Mechanisms of the triplet repeat expansion in eukaryotic chromatin.

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Since 1991, more than 10 human hereditary diseases, including Fragile X syndrome, myotonic dystrophy, and Friedreichs's ataxia have been identified to be associated with the expansion of (CTG)_n, (CGG)_n, and (GAA)_n. Several mechanisms of expansions have been proposed based on the features that these repeats can form a variety of alternative DNA conformations including hairpins, slipped strand structures, quadruplex DNA and intramolecular triplexes, yet exact mechanisms are unknown. Since in eukaryotic cells DNA is packaged into arrays of nucleosomes, chromatin may be involved in the triplet repeat diseases.The purpose of this project is to elucidate mechanisms of triplet repeat expansion causing genetic diseases. To address this issue, we examined effects of triplet repeat sequences on the chromatin organization and gene expression in vivo. We developed an assay system with which to examine the effects of DNA sequences on the nucleosome formation in yeast minichromosomes in vivo. Usi … More ng this system, we show that (CGG)12 disrupts an array of positioned nucleosomes. Such destabilization of nucleosomes can be accounted for in terms of formation of stable higher-order DNA structures (haipins, quadruplex or sllipped structures) or high energy cost of nucleosome formation with the sequences. In contrast, it was shown that CTG repeats promote nucleosome formation. Since nuclesomes acts as a general represser for gene expression, the CTG repeats may modulate expression of disease-causing genes via chromatin structural changes. A (GAA)_<12> sequence was incorporated in a nucleosome, but nuclesome positioning in the neighboring region was altered. We also find that (CGG)_<12>, but not (CTG)_<12> and (GAA)_<12> increased a reporter lacZ expression from UAS-less promoters. This is consistent with the finding that (CGG)_<12> destabilizes nucleosomes. In summary, triplet repeat sequences can affect the chromatin organization as well as gene expresion in vivo, which may contribute to the expansion phenomenon and the disease phynotypes. Less