title: A persistent RNA.DNA hybrid formed by transcription of the Friedreich ataxia triplet repeat in live bacteria, and by T7 RNAP in vitro.
abstract: Expansion of an unstable GAA.TTC repeat in the first intron of the FXN gene causes Friedreich ataxia by reducing frataxin expression. Deficiency of frataxin, an essential mitochondrial protein, leads to progressive neurodegeneration and cardiomyopathy. The degree of frataxin reduction correlates with GAA.TTC tract length, but the mechanism of reduction remains controversial. Here we show that transcription causes extensive RNA.DNA hybrid formation on GAA.TTC templates in bacteria as well as in defined transcription reactions using T7 RNA polymerase in vitro. RNA.DNA hybrids can also form to a lesser extent on smaller, so-called 'pre-mutation' size GAA.TTC repeats, that do not cause disease, but are prone to expansion. During in vitro transcription of longer repeats, T7 RNA polymerase arrests in the promoter distal end of the GAA.TTC tract and an extensive RNA.DNA hybrid is tightly linked to this arrest. RNA.DNA hybrid formation appears to be an intrinsic property of transcription through long GAA.TTC tracts. RNA.DNA hybrids have a potential role in GAA.TTC tract instability and in the mechanism underlying reduced frataxin mRNA levels in Friedreich Ataxia.
authors: Department of Genetics, Louisiana State University Health Sciences Center, 533 Bolivar Street, New Orleans, LA 70112, USA. egrabc@lsuhsc.edu
authors: Ed
authors: E
authors: Grabczyk
authors: Miriam
authors: M
authors: Mancuso
authors: Mimi C
authors: MC
authors: Sammarco
mesh_terms: D001419:Bacteria / Q000235:genetics; D004247:DNA / Q000096:biosynthesis*; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D033862:Iron-Binding Proteins / Q000235:genetics*; D012313:RNA / Q000096:biosynthesis*; D013698:Templates, Genetic; D014158:Transcription, Genetic; D018911:Trinucleotide Repeats; D014764:Viral Proteins / Q000378:metabolism*; D000096923:Frataxin
chemical_list: D004247:DNA; D012321:DNA-Directed RNA Polymerases; D033862:Iron-Binding Proteins; D012313:RNA; D014764:Viral Proteins; D000096923:Frataxin; C062210:bacteriophage T7 RNA polymerase
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