Letter | Published:

Mutant huntingtin binds the mitochondrial fission GTPase dynamin-related protein-1 and increases its enzymatic activity

Nature Medicine volume 17, pages 377382 (2011) | Download Citation

Abstract

Huntington's disease is an inherited and incurable neurodegenerative disorder caused by an abnormal polyglutamine (polyQ) expansion in huntingtin (encoded by HTT). PolyQ length determines disease onset and severity, with a longer expansion causing earlier onset. The mechanisms of mutant huntingtin-mediated neurotoxicity remain unclear; however, mitochondrial dysfunction is a key event in Huntington's disease pathogenesis1,2. Here we tested whether mutant huntingtin impairs the mitochondrial fission-fusion balance and thereby causes neuronal injury. We show that mutant huntingtin triggers mitochondrial fragmentation in rat neurons and fibroblasts of individuals with Huntington's disease in vitro and in a mouse model of Huntington's disease in vivo before the presence of neurological deficits and huntingtin aggregates. Mutant huntingtin abnormally interacts with the mitochondrial fission GTPase dynamin-related protein-1 (DRP1) in mice and humans with Huntington's disease, which, in turn, stimulates its enzymatic activity. Mutant huntingtin–mediated mitochondrial fragmentation, defects in anterograde and retrograde mitochondrial transport and neuronal cell death are all rescued by reducing DRP1 GTPase activity with the dominant-negative DRP1 K38A mutant. Thus, DRP1 might represent a new therapeutic target to combat neurodegeneration in Huntington's disease.

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Acknowledgements

We thank S. Finkbeiner (University of California–San Francisco) for the pGW1 plasmids encoding huntingtinex1-Q17-GFP, huntingtinex1-Q46-GFP, huntingtinex1-Q97-GFP; L. Thompson (University of California–Irvine) for the huntingtin plasmids pcDNA3.1-Q25-GFP and pcDNA3.1-Q97-GFP; U. Hartl (Max Planck Institute of Biochemistry) for the GST-huntingtinex1-Q20 and -Q53 constructs; A. van der Bliek (University of California–Los Angeles) for the DRP1 K38A cDNA in baculovirus expression vector (US National Center for Biotechnology Information accession number NM_005690.3); R. Youle (US National Institutes of Health (NIH)) for the YFP-DRP1 plasmid; R. Slack (University of Ottawa) for the MFN2 RasG12V (p82-FzoRV12pECFP-C1) expression plasmid; S. Strack (University of Iowa) for his pcDNA3.1β-DRP1shRNA vector; S. Lubitz, J. Johnson, V. DeAssis, C. Eldon and B. Kincaid for technical assistance; and A. Knott for manuscript development and editing. This work is supported by NIH grants to E.B.-W. (R01EY016164 and R01NS055193), a fellowship from the Hereditary Disease Foundation (to G.L.), grants to I.R., M.A.P. and M.R.H. from the Canadian Institutes of Health Research, and support from CHDI to M.R.H. The electron microscope tomography work was carried out in facilities of the US National Center for Microscopy and Imaging Research, supported by NIH grant P41RR004050 awarded to M.E.

Author information

Affiliations

  1. Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida, USA.

    • Wenjun Song
    • , Jin Chen
    • , Alejandra Petrilli
    • , Geraldine Liot
    • , Yue Zhou
    • , Blaise Bossy
    •  & Ella Bossy-Wetzel
  2. Structural Biology Group, Department of Molecular Biology, University of Salzburg, Salzburg, Austria.

    • Eva Klinglmayr
    •  & Robert Schwarzenbacher
  3. National Center for Microscopy and Imaging Research, University of California–San Diego, San Diego, California, USA.

    • Patrick Poquiz
    • , Jonathan Tjong
    • , Mark Ellisman
    •  & Guy Perkins
  4. Centre for Molecular Medicine and Therapeutics, University of British Columbia, Vancouver, British Columbia, Canada.

