Cytosolic chaperonin prevents polyglutamine toxicity with altering the aggregation state

Polyglutamine (polyQ)-expansion proteins cause neurodegenerative disorders including Huntington's disease, Kennedy's disease and various ataxias. The cytotoxicity of these proteins is associated with the formation of aggregates or other conformationally toxic species. Here, we show that the cytosolic chaperonin CCT (also known as TRiC) can alter the course of aggregation and cytotoxicity of huntingtin (Htt)–polyQ proteins in mammalian cells. Disruption of the CCT complex by RNAi-mediated knockdown enhanced Htt–polyQ aggregate formation and cellular toxicity. Analysis of the aggregation states of the Htt–polyQ proteins by fluorescence correlation spectroscopy revealed that CCT depletion results in the appearance of soluble Htt–polyQ aggregates. Similarly, overexpression of all eight subunits of CCT suppressed Htt aggregation and neuronal cell death. These results indicate that CCT has an essential role in protecting against the cytotoxicity of polyQ proteins by affecting the course of aggregation.

[1]  Mark Turmaine,et al.  Formation of Neuronal Intranuclear Inclusions Underlies the Neurological Dysfunction in Mice Transgenic for the HD Mutation , 1997, Cell.

[2]  Ulrike M K Böttcher,et al.  TRiC/CCT cooperates with different upstream chaperones in the folding of distinct protein classes , 2003, The EMBO journal.

[3]  J. Frydman,et al.  Closing the Folding Chamber of the Eukaryotic Chaperonin Requires the Transition State of ATP Hydrolysis , 2003, Cell.

[4]  P. Muchowski,et al.  Modulation of neurodegeneration by molecular chaperones , 2005, Nature Reviews Neuroscience.

[5]  Hans Lehrach,et al.  Huntingtin-Encoded Polyglutamine Expansions Form Amyloid-like Protein Aggregates In Vitro and In Vivo , 1997, Cell.

[6]  Christopher A Ross,et al.  Polyglutamine Pathogenesis Emergence of Unifying Mechanisms for Huntington's Disease and Related Disorders , 2002, Neuron.

[7]  J. Frydman,et al.  Tumorigenic mutations in VHL disrupt folding in vivo by interfering with chaperonin binding. , 2003, Molecular cell.

[8]  Richard I. Morimoto,et al.  Progressive Disruption of Cellular Protein Folding in Models of Polyglutamine Diseases , 2006, Science.

[9]  Jason C. Young,et al.  Pathways of chaperone-mediated protein folding in the cytosol , 2004, Nature Reviews Molecular Cell Biology.

[10]  K. Nagata,et al.  Cytosolic chaperonin protects folding intermediates of Gβ from aggregation by recognizing hydrophobic β-strands , 2006 .

[11]  F. Hartl,et al.  Principles of Chaperone-Assisted Protein Folding: Differences Between in Vitro and in Vivo Mechanisms , 1996, Science.

[12]  T. Misteli,et al.  High mobility of proteins in the mammalian cell nucleus , 2000, Nature.

[13]  H. Yanagi,et al.  Cytosolic Chaperonin Is Up-regulated during Cell Growth , 1999, The Journal of Biological Chemistry.

[14]  R. Rigler,et al.  Fluorescence correlation spectroscopy with high count rate and low background: analysis of translational diffusion , 1993, European Biophysics Journal.

[15]  Andreas Matouschek,et al.  Inefficient degradation of truncated polyglutamine proteins by the proteasome , 2004, The EMBO journal.

[16]  Hiroshi Kimura,et al.  Kinetics of Core Histones in Living Human Cells , 2001, The Journal of cell biology.

[17]  E. Wanker,et al.  Hsp70 and hsp40 chaperones can inhibit self-assembly of polyglutamine proteins into amyloid-like fibrils. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[18]  Alejandro Chavez,et al.  Genome-wide RNA interference screen identifies previously undescribed regulators of polyglutamine aggregation. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[19]  Huda Y. Zoghbi,et al.  Diseases of Unstable Repeat Expansion: Mechanisms and Common Principles , 2005, Nature Reviews Genetics.

[20]  J Martín-Benito,et al.  Eukaryotic chaperonin CCT stabilizes actin and tubulin folding intermediates in open quasi‐native conformations , 2000, The EMBO journal.

[21]  Richard I. Morimoto,et al.  Polyglutamine protein aggregates are dynamic , 2002, Nature Cell Biology.

[22]  Peter Breuer,et al.  Cellular toxicity of polyglutamine expansion proteins: mechanism of transcription factor deactivation. , 2004, Molecular cell.

[23]  Arthur L Horwich,et al.  Chaperonin-mediated protein folding: fate of substrate polypeptide , 2003, Quarterly Reviews of Biophysics.

[24]  R. Carraway,et al.  Huntingtin is a cytoplasmic protein associated with vesicles in human and rat brain neurons , 1995, Neuron.

[25]  Judith Frydman,et al.  Mechanism of the eukaryotic chaperonin: protein folding in the chamber of secrets. , 2004, Trends in cell biology.

[26]  A. Ashworth,et al.  Identification of six Tcp-1-related genes encoding divergent subunits of the TCP-1-containing chaperonin , 1994, Current Biology.

[27]  N. Hirokawa,et al.  Oligomeric Tubulin in Large Transporting Complex Is Transported via Kinesin in Squid Giant Axons , 2000, Cell.

[28]  R. Tsien,et al.  A monomeric red fluorescent protein , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[29]  J. Lippincott-Schwartz,et al.  Studying protein dynamics in living cells , 2001, Nature Reviews Molecular Cell Biology.

[30]  R. Morimoto,et al.  Huntingtin and Mutant SOD1 Form Aggregate Structures with Distinct Molecular Properties in Human Cells* , 2006, Journal of Biological Chemistry.

[31]  G. Farr,et al.  Chaperonin-Mediated Folding in the Eukaryotic Cytosol Proceeds through Rounds of Release of Native and Nonnative Forms , 1997, Cell.

[32]  J T Finch,et al.  Amyloid fibers are water-filled nanotubes , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[33]  Manish S. Shah,et al.  A novel gene containing a trinucleotide repeat that is expanded and unstable on Huntington's disease chromosomes , 1993, Cell.