title: Polypurine reverse-Hoogsteen (PPRH) oligonucleotides can form triplexes with their target sequences even under conditions where they fold into G-quadruplexes.
abstract: Polypurine reverse-Hoogsteen (PPRH) oligonucleotides are non-modified DNA molecules composed of two mirror-symmetrical polypurine stretches linked by a five-thymidine loop. They can fold into reverse-Hoogsteen hairpins and bind to their polypyrimidine target sequence by Watson-Crick bonds forming a three-stranded structure. They have been successfully used to knockdown gene expression and to repair single-point mutations in cells. In this work, we provide an in vitro characterization (UV and fluorescence spectroscopy, gel electrophoresis and nuclease assays) of the structure and stability of two repair-PPRH oligonucleotides and of the complexes they form with their single-stranded targets. We show that one PPRH oligonucleotide forms a hairpin, while the other folds, in potassium, into a guanine-quadruplex (G4). However, the hairpin-prone oligonucleotide does not form a triplex with its single-stranded target, while the G4-prone oligonucleotide converts from a G4 into a reverse-Hoogsteen hairpin forming a triplex with its target sequence. Our work proves, in particular, that folding of a PPRH oligonucleotide into a G4 does not necessarily impair sequence-specific DNA recognition by triplex formation. It also illustrates an original example of DNA structural conversion of a G4 into a reverse-Hoogsteen hairpin driven by triplex formation; this kind of conversion might occur at particular loci of genomic DNA.
authors.affiliation: Department of Biochemistry and Physiology, School of Pharmacy, University of Barcelona, Barcelona, Spain.
authors.forename: Anna
authors.initials: A
authors.lastname: Solé
authors.affiliation: Department of Biochemistry and Physiology, School of Pharmacy, University of Barcelona, Barcelona, Spain.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Delagoutte
authors.affiliation: Department of Biochemistry and Physiology, School of Pharmacy, University of Barcelona, Barcelona, Spain.
authors.forename: Carlos J
authors.initials: CJ
authors.lastname: Ciudad
authors.affiliation: Department of Biochemistry and Physiology, School of Pharmacy, University of Barcelona, Barcelona, Spain.
authors.forename: Véronique
authors.initials: V
authors.lastname: Noé
authors.affiliation: Structure et Instability of Genomes, Sorbonne Universités, Muséum National d'Histoire Naturelle, Inserm U 1154, CNRS UMR 7196, Paris, France.
authors.forename: Patrizia
authors.initials: P
authors.lastname: Alberti
mesh_terms: D020816:Amino Acid Motifs / Q000235:genetics; D001483:Base Sequence; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D004260:DNA Repair; D005136:Eye Proteins / Q000235:genetics; D054856:G-Quadruplexes; D016678:Genome; D018398:Homeodomain Proteins / Q000235:genetics; D009419:Nerve Tissue Proteins / Q000235:genetics; D009690:Nucleic Acid Conformation; D009841:Oligonucleotides / Q000235:genetics*; D017354:Point Mutation / Q000235:genetics; D013050:Spectrometry, Fluorescence; D000097607:Homeobox Protein SIX3
chemical_list: D004247:DNA; D005136:Eye Proteins; D018398:Homeodomain Proteins; D009419:Nerve Tissue Proteins; D009841:Oligonucleotides; D000097607:Homeobox Protein SIX3; C051802:triplex DNA