Abstract
Huntington’s disease is the result of a long polyglutamine tract in the gene encoding huntingtin protein, which in turn causes a large number of cellular changes and ultimately results in neurodegeneration of striatal neurons. Although many theories have been proposed, the precise mechanism by which the polyglutamine expansion causes cellular changes is not certain. Some evidence supports the hypothesis that the long polyglutamine tract inhibits the proteasome, a multiprotein complex involved in protein degradation. However, other studies report normal proteasome function in cells expressing long polyglutamine tracts. The controversy may be due to the methods used to examine proteasome activity in each of the previous studies. In the present study, we measured proteasome function by examining levels of endogenous peptides that are products of proteasome cleavage. Peptide levels were compared among mouse striatal cell lines expressing either 7 glutamines (STHdh Q7/Q7) or 111 glutamines in the huntingtin protein, either heterozygous (STHdh Q7/Q111) or homozygous (STHdh Q111/Q111). Both of the cell lines expressing huntingtin with 111 glutamines showed a large reduction in nearly all of the peptides detected in the cells, relative to levels of these peptides in cells homozygous for 7 glutamines. Treatment of STHdh Q7/Q7 cells with proteasome inhibitors epoxomicin or bortezomib also caused a large reduction in most of these peptides, suggesting that they are products of proteasome-mediated cleavage of cellular proteins. Taken together, these results support the hypothesis that proteasome function is impaired by the expression of huntingtin protein containing long polyglutamine tracts.
Highlights
Huntington’s disease (HD) is an autosomal dominant neurodegenerative disorder caused by expansion of CAG trinucleotide repeats within exon 1 of the HTT gene that encodes huntingtin protein [1,2,3]
Some studies found evidence in support of this theory, while other studies did not [9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29]. These previous studies used a variety of experimental approaches to assay proteasome activity; some measured cleavage of fluorogenic peptides that are proteasome substrates while others measured levels of reporter proteins expressed with or without a degradation signal. Each of these approaches have yielded insights about the role of proteasome function in HD, but each method explores a different component of the ubiquitinproteasome pathway and is limited by the ability to examine the degradation of only a few select substrates
Q7Q7, Q7Q111, and Q111Q111 cell lines were previously established from wild-type and Hdh (Q111) knock-in embryos, as described by Trettel et al [36]
Summary
Huntington’s disease (HD) is an autosomal dominant neurodegenerative disorder caused by expansion of CAG trinucleotide repeats within exon 1 of the HTT gene that encodes huntingtin protein [1,2,3]. Some studies found evidence in support of this theory, while other studies did not [9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29] These previous studies used a variety of experimental approaches to assay proteasome activity; some measured cleavage of fluorogenic peptides that are proteasome substrates while others measured levels of reporter proteins expressed with or without a degradation signal. A more accurate understanding of endogenous proteasome function can come from measuring levels of endogenous products of the proteasome, i.e. intracellular peptides
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