Background: The Repeat Expansion Diseases are caused by intergenerational expansions of a specific tandem repeat. More than 20 such diseases that belong to this group have been identified thus far. The Fragile X-related disorders (FXDs) arise from expansion of a CGG.CCG-repeat in the 5'UTR of the X-linked FMR1 gene. Carriers of alleles with 55-200 repeats, so-called premutation (PM) alleles, are at risk for a neurodegenerative disorder, Fragile X-associated tremor/ataxia syndrome (FXTAS) and a form of ovarian dysfunction known as FX-associated primary ovarian insufficiency (FXPOI). Furthermore, in females, the premutation allele can undergo expansion on intergenerational transfer that can result in their children having alleles with >200 repeats. This expanded allele is known as a full mutation (FM) and individuals who inherit such alleles almost always have Fragile X syndrome (FXS), which is the leading heritable cause of intellectual disability. FM alleles become silenced. This results in a deficiency of the protein product of this gene, FMRP, which is involved in the regulation of translation of a subset of mRNAs. The FMRP deficiency in brain results in aberrant dendritic spine morphology and a defective response to synaptic activation. The mechanism of gene silencing is unknown, but may show parallels to Friedreich ataxia, a related disorder that also shows repeat-mediated gene silencing. Progress report: We have previously identified a number of steps in the Fragile X gene silencing process including some that precede DNA methylation and some that occur very late in the silencing process (Biacsi, Kumari and Usdin, 2008). One of the late steps in FX gene silencing turns out to be the deacetylation of histone H4 on lysine 16, a step we showed to be carried out by SIRT1, a class III histone deacetylase. We have shown that SIRT1 inhibition reactivates the FMR1 gene without requiring DNA demethylation (Biacsi, Kumari and Usdin, 2008). It may thus be useful in cells like neurons where the effect of gene silencing is most apparent. More recently we have shown that reactivation of silenced alleles either with SIRT1 inhibitors or the DNA methyltransferase inhibitor 5-azadeoxycytidine is associated with an increase in the levels of a mark of repressed heterochromatin, H3K27me3 (Kumari and Usdin, 2014). This mark is normally deposited by components of Polycomb Repressive Complex 2. We have shown that H3K27me3 deposition is dependent on the presence of FMR1 mRNA and that knocking down the FMR1 transcript prevents gene resilencing. Thus a catch-22 situation exists in FXS, where transcription of the FX allele triggers the recruitment of repressive protein complexes that lead to the gene being switched off. We have also developed a number of induced pluripotent stem cell (iPSCs) lines from individuals with FXS as well as FXTAS and FXPOI. We are using these cells to derived disease-relevant cell types for use in understanding disease pathology and screening for small molecules able to ameliorate the disease phenotype.
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