A GAA repeat expansion reporter model of friedreich's ataxia recapitulates the genomic context and allows rapid screening of therapeutic compounds

Abstract

Friedreich's ataxia (FRDA) is caused by large GAA expansions in intron 1 of the frataxin gene (FXN), which lead to reduced FXNexpression through amechanism not fully understood. Understanding such mechanismis essential for the identification of novel therapies for FRDA and this can be accelerated by the development of cell models which recapitulate the genomic context of the FXN locus and allow direct comparison of normal and expanded FXN loci with rapid detection of frataxin levels. Here we describe the development of the first GAA-expanded FXN genomic DNA reporter model of FRDA. We modified BAC vectors carrying the whole FXN genomic DNA locusbyinsertingtheluciferasegeneinexon5aof theFXNgene(pBAC-FXN-Luc)andreplacingthesixGAArepeats present in the vector with an ~310GAA repeat expansion (pBAC-FXN-GAA-Luc).We generated human clonal cell lines carrying the two vectors using site-specific integration to allow direct comparison of normal and expanded FXN loci. We demonstrate that the presence of expanded GAA repeats recapitulates the epigeneticmodifications andrepressionofgeneexpression seen in FRDA. We appliedtheGAA-expandedreportermodel tothescreeningof a library of novel smallmolecules and identified one molecule which up-regulatesFXN expression inFRDApatient primary cells andrestores normal histone acetylation around theGAArepeats. These results suggest the potential use of genomic reporter cell models for the study of FRDA and the identification of novel therapies, combining physiologically relevant expression with the advantages of quantitative reporter gene expression. © The Author 2013.