    • Mahmoud A Pouladi
    •  & Michael R Hayden
  5. University of California–San Diego, La Jolla, California, USA.

    • Eliezer Masliah
  6. Department of Anatomy and Cell Biology, McGill University, Montreal, Québec, Canada.

    • Isabelle Rouiller

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Contributions

W.S. performed all imaging and participated in the mitochondrial fragmentation and cell death analyses. J.C. performed the GTPase assays, some of the immune precipitations and the electron microscopy analysis. A.P. performed some of the neuronal cell death and immune precipitations. G.L. performed the electron microscopy stereology and generated some of the preliminary data. E.K. purified, cloned and prepared the recombinant DRP1 protein. Y.Z. performed western blotting for the DRP1 knockdown. P.P. and J.T. participated in the electron microscope tomography. M.A.P. and M.R.H. provided the YAC18 and YAC128 mice and advice on huntingtin coimmunoprecipitations. E.M. provided human postmortem brain samples. R.S. led the DRP1 protein purification. M.E. and G.P. led the electron microscope tomography experiment. B.B. performed GTPase assays, prepared samples for electron microscopy and purified the huntingtin protein. I.R. performed electron microscope negative-stain experiments. E.B.-W. conceived the project and wrote the article. All authors participated in the data analysis and interpretation of the results.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Ella Bossy-Wetzel.

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–10, Supplementary Table 1 and Supplementary Methods

Videos

  1. 1.

    Supplementary Video 1

    Mitochondrial movement in a neuron expressing HTT exon1-Q17-GFP and DsRed2-Mito. Movie corresponds to the kymograph in Figure 1f, top panel and shows mitochondrial transport. The movie lasts 5 min and is played back accelerated (original: 5 s frame−1, playback: 1/6 s frame−1).

  2. 2.

    Supplementary Video 2

    Mitochondrial movement in a neuron expressing HTT exon1-Q46-GFP and DsRed2-Mito. Movie corresponds to the kymograph in Figure 1f, center panel and shows a clear decrease in mitochondrial transport. The movie lasts 5 min and is played back accelerated (original: 5 s frame−1, playback: 1/6 s frame−1).

  3. 3.

    Supplementary Video 3

    Mitochondrial movement in a neuron expressing HTT exon1-Q97-GFP and DsRed2-Mito. Movie corresponds to the kymograph in Figure 1f, bottom panel and shows more pronounced arrest in mitochondrial transport. The movie lasts 5 min and is played back accelerated (original: 5 s frame−1, playback: 1/6 s frame−1).

  4. 4.

    Supplementary Video 4

    Electron tomography of a control mitochondrion in a medium spiny neuron. Movie showing the three-dimensional details of a mitochondrion in a medium spiny neuron reconstructed using electron tomography. These mitochondria are typically elongated along the direction of the axonal long axis. Clip 1: a rapid sequence through 190 slices (2.2 nm slice−1) of the tomographic volume that shows nearly the entire mitochondrial volume. There are 84 cristae. Clip 2: rotations and zooms of the surface-rendered volume after segmentation of the inner and outer membranes. The blue outer membrane is translucent to visualize the cristae displayed in various colors. Clip 3: rotation of the cristae after removal of the outer membrane to better distinguish the variety of shapes and sizes.

  5. 5.

    Supplementary Video 5

    Electron tomography of a fissioning YAC128 mitochondrion in a medium spiny neuron. Movie showing the three-dimensional details of a mitochondrion fissioning into three parts in a medium spiny neuron reconstructed using electron tomography. Clip 1: a rapid sequence through 210 slices (2.2 nm slice−1) of the tomographic volume. There are 223 cristae, many of which are small. Clip 2: rotation showing the outer membrane and the widths of the two constriction sites. Clip 3: rotations showing the cristae in each of the three parts. Clip 4: rotations and zooms highlighting the cristae and the constriction sites. The blue outer membrane is translucent to visualize the cristae displayed in various colors.

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DOI

https://doi.org/10.1038/nm.2313

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