Insoluble detergent-resistant aggregates form between pathological and nonpathological lengths of polyglutamine in mammalian cells.
Pathological degeneration of neurons in Huntington's disease and associated neurodegenerative disorders is directly correlated with the expansion of CAG repeats encoding polyglutamines of extended length. The physical properties of extended polyglutamines and the intracellular consequences of expression of polyglutamine expansion have been the object of intensive investigation. We have extended the range of lengths of polyglutamine produced by recombinant DNA methodology by constructing a library of CAG/CAA repeats coding for a range of 25-300 glutamine residues. We have investigated the subcellular localization, interaction with other polyglutamine-containing polypeptides, and the physical properties of aggregated forms of polyglutamine in the cell. Extended polyQ aggregated in the cytoplasm and was only transported to the nucleus when a strong nuclear localization signal was present. Polyglutamine below pathological lengths could be captured in aggregates and transported to ectopic cell locations. The CREB-binding protein (CBP), containing a homopolymeric stretch of 19 glutamines, was likewise found to coaggregate in a polyglutamine-dependent manner, suggesting that pathology in polyglutamine disease may result from cellular depletion of normal proteins containing polyglutamine. We have observed a striking detergent resistance in aggregates produced from polyglutamine of pathological length. This observation has led to the development of a fluorescence-based assay exploiting the detergent resistance of polyglutamine aggregates that should facilitate high-throughput screening for agents that suppress polyglutamine aggregation in cells.
Kazantsev
A
A
Center for Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Preisinger
E
E
Dranovsky
A
A
Goldgaber
D
D
Housman
D
D
D000818:Animals; D019556:COS Cells; D050882:CREB-Binding Protein; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000737:chemistry; D003902:Detergents / Q000494:pharmacology*; D006816:Huntington Disease / Q000235:genetics; D009687:Nuclear Proteins / Q000737:chemistry; D010455:Peptides / Q000737:chemistry* / Q000235:genetics; D011487:Protein Conformation; D011489:Protein Denaturation; D015534:Trans-Activators / Q000737:chemistry; D018911:Trinucleotide Repeats
D003902:Detergents; D009687:Nuclear Proteins; D010455:Peptides; D015534:Trans-Activators; C097188:polyglutamine; D050882:CREB-Binding Protein