Expression of Human Frataxin Is Regulated by Transcription Factors SRF and TFAP2

Kuanyu Li,Liang Pan,Xiaoman Dai,Zhuangzhuang Cong,Daniel R Crooks,Dung Ha,Anamika Singh,Tracey A Rouault,Sumitra Deb

doi:10.1371/journal.pone.0012286

Abstract

BackgroundFriedreich ataxia is an autosomal recessive neurodegenerative disease caused by reduced expression levels of the frataxin gene (FXN) due to expansion of triplet nucleotide GAA repeats in the first intron of FXN. Augmentation of frataxin expression levels in affected Friedreich ataxia patient tissues might substantially slow disease progression.Methodology/Principal FindingsWe utilized bioinformatic tools in conjunction with chromatin immunoprecipitation and electrophoretic mobility shift assays to identify transcription factors that influence transcription of the FXN gene. We found that the transcription factors SRF and TFAP2 bind directly to FXN promoter sequences. SRF and TFAP2 binding sequences in the FXN promoter enhanced transcription from luciferase constructs, while mutagenesis of the predicted SRF or TFAP2 binding sites significantly decreased FXN promoter activity. Further analysis demonstrated that robust SRF- and TFAP2-mediated transcriptional activity was dependent on a regulatory element, located immediately downstream of the first FXN exon. Finally, over-expression of either SRF or TFAP2 significantly increased frataxin mRNA and protein levels in HEK293 cells, and frataxin mRNA levels were also elevated in SH-SY5Y cells and in Friedreich ataxia patient lymphoblasts transfected with SRF or TFAP2.Conclusions/SignificanceWe identified two transcription factors, SRF and TFAP2, as well as an intronic element encompassing EGR3-like sequence, that work together to regulate expression of the FXN gene. By providing new mechanistic insights into the molecular factors influencing frataxin expression, our results should aid in the discovery of new therapeutic targets for the treatment of Friedreich ataxia.

Highlights

Friedreich’s ataxia, the most common inherited ataxia, is an autosomal recessive neurodegenerative disease caused by expansion of triplet nucleotide GAA repeats in the first intron of the frataxin gene (FXN) gene
We initiated the current study by using Genomatix software to identify putative transcription factor binding sites within the promoter that might serve as transcriptional regulatory elements of the FXN gene
It has been suggested that restoration of frataxin gene expression levels in Friedreich ataxia patients to levels observed in heterozygotes may substantially slow disease progression

Summary

Introduction

Friedreich’s ataxia, the most common inherited ataxia, is an autosomal recessive neurodegenerative disease caused by expansion of triplet nucleotide GAA repeats in the first intron of the FXN gene. Deficiency of iron-sulfur (Fe-S) clustercontaining mitochondrial respiratory chain enzymes is a feature found both in patient cardiac biopsies and in Yfh1p-deficient S. cerevisiae [3] These seminal findings regarding frataxin function have led to further work suggesting potential roles for human frataxin (and its homologues in lower organisms) in cellular functions including as an iron donor for heme biosynthesis [4], as an iron storage protein [5], as an iron chaperone [6] or accessory protein [7] important for Fe-S cluster assembly. Friedreich ataxia is an autosomal recessive neurodegenerative disease caused by reduced expression levels of the frataxin gene (FXN) due to expansion of triplet nucleotide GAA repeats in the first intron of FXN. Augmentation of frataxin expression levels in affected Friedreich ataxia patient tissues might substantially slow disease progression

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