title: Disruption of Higher Order DNA Structures in Friedreich's Ataxia (GAA)n Repeats by PNA or LNA Targeting.
abstract: Expansion of (GAA)n repeats in the first intron of the Frataxin gene is associated with reduced mRNA and protein levels and the development of Friedreich's ataxia. (GAA)n expansions form non-canonical structures, including intramolecular triplex (H-DNA), and R-loops and are associated with epigenetic modifications. With the aim of interfering with higher order H-DNA (like) DNA structures within pathological (GAA)n expansions, we examined sequence-specific interaction of peptide nucleic acid (PNA) with (GAA)n repeats of different lengths (short: n=9, medium: n=75 or long: n=115) by chemical probing of triple helical and single stranded regions. We found that a triplex structure (H-DNA) forms at GAA repeats of different lengths; however, single stranded regions were not detected within the medium size pathological repeat, suggesting the presence of a more complex structure. Furthermore, (GAA)4-PNA binding of the repeat abolished all detectable triplex DNA structures, whereas (CTT)5-PNA did not. We present evidence that (GAA)4-PNA can invade the DNA at the repeat region by binding the DNA CTT strand, thereby preventing non-canonical-DNA formation, and that triplex invasion complexes by (CTT)5-PNA form at the GAA repeats. Locked nucleic acid (LNA) oligonucleotides also inhibited triplex formation at GAA repeat expansions, and atomic force microscopy analysis showed significant relaxation of plasmid morphology in the presence of GAA-LNA. Thus, by inhibiting disease related higher order DNA structures in the Frataxin gene, such PNA and LNA oligomers may have potential for discovery of drugs aiming at recovering Frataxin expression.
authors.affiliation: Department of Medical Biochemistry and Microbiology, Microbiology-Immunology, Uppsala University, Uppsala, Sweden.|Department of Laboratory Medicine, Clinical Research Center, Karolinska Institutet, SE-141 86, Huddinge, Sweden.
authors.forename: Helen
authors.initials: H
authors.lastname: Bergquist
authors.affiliation: Department of Laboratory Medicine, Clinical Research Center, Karolinska Institutet, SE-141 86, Huddinge, Sweden.
authors.forename: Cristina S J
authors.initials: CS
authors.lastname: Rocha
authors.affiliation: KTH Royal Institute of Technology, School of Chemical Science and Engineering, Department of Chemistry, Stockholm, Sweden.
authors.forename: Rubén
authors.initials: R
authors.lastname: Álvarez-Asencio
authors.affiliation: Laboratoire de Pharmacochimie, Institut Curie, PSL Research University, UMR 9187 - U 1196 CNRS-Institut Curie, INSERM, Centre Universitaire, Orsay, France.
authors.forename: Chi-Hung
authors.initials: CH
authors.lastname: Nguyen
authors.affiliation: KTH Royal Institute of Technology, School of Chemical Science and Engineering, Department of Chemistry, Stockholm, Sweden.
authors.forename: Mark W
authors.initials: MW
authors.lastname: Rutland
authors.affiliation: Department of Laboratory Medicine, Clinical Research Center, Karolinska Institutet, SE-141 86, Huddinge, Sweden.
authors.forename: C I Edvard
authors.initials: CI
authors.lastname: Smith
authors.affiliation: Department of Pathology and Infectious Diseases, Royal Veterinary College, University of London, United Kingdom.
authors.forename: Liam
authors.initials: L
authors.lastname: Good
authors.affiliation: Department of Cellular and Molecular Medicine, Faculty of Health and Medical Sciences, University of Copenhagen, The Panum Institute, Copenhagen, Denmark.
authors.forename: Peter E
authors.initials: PE
authors.lastname: Nielsen
authors.affiliation: Department of Laboratory Medicine, Clinical Research Center, Karolinska Institutet, SE-141 86, Huddinge, Sweden.|Department of Clinical Genetics, Centre for Rare Diseases, Karolinska University Hospital, SE-171 76, Stockholm, Sweden.
authors.forename: Rula
authors.initials: R
authors.lastname: Zain
mesh_terms: D001483:Base Sequence; D004247:DNA / Q000737:chemistry*; D005621:Friedreich Ataxia / Q000235:genetics*; D006801:Humans; D009690:Nucleic Acid Conformation; D059372:Nucleotide Motifs / Q000235:genetics; D009841:Oligonucleotides / Q000378:metabolism*; D020135:Peptide Nucleic Acids / Q000378:metabolism*; D019680:Trinucleotide Repeat Expansion / Q000235:genetics*
chemical_list: D009841:Oligonucleotides; D020135:Peptide Nucleic Acids; C477371:locked nucleic acid; C051802:triplex DNA; D004247:DNA