Friedreich's ataxia – a case of aberrant transcription termination?

Abstract

R educed expression of the mitochondrial protein Frataxin (FXN) is the underlying cause of Friedreich's ataxia. We propose a model of premature termination of FXN transcription induced by pathogenic expanded GAA repeats that links R-loop structures, antisense transcription, and heterochromatin formation as a novel mechanism of transcriptional repression in Friedreich's ataxia.Expanded GAA Repeats Induce an Altered Chromatin Environment at the FXN Locus in Friedreich's AtaxiaTandem repeats make up »3% of the human genome.1 These sequences are often highly polymorphic and both their normal variability as well as pathological changes can drastically affect phenotype, predominantly via altering gene expression. Mutations in selected short tandem repeats composed of 3-6 nucleotide units are responsible for »30 severe neurological and neuromuscular diseases, including fragile X syndrome, myotonic dystrophy, Huntington's disease, and Friedreich's ataxia (FRDA).2 Friedreich's ataxia, the most common inherited form of ataxia, is a fatal neurodegenerative disease that results from large expansions of GAA trinucleotide repeat sequences in intron 1 of the Frataxin (FXN) gene, leading to transcriptional repression.3 While unaffected individuals carry less than 40 repeats, patients have large GAA tracts reaching several hundreds of GAAs. 4 demonstrated that insertion of these expanded polypurine-polypyrimidine repeats induce position effect variegation leading to transcriptional silencing of the nearby located transgenes. Detailed histone modification analyses conducted in various FRDA animal and cell line models, as well as in FRDA patient cells, readily confirmed the heterochromatin-like characteristics of this locus, featuring enriched histone H3 lysine 9 methylation (H3K9me2/me3) and decreased histone H3 and H4 acetylation.5-7 These GAAinduced chromatin changes were localized in direct proximity of the GAA repeats and did not spread more than 2 kb from the expanded repeats.Although significant downregulation of FXN expression (~75-95%) is undoubtedly the underlying cause of the FRDA, the molecular trigger as well as mechanisms of the transcriptional repression induced by the expanding intronic GAA repeat tract are unclear. Importantly, transcript stability and FXN pre-mRNA splicing are not affected by GAA expansion, nor has formation of GAA repeat-containing RNA foci been detected in FRDA samples.8 A consensus in the field exists regarding significant changes in histone modification patterns induced by GAAexpansions. Treatment with particular histone deacetylase inhibitors results in the moderate elevation of frataxin mRNA levels and is accompanied by increased histone acetylation both in the promoter and vicinity of the GAAs. 8,9 On the contrary, treatment with the G9a histone methyltransferase inhibitor BIX-01294 decreases the level of H3K9me2 but fails to reactivate FXN expression.10 Additional investigation into potential combinatorial approaches is necessary to fully elucidate the effects of histone modification-targ...

Activating frataxin expression by repeat-targeted nucleic acids

“…These protein stability data, together with our data showing unchanged recruitment of RNAP2, suggests that RNA activation by anti-GAA duplex RNAs occurs at the level of RNA synthesis but not at the level of polymerase binding. Increased expression that is not dependent on increased recruitment of RNAP2 is consistent with previous observations of the mechanism of impaired elongation in FRDA patient-derived cells 20 21 30 . Our observation of RNA-induced histone modifications is also consistent with relief of heterochromatin formation, suggesting that transcript elongation is being enhanced.…”

“…This R-loop may then interfere with transcription and reduce FXN expression ( Fig. 1b ), possibly by inducing silencing chromatin modifications that impeded transcriptional elongation 20 21 . Synthetic nucleic acids complementary to the GAA repeat might bind to the expanded RNA, block R-loop formation and remove the trigger for transcriptional silencing.…”

A Defective mRNA Cleavage and Polyadenylation Complex Facilitates Expansions of Transcribed (GAA)n Repeats Associated with Friedreich’s Ataxia

“…Slow transcription elongation is known to stimulate R-loop formation at various sequences, including (GAA) n repeats (Butler and Napierala, 2015; Groh et al, 2014). It was foreseeable, therefore, that increased R-loop formation at (GAA) n repeats in the ysh1-L14S mutant could ultimately promote repeat expansions.…”

Regulation of transcription termination by FUS and TDP-43

“…G-rich DNA tracts can frequently be found in promoters and termination regions, as well as regions with trinucleotide repeat expansions that are responsible for various neuronal diseases (e.g. ALS/FTD, Fragile X syndrome, and Friedreich's Ataxia) (58)(59)(60)(61). Such tracts are known to induce DNA damage and mutations, as they are often stabilized upon the knock-down of certain factors involved in the transcription-coupled nucleotide excision repair (48), homologous recombination (62), and the Fanconi Anemia pathway (53,63).…”