Background: Machado-Joseph disease is the most frequent dominant ataxia worldwide. Despite its frequency and presence in many populations, only 2 founder mutations have been suggested to explain its current geographic distribution.Objectives: To trace back in history the main mutational events in Machado-Joseph disease, we aimed to assess ancestral haplotypes and population backgrounds, to date the mutations, and to trace the routes and time of introduction of the founder haplotypes in different populations.Design, Setting, and Participants: We studied 264 families with Machado-Joseph disease from 20 different populations. Six intragenic single-nucleotide polymorphisms were used to determine ancestral mutational events; 4 flanking short tandem repeats were used to construct extended haplotypes and measure accumulation of genetic diversity over time within each lineage. Results:The worldwide-spread lineage, TTACAC, had its highest diversity in the Japanese population, where we identified the ancestral short tandem repeat-based haplotype. Accumulated variability suggested a postneolithic mutation, about 5774±1116 years old, with more recent introductions in North America, Germany, France, Portugal, and Brazil. As to the second mutational event, in the GTGGCA lineage, only 7 families (of 71 families) did not have Portuguese ancestry, although gene diversity was again smaller in Portuguese families (0.44) than in non-Portuguese families (0.93). Conclusions:The worldwide-spread mutation may have first occurred in Asia and later been diffused throughout Europe, with a founder effect accounting for its high prevalence in Portugal; the other Machado-Joseph disease lineage is more recent, about 1416±434 years old, and its dispersion may be explained mainly by recent Portuguese emigration.

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To analyze the ataxin 2 (ATXN2) CAG repeat size in a cohort of patients with amyotrophic lateral sclerosis (ALS) and healthy controls. Large (CAG) n alleles of the ATXN2 gene (27-33 repeats) were recently reported to be associated with an increased risk of ALS.

Spinocerebellar ataxia type 10 (SCA10) is an autosomal dominant neurodegenerative disease characterized by cerebellar ataxia and seizures. The disease is caused by a large ATTCT repeat expansion in the ATXN10 gene. The first families reported with SCA10 were of Mexican origin, but the disease was soon after described in Brazilian families of mixed Portuguese and Amerindian ancestry. The origin of the SCA10 expansion and a possible founder effect that would account for its geographical distribution have been the source of speculation over the last years. To unravel the mutational origin and spread of the SCA10 expansion, we performed an extensive haplotype study, using closely linked STR markers and intragenic SNPs, in families from Brazil and Mexico. Our results showed (1) a shared disease haplotype for all Brazilian and one of the Mexican families, and (2) closely-related haplotypes for the additional SCA10 Mexican families; (3) little or null genetic distance in small normal alleles of different repeat sizes, from the same SNP lineage, indicating that they are being originated by a single step mechanism; and (4) a shared haplotype for pure and interrupted expanded alleles, pointing to a gene conversion model for its generation. In conclusion, we show evidence for an ancestral common origin for SCA10 in Latin America, which might have arisen in an ancestral Amerindian population and later have been spread into the mixed populations of Mexico and Brazil.

Despite the intense debate around the repeat instability reported on the large group of neurological disorders caused by trinucleotide repeat expansions, little is known about the mutation process underlying alleles in the normal range that, ultimately, expand to pathological size. In this study, we assessed the mutation mechanisms by which wild-type Machado -Joseph disease (MJD) alleles have been generated throughout human evolution. Haplotypes including the CAG repeat, six intragenic SNPs and four flanking microsatellites were analysed in 431 normal chromosomes of European, Asian and African origin. A bimodal CAG repeat length frequency distribution was found in the four most frequent wild-type lineages (H1-GCGGCA; H2-GTGGCA; H3-TTAGAC and H4-TTACAC). Based on flanking microsatellite haplotypes, the variance calculated by analysis of molecular variance between modal (CAG) n alleles was little or null in lineages H1, H2 and H4, as were the pairwise differences. Moreover, genetic distances among all the alleles from each lineage did not reflect the allele sizes differences, as expected if a stepwise mutation model was the main process of evolution. On the contrary, when exposed in maximum parsimonious phylogenetic trees, a large number of mutation steps separated same-size alleles, whereas several microsatellite haplotypes were shared by modal CAGs. In conclusion, our results suggest that the main mutation mechanism occurring in the evolution of the polymorphic CAG region at MJD/SCA3 locus is a multistep one, either by gene conversion or DNA slippage; repeats with 14, 21, 23 and 27 CAGs are the main alleles involved in this process.

