Journal of Biological Chemistry

Journal of Biological Chemistry

Volume 293, Issue 40, 5 October 2018, Pages 15581-15593
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Molecular Bases of Disease
Heat shock promotes inclusion body formation of mutant huntingtin (mHtt) and alleviates mHtt-induced transcription factor dysfunction

https://doi.org/10.1074/jbc.RA118.002933Get rights and content
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PolyQ-expanded huntingtin (mHtt) variants form aggregates, termed inclusion bodies (IBs), in individuals with and models of Huntington's disease (HD). The role of IB versus diffusible mHtt in neurotoxicity remains unclear. Using a ponasterone (PA)-inducible cell model of HD, here we evaluated the effects of heat shock on the appearance and functional outcome of Htt103QExon1–EGFP expression. Quantitative image analysis indicated that 80–90% of this mHtt protein initially appears as “diffuse” signals in the cytosol, with IBs forming at high mHtt expression. A 2-h heat shock during the PA induction reduced the diffuse signal, but greatly increased mHtt IB formation in both cytosol and nucleus. Dose- and time-dependent mHtt expression suggested that nucleated polymerization drives IB formation. RNA-mediated knockdown of heat shock protein 70 (HSP70) and heat shock cognate 70 protein (HSC70) provided evidence for their involvement in promoting diffuse mHtt to form IBs. Reporter gene assays assessing the impacts of diffuse versus IB mHtt showed concordance of diffuse mHtt expression with the repression of heat shock factor 1, cAMP-responsive element-binding protein (CREB), and NF-κB activity. CREB repression was reversed by heat shock coinciding with mHtt IB formation. In an embryonic striatal neuron–derived HD model, the chemical chaperone sorbitol similarly promoted the structuring of diffuse mHtt into IBs and supported cell survival under stress. Our results provide evidence that mHtt IB formation is a chaperone-supported cellular coping mechanism that depletes diffusible mHtt conformers, alleviates transcription factor dysfunction, and promotes neuron survival.

Huntington's disease
aggregation
heat shock protein (HSP)
chaperone
heat shock factor protein 1 (HSF1)
neurodegeneration
HSP chaperones
heat shock
inclusion bodies
polyQ-mHtt protein
transcription factor sequestration
CREB

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This work was supported by internal University funding (to A. Y. C. L.), the Chemistry and Chemical Biology Fund 800177 of Rutgers (to K. Y. C.), and National Institutes of Health Grant R01 5DC15000 (to K. K.). M. P., H. S., and D. B. were supported by Aresty Research Fellowships from Rutgers University while working on this research project. The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

We dedicate this work to the memory of Dr. J Fred “Paulo” Dice. It was through his generosity and introduction that we secured the HSC70 antisense construct from Dr. Janice Blum's laboratory of Indiana University School of Medicine in 2006.

This article contains supporting Figs. S1–S6.

1

These authors contributed equally to this work.

2

Present address: Joint Dept. of Biomedical Engineering, University of North Carolina and North Carolina State University, Raleigh, NC 27695.

3

Present address: Lewis Katz School of Medicine, Temple University, Philadelphia, PA 19140.

4

Present address: Helen Wills Neuroscience Institute, University of California, Berkeley, 132 Barker Hall, Berkeley, CA 94720.

© 2018 Chen et al.