DNA elements important for CAG•CTG repeat thresholds in Saccharomyces cerevisiae

Trinucleotide repeat (TNR) instability is of interest because of its central role in human diseases such as Huntington's and its unique genetic features. One distinctive characteristic of TNR instability is a threshold, defined as a minimal repeat length that confers frequent mutations. While thresholds are well established, important risk determinants for disease-causing mutations, their mechanistic analysis has been delayed by the lack of suitably tractable experimental systems. In this study, we directly compared for the first time three DNA elements-TNR sequence, purity and flanking sequence-all of which are suggested in the literature to contribute to thresholds. In a yeast model system, we find that CAG repeats require a substantially longer threshold to contract than CTG tracts, indicating that the lagging template repeat sequence helps determine the threshold. In contrast, ATG interruptions within a CTG run do not inhibit contractions via a threshold mechanism, but by altering the likelihood of forming a hairpin intermediate. The presence of a GC-rich flanking sequence, similar to a haplotype found in some Huntington's patients, does not detectably alter expansions of Okazaki fragment CTG tracts, suggesting no role for this flanking sequence on thresholds. Together these results help better define TNR thresholds by delineating sequence elements that modulate instability.