More than 30 human neurodegenerative diseases, including Huntington's disease, spinocerebellar ataxias, and fragile X syndrome, involve expanded poly-glutamine [poly(Q)] repeats of trios of nucleotides, such as CAG and CGG. When they occur within coding regions of genes, the expanded poly(Q) domain may cause misfolding of the affected protein and its aggregation into visible abnormal nuclear inclusions. Repeat poly(Q) lengths are also unstable, undergoing shrinkage and expansion, in some cases involving tens of repeats, upon transmission from parent to child. Expansion of repeat lengths is associated with more severe disease, and in some cases their instability accounts for ''genetic anticipation'' in which disease symptoms in successive generations become progressively more severe.
Trinucleotide-repeat instability has been viewed largely as a matter of DNA metabolism, but recent data suggest that repeat instability may be influenced by poly(Q) protein toxicity. Jung and Bonini have shown that repeat instability can be replicated in the fruit fly, Drosophila melanogaster, and they exploited this organism to better understand the mechanism of instability. Their study provided fresh insights into the role of gene transcription, DNA repair, and the potential complicity of pathogenic poly(Q) proteins that underlie repeat instability.147,148 By showing that changes in repeat length that require decades in humans could be monitored in weeks in the fly, the study opened the way for genetic analysis. Based on the idea that transcription of CAG repeats might be critical for generating instability, they reasoned that forcing a repeat-containing transgene to be active in germ cells might trigger changes in repeat length; they also expected that germline events would be transmitted from one generation to the next as occurs in ''genetic anticipation." They suspected that previous fly models that were more suitable for studying the neurodegenerative phenomenon might have failed to uncover instability because they relied on the expression of repeats in either developing or fully differentiated tissues.
To test their idea, Jung and Bonini expressed a transgene bearing 51-78 CAG repeats in germ cells, and tracked transgenes over nine generations. They found that the overall rate of instability was 20% and that the majority of changes were small, involving expansion or shrinkage of one to three CAG units. However, 10% of changes involved larger (>10) repeat expansions or shrinkages, and expansions consistently outweighed shrinkages similar to what is observed in human poly(Q) diseases.
Jung and Bonini then asked how transcription might influence the size of trinucleotide repeats. They found that fly variants deficient in transcription-coupled DNA repair dampened rates of repeat instability, while decreased levels of adenosine 3,' 5'-monophosphate (cAMP) response element-binding protein (CREB)-binding protein (CBP), a protein found in poly(Q) inclusions that regulates many transcription factors, increasing repeat instability. Because the inhibitory effect of pathogenic poly(Q) proteins on CBP was thought to be due to the loss of histone acetylase activity, the investigators expected that treating flies with histone deacetylase inhibitors to normalize acetylation levels might protect against poly(Q) protein pathogenesis. They found that flies treated with the histone deacetylase inhibitor trichostatin A countered the loss of CBP activity and protected against poly(Q) instability for maternal and paternal transmissions. Although trichostatin can affect multiple pathways, analysis of the pattern of repeat changes suggested that reduced instability was, at least in part, due to compensation for decreased CPB and/or histone acetylase activity because both CBP gene changes and trichostatin treatment preferentially modulated poly(Q) expansions relative to other events.
Jung and Bonini extended these findings by examining the effects of transcription and the contribution of CBP protein to models of Huntington's disease, and by examining instability in a model for CGG premutation expansions in fragile X syndrome, an unstable noncoding trinucleotide repeat. Repeat instability with germline transcription was enhanced in both models, from which they concluded that features of repeat instability, including transcriptional dependence, might be a fundamental property of trinucleotide instability seen in several disease models.
These studies recapitulate several features of human CAG repeat instability including the wide range of repeat changes and the strong bias toward repeat expansions. Finding that poly(Q) protein pathology, that is, a decrease of CBP activity via sequestration or inhibition, might enhance repeat instability was an important consequence of Jung and Bonini's work. They noted that CAG expansions occurred not only generationally, but also somatically in Huntington's disease, and pointed out that modifiers such as Msh2 modulate instability in both germline and somatic tissues. This suggested that findings with respect to germline instability might apply to somatic instability. Finally, Jung and colleagues suggested that repeat instability might be influenced by poly(Q) protein toxicity, and that treatments that restrict repeat instability might also provide an avenue to curb poly(Q) protein toxicity. Among these, histone deacetylase inhibitors are in clinical trials.149
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