UC Davis

I. Abstract

Allele instability in trinucleotides repeat disorders has been associated with many different physical and psychological conditions, including inherited forms of autism and neurodegenerative disorders. This form of instability is observed in Fragile X Syndrome, a trinucleotide disorder, in which a CGG repeat located in the 5’UTR of the fragile X messenger ribonucleoprotein 1 (FMR1) gene is greater than 200 CGG repeats. This leads to methylation of the gene, transcriptional silencing and consequent absence or reduction of the encoded protein, FMRP. In FMR1 premutation (PM) carriers, who have an allele containing between 55 – 200 CGG repeat length, a CGG repeat expansion during transmission to the offspring, leads to Fragile X Syndrome. Although allele instability has been observed mainly in full mutation alleles (>200 CGG repeats), it has been observed throughout the CGG range, and leading to somatic mosaicism.

However, it is unclear what molecular factors are associated with the risk of FMR1 CGG repeat expansion and if instability correlates with clinical conditions throughout the lifespan of individuals carrying an expanded allele. Furthermore, it is unknown if these factors confer difference in the degrees of risk based on the individual’s biological sex, or age, or if instability or changes may occur over time within individuals.

In this study, we investigated 426 PM female and 454 PM male carriers. Within these two cohorts, we observed that CGG repeat size correlates with FMR1 mRNA levels, and with the number of AGG interruptions, confirming previous reports. Interestingly, a lower number of AGG interruptions and higher CGG repeat size increases the risk of expansion from mother to offspring during transmission, relevant to the new observation here reported. When studying CGG instability over time, we found that eight PM females (n=24) underwent allele expansion as they aged, with three individuals displaying an increase of three or greater repeats in CGG repeat number. Likewise, in the PM male group (n=50), 19 individuals showed an increased CGG repeat number over time, with two undergoing CGG allele size decrease. We also found that the expanded unmethylated regions, significantly correlated with CGG repeat size and AGG interruptions in both females and males.

Thus, CGG repeat size and AGG interruptions are significantly correlated with somatic instability, regardless of gender, although differences in allele expansion patterns between PM males and females exist. Trans molecular factors such as DNA repair-associated genes are also capable of affecting risk of expansion. Indeed, our preliminary observations found a significant correlation between allele in two genes, MSH3 and FAN1, both of which play a role in DNA repair.

These findings carry the implications that molecular measures could be used to determine individuals with higher likelihood of allele instability, which may influence the phenotypic expression. Overall, this study can help future diagnostics in determining which PM individuals, both males and females, are more likely to experience allele expansion and be at risk of developing Fragile X associated conditions.