Aggregation of huntingtin in yeast varies with the length of the polyglutamine expansion and the expression of chaperone proteins.
Huntington's disease (HD) is an inherited neurodegenerative disorder caused by polyglutamine (polyQ) expansions in the huntingtin (Ht) protein. A hallmark of HD is the proteolytic production of an N-terminal fragment of Ht, containing the polyQ repeat, that forms aggregates in the nucleus and cytoplasm of affected neurons. Proteins with longer polyQ repeats aggregate more rapidly and cause disease at an earlier age, but the mechanism of aggregation and its relationship to disease remain unclear. To provide a new, genetically tractable model system for the study of Ht, we engineered yeast cells to express an N-terminal fragment of Ht with different polyQ repeat lengths of 25, 47, 72, or 103 residues, fused to green fluorescent protein. The extent of aggregation varied with the length of the polyQ repeat: at the two extremes, most HtQ103 protein coalesced into a single large cytoplasmic aggregate, whereas HtQ25 exhibited no sign of aggregation. Mutations that inhibit the ubiquitin/proteasome pathway at three different steps had no effect on the aggregation of Ht fragments in yeast, suggesting that the ubiquitination of Ht previously noted in mammalian cells may not inherently be required for polyQ length-dependent aggregation. Changing the expression levels of a wide variety of chaperone proteins in yeast neither increased nor decreased Ht aggregation. However, Sis1, Hsp70, and Hsp104 overexpression modulated aggregation of HtQ72 and HtQ103 fragments. More dramatically, the deletion of Hsp104 virtually eliminated it. These observations establish yeast as a system for studying the causes and consequences of polyQ-dependent Ht aggregation.
Krobitsch
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Howard Hughes Medical Institute, University of Chicago, Chicago, IL 60637, USA.
Lindquist
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D002499:Centrifugation, Density Gradient; D003546:Cysteine Endopeptidases / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006360:Heat-Shock Proteins / Q000378:metabolism; D008164:Luminescent Proteins; D008856:Microscopy, Fluorescence; D018832:Molecular Chaperones / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000378:metabolism; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000235:genetics*; D009687:Nuclear Proteins / Q000235:genetics*; D010446:Peptide Fragments / Q000737:chemistry; D010455:Peptides / Q000235:genetics*; D010957:Plasmids; D046988:Proteasome Endopeptidase Complex; D029701:Saccharomyces cerevisiae Proteins; D019680:Trinucleotide Repeat Expansion / Q000235:genetics*; D015003:Yeasts
D006360:Heat-Shock Proteins; D008164:Luminescent Proteins; D018832:Molecular Chaperones; D009097:Multienzyme Complexes; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D010446:Peptide Fragments; D010455:Peptides; D029701:Saccharomyces cerevisiae Proteins; C070972:HsP104 protein, S cerevisiae; D049452:Green Fluorescent Proteins; C097188:polyglutamine; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex