The Triplet Expansion Diseases (TREDs) are a group of genetic disorders that result from an expansion or increase in the size of a triplet repeat array in the coding or regulatory region of the affected gene. These disorders include Fragile X syndrome, the most common heritable form of mental retardation, and Friedreich's ataxia, the most common recessive ataxia. All currently known TREDs involve the triplets CGG, CAG, or GAA, but it is not clear whether this reflects ascertainment bias, or some fundamental property of these triplets. We are interested in both the mechanism of expansion and the consequences of expansion in these two disorders. We and others have shown that these repeats form a variety of intrastrand structures, and that the ability to form such structures is a common property of hypervariable sequences. In the course of this work we have characterized a variety of new DNA structures that lead us to believe that the number of sequences in the human genome that are able to form secondary structures may be much larger than previously appreciated. For example we have found that the mouse Ms6-hm locus forms an usual hairpin containing a mixture of G.C, G.G and G.A basepairs, as well as a tetraplex containing extrahelical adenines and neutral C.C pairs, and a tetraplex containing G.A pairs and hemiprotonated C.C pairs. We have also shown that repeats of certain triplets that are currently not associated with a TRED form structures similar to those that are. In particular we have shown that the repeats CGG, TGG and AGG form tetraplexes in which the guanines are all involved in G4-tetrads with the non-G bases stacked within the helix. This finding suggests either that the category of triplet expansion diseases could be larger than currently appreciated, or that structure formation is not necessary or sufficient for expansion. We have generated mice containing transgenes corresponding to alleles of the human FMR1 gene that have a ~100% likelihood of expansion in the offspring of maternal carriers of Fragile X syndrome. These transgenes show no evidence of expansion in mice after more than 5 generations in the heterozygous state. No expansion was seen in mice homozygous for the transgene either. The CGG-tracts do have a mutation rate approaching 100% but these mutations involve only the gain or loss of a small number of repeats. Our data support the idea that other factors are important for expansion. We are currently trying to identify these factors using a variety of approaches including crossing our existing transgenic mice to mice deficient in various DNA damage repair/survelliance systems. Most cases of Friedreich's ataxia result from an expansion of a GAA-tract in the first intron of the frataxin gene. Transcriptional analysis of templates containing long GAA-tracts has given us insight into the molecular basis of the disease etiology and has identified a novel means by which gene expression can be modulated.
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