danio rerio) model of MJD by expressing human ataxin-3 protein containing either 23 glutamines (23Q, wild-type) or 84Q (MJD-causing) within neurons. Phenotypic characterization of the zebrafish (male and female) revealed that the ataxin-3-84Q zebrafish have decreased survival compared with ataxin-3-23Q and develop ataxin-3 neuropathology, ataxin-3 cleavage fragments and motor impairment. Ataxin-3-84Q zebrafish swim shorter distances than ataxin-3-23Q zebrafish as early as 6 days old, even if expression of the human ataxin-3 protein is limited to motor neurons. This swimming phenotype provides a valuable readout for drug treatment studies. Treating the EGFP-ataxin-3-84Q zebrafish with the calpain inhibitor compound calpeptin decreased levels of ataxin-3 cleavage fragments, but also removed all human ataxin-3 protein (confirmed by ELISA) and prevented the early MJD zebrafish motor phenotype. We identified that this clearance of ataxin-3 protein by calpeptin treatment resulted from an increase in autophagic flux (indicated by decreased p62 levels and increased LC3II). Cotreatment with the autophagy inhibitor chloroquine blocked the decrease in human ataxin-3 levels and the improved movement produced by calpeptin treatment. This study demonstrates that this first transgenic zebrafish model of MJD is a valuable tool for testing potential treatments for MJD. Calpeptin treatment is protective in this model of MJD and removal of human ataxin-3 through macro-autophagy plays an important role in this beneficial effect.",
keywords = "autophagy, calpain, Joseph disease, machado, neurodegeneration, spinocerebellar ataxia-3, zebrafish",
author = "Maxinne Watchon and Yuan, {Kristy C.} and Nick Mackovski and Svahn, {Adam J.} and Cole, {Nicholas J.} and Claire Goldsbury and Silke Rinkwitz and Becker, {Thomas S.} and Nicholson, {Garth A.} and Laird, {Angela S.}",
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Watchon, M, Yuan, KC, Mackovski, N, Svahn, AJ, Cole, NJ, Goldsbury, C, Rinkwitz, S, Becker, TS, Nicholson, GA & Laird, AS 2017, 'Calpain inhibition is protective in Machado–Joseph disease zebrafish due to induction of autophagy', Journal of Neuroscience, vol. 37, no. 32, pp. 7782-7794. https://doi.org/10.1523/JNEUROSCI.1142-17.2017

Calpain inhibition is protective in Machado–Joseph disease zebrafish due to induction of autophagy. / Watchon, Maxinne; Yuan, Kristy C.; Mackovski, Nick; Svahn, Adam J.; Cole, Nicholas J.; Goldsbury, Claire; Rinkwitz, Silke; Becker, Thomas S.; Nicholson, Garth A.; Laird, Angela S.

In: Journal of Neuroscience, Vol. 37, No. 32, 2017, p. 7782-7794.

Research output: Contribution to journalArticleResearchpeer-review

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T1 - Calpain inhibition is protective in Machado–Joseph disease zebrafish due to induction of autophagy

AU - Watchon, Maxinne

AU - Yuan, Kristy C.

AU - Mackovski, Nick

AU - Svahn, Adam J.

AU - Cole, Nicholas J.

AU - Goldsbury, Claire

AU - Rinkwitz, Silke

AU - Becker, Thomas S.

AU - Nicholson, Garth A.

AU - Laird, Angela S.

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N2 - The neurodegenerative disease Machado–Joseph disease (MJD), also known as spinocerebellar ataxin-3, affects neurons of the brain and spinal cord, disrupting control of the movement of muscles. We have successfully established the first transgenic zebrafish (Danio rerio) model of MJD by expressing human ataxin-3 protein containing either 23 glutamines (23Q, wild-type) or 84Q (MJD-causing) within neurons. Phenotypic characterization of the zebrafish (male and female) revealed that the ataxin-3-84Q zebrafish have decreased survival compared with ataxin-3-23Q and develop ataxin-3 neuropathology, ataxin-3 cleavage fragments and motor impairment. Ataxin-3-84Q zebrafish swim shorter distances than ataxin-3-23Q zebrafish as early as 6 days old, even if expression of the human ataxin-3 protein is limited to motor neurons. This swimming phenotype provides a valuable readout for drug treatment studies. Treating the EGFP-ataxin-3-84Q zebrafish with the calpain inhibitor compound calpeptin decreased levels of ataxin-3 cleavage fragments, but also removed all human ataxin-3 protein (confirmed by ELISA) and prevented the early MJD zebrafish motor phenotype. We identified that this clearance of ataxin-3 protein by calpeptin treatment resulted from an increase in autophagic flux (indicated by decreased p62 levels and increased LC3II). Cotreatment with the autophagy inhibitor chloroquine blocked the decrease in human ataxin-3 levels and the improved movement produced by calpeptin treatment. This study demonstrates that this first transgenic zebrafish model of MJD is a valuable tool for testing potential treatments for MJD. Calpeptin treatment is protective in this model of MJD and removal of human ataxin-3 through macro-autophagy plays an important role in this beneficial effect.

AB - The neurodegenerative disease Machado–Joseph disease (MJD), also known as spinocerebellar ataxin-3, affects neurons of the brain and spinal cord, disrupting control of the movement of muscles. We have successfully established the first transgenic zebrafish (Danio rerio) model of MJD by expressing human ataxin-3 protein containing either 23 glutamines (23Q, wild-type) or 84Q (MJD-causing) within neurons. Phenotypic characterization of the zebrafish (male and female) revealed that the ataxin-3-84Q zebrafish have decreased survival compared with ataxin-3-23Q and develop ataxin-3 neuropathology, ataxin-3 cleavage fragments and motor impairment. Ataxin-3-84Q zebrafish swim shorter distances than ataxin-3-23Q zebrafish as early as 6 days old, even if expression of the human ataxin-3 protein is limited to motor neurons. This swimming phenotype provides a valuable readout for drug treatment studies. Treating the EGFP-ataxin-3-84Q zebrafish with the calpain inhibitor compound calpeptin decreased levels of ataxin-3 cleavage fragments, but also removed all human ataxin-3 protein (confirmed by ELISA) and prevented the early MJD zebrafish motor phenotype. We identified that this clearance of ataxin-3 protein by calpeptin treatment resulted from an increase in autophagic flux (indicated by decreased p62 levels and increased LC3II). Cotreatment with the autophagy inhibitor chloroquine blocked the decrease in human ataxin-3 levels and the improved movement produced by calpeptin treatment. This study demonstrates that this first transgenic zebrafish model of MJD is a valuable tool for testing potential treatments for MJD. Calpeptin treatment is protective in this model of MJD and removal of human ataxin-3 through macro-autophagy plays an important role in this beneficial effect.

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KW - calpain

KW - Joseph disease

KW - machado

KW - neurodegeneration

KW - spinocerebellar ataxia-3

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