Abstract
BackgroundFriedreich ataxia, an autosomal recessive neurodegenerative and cardiac disease, is caused by abnormally low levels of frataxin, an essential mitochondrial protein. All Friedreich ataxia patients carry a GAA⋅TTC repeat expansion in the first intron of the frataxin gene, either in the homozygous state or in compound heterozygosity with other loss-of-function mutations. The GAA expansion inhibits frataxin expression through a heterochromatin-mediated repression mechanism. Histone modifications that are characteristic of silenced genes in heterochromatic regions occur at expanded alleles in cells from Friedreich ataxia patients, including increased trimethylation of histone H3 at lysine 9 and hypoacetylation of histones H3 and H4.Methodology/Principal FindingsBy chromatin immunoprecipitation, we detected the same heterochromatin marks in homozygous mice carrying a (GAA)230 repeat in the first intron of the mouse frataxin gene (KIKI mice). These animals have decreased frataxin levels and, by microarray analysis, show significant gene expression changes in several tissues. We treated KIKI mice with a novel histone deacetylase inhibitor, compound 106, which substantially increases frataxin mRNA levels in cells from Friedreich ataxia individuals. Treatment increased histone H3 and H4 acetylation in chromatin near the GAA repeat and restored wild-type frataxin levels in the nervous system and heart, as determined by quantitative RT-PCR and semiquantitative western blot analysis. No toxicity was observed. Furthermore, most of the differentially expressed genes in KIKI mice reverted towards wild-type levels.Conclusions/SignificanceLack of acute toxicity, normalization of frataxin levels and of the transcription profile changes resulting from frataxin deficiency provide strong support to a possible efficacy of this or related compounds in reverting the pathological process in Friedreich ataxia, a so far incurable neurodegenerative disease.
Highlights
Friedreich ataxia (FRDA, OMIM 229300) is an inherited recessive disorder characterized by progressive neurological disability and heart disease [1]
Results fxn (GAA)230/(GAA)230 (KIKI) mice recapitulate the genetic and epigenetic features of FRDA and have an altered gene expression profile as a consequence of mild frataxin deficiency We previously generated a knock-in mouse carrying a (GAA)230 repeat in the first intron of the endogenous frataxin gene [24], and we showed that homozyogous fxn(GAA)230/(GAA)230 (KIKI) mice have mildly but significantly lower frataxin levels than wildtype (WT) animals with the same strain background (C57Bl/6), as determined by densitometry of western blots and semiquantitave RT-PCR [24]
We found changes that correspond to those occurring in cells from FRDA patients [22], though they are quantitatively less marked, as expected given the relatively short GAA repeats in KIKI mice compared to a typical FRDA patient (Figure 1B)
Summary
Friedreich ataxia (FRDA, OMIM 229300) is an inherited recessive disorder characterized by progressive neurological disability and heart disease [1]. A group of 12 KIKI mice received subcutaneous injections of 150 mg/kg of 106 every 24 hours for three consecutive days to evaluate the effects of the compound on fxn mRNA and frataxin protein levels and on histone modifications near the GAA repeat. We checked whether in KIKI mice, as observed in lymphocytes from FRDA patients, 106 treatment was accompanied by changes in histone modifications in chromatin near the GAA repeat that could lead to a more transcription-permissive state. The most striking finding concerned the set of genes that were differentially expressed in untreated KIKI vs WT mice Most of these genes (67% in brain, 84% in cerebellum, 67% in heart) showed coordinate changes towards normal levels after 106 treatment (Figure 4B). Treatment with 106 modified the global gene expression profile in KIKI animals to resemble the WT profile
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