The spinocerebellar ataxia type 2 (SCA2) is an autosomal dominant neurodegenerative disease characterized by gait and limb ataxia. This disease is caused by the expansion of a (CAG)(n) located in the ATXN2, that encodes a polyglutamine tract of more than 34 repeats. Lately, alleles with 32-33 CAGs have been associated to late-onset disease cases. Repeat interruptions by CAA triplets are common in normal alleles, while expanded alleles usually contain a pure repeat tract. To investigate the mutational origin and the instability associated to the ATXN2 repeat, we performed an extensive haplotype study and sequencing of the CAG/CAA repeat, in a cohort of families of different geographic origins and phenotypes. Our results showed (1) CAA interruptions also in expanded ATXN2 alleles; (2) that pathological CAA interrupted alleles shared an ancestral haplotype with pure expanded alleles; and (3) higher genetic diversity in European SCA2 families, suggesting an older European ancestry of SCA2. In conclusion, we found instability towards expansion in interrupted ATXN2 alleles and a shared ancestral ATXN2 haplotype for pure and interrupted expanded alleles; this finding has strong implications in mutation diagnosis and counseling. Our results indicate that interrupted alleles, below the pathological threshold, may be a reservoir of mutable alleles, prone to expansion in subsequent generations, leading to full disease mutation.

Twelve neurological disorders are caused by gene-specific CAG/CTG repeat expansions that are highly unstable upon transmission to offspring. This intergenerational repeat instability is clinically relevant since disease onset, progression and severity are associated with repeat size. Studies of model organisms revealed the involvement of some DNA replication and repair genes in the process of repeat instability, however, little is known about their role in patients. Here, we used an association study to search for genetic modifiers of (CAG)n instability in 137 parent-child transmissions in Machado-Joseph disease (MJD/SCA3). With the hypothesis that variants in genes involved in DNA replication, repair or recombination might alter the MJD CAG instability patterns, we screened 768 SNPs from 93 of these genes. We found a variant in ERCC6 (rs2228528) associated with an expansion bias of MJD alleles. When using a gene-gene interaction model, the allele combination G-A (rs4140804-rs2972388) of RPA3-CDK7 is also associated with MJD instability in a direction-dependent manner. Interestingly, the transcription-coupled repair factor ERCC6 (aka CSB), the single-strand binding protein RPA, and the CDK7 kinase part of the TFIIH transcription repair complex, have all been linked to transcription-coupled repair. This is the first study performed in patient samples to implicate specific modifiers of CAG instability in humans. In summary, we found variants in three transcription-coupled repair genes associated with the MJD mutation that points to distinct mechanisms of (CAG)n instability.

In repeat expansion disorders, the size of pathological alleles is the most relevant factor accounting for the disease severity and age-at-onset, emphasizing the clinical significance of their underlying intergenerational instability. In one of these diseases, Machado-Joseph disease (MJD), the sex of transmitting progenitor and the C(987)GG/G(987)GG polymorphism are the best studied factors acting on intergenerational instability of expanded alleles. Here, we assessed the influence of other cis and inter-allelic acting factors, at the ATXN3 locus, through the analysis of MJD lineages, flanking STR-based haplotypes, the initial repeat size and parental age. A total of 100 transmissions of the expanded MJD allele were analyzed according to the sex of the transmitting parent. We have shown that independent origin mutations (identified by intragenic SNP-based haplotypes) behave differently, as the status of instability (contraction, no change or further expansion) is concerned. Indeed, 72% of expansions were associated to the worldwide spread TTACAC lineage, whereas the GTGGCA displayed 75% of all contractions observed. The analysis of flanking recombinant haplotypes did not suggest any further distant cis elements acting up- or downstream the ATXN3 locus. Considering the increased amplitude of expansions seen in older transmitting fathers, a repair-based mechanism may be suggested for the meiotic instability at this locus; furthermore, the lack of correlation between the initial repeat size and degree of instability did not support a replication-based mechanism. In summary, our findings point to different mechanisms of instability underlying male and female meioses, as well as contraction and expansion processes in MJD.