title: A combinatorial approach to the repertoire of RNA kissing motifs; towards multiplex detection by switching hairpin aptamers.
abstract: Loop-loop (also known as kissing) interactions between RNA hairpins are involved in several mechanisms in both prokaryotes and eukaryotes such as the regulation of the plasmid copy number or the dimerization of retroviral genomes. The stability of kissing complexes relies on loop parameters (base composition, sequence and size) and base combination at the loop-loop helix - stem junctions. In order to identify kissing partners that could be used as regulatory elements or building blocks of RNA scaffolds, we analysed a pool of 5.2 × 10(6) RNA hairpins with randomized loops. We identified more than 50 pairs of kissing RNA hairpins. Two kissing motifs, 5'CCNY and 5'RYRY, generate highly stable complexes with KDs in the low nanomolar range. Such motifs were introduced in the apical loop of hairpin aptamers that switch between unfolded and folded state upon binding to their cognate target molecule, hence their name aptaswitch. The aptaswitch-ligand complex is specifically recognized by a second RNA hairpin named aptakiss through loop-loop interaction. Taking advantage of our kissing motif repertoire we engineered aptaswitch-aptakiss modules for purine derivatives, namely adenosine, GTP and theophylline and demonstrated that these molecules can be specifically and simultaneously detected by surface plasmon resonance or by fluorescence anisotropy.
authors.affiliation: University of Bordeaux, ARNA Laboratory, 146 rue Léo Saignat, 33076 Bordeaux, France Inserm U1212, 146 rue Léo Saignat, 33076 Bordeaux, France CNRS UMR5320, 146 rue Léo Saignat, 33076 Bordeaux, France.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Durand
authors.affiliation: University of Bordeaux, ARNA Laboratory, 146 rue Léo Saignat, 33076 Bordeaux, France Inserm U1212, 146 rue Léo Saignat, 33076 Bordeaux, France CNRS UMR5320, 146 rue Léo Saignat, 33076 Bordeaux, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Dausse
authors.affiliation: University Grenoble Alpes, Département de Pharmacochimie Moléculaire, CNRS UMR5063, 38400 St Martin d'Hères, France.
authors.forename: Emma
authors.initials: E
authors.lastname: Goux
authors.affiliation: University Grenoble Alpes, Département de Pharmacochimie Moléculaire, CNRS UMR5063, 38400 St Martin d'Hères, France.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Fiore
authors.affiliation: University Grenoble Alpes, Département de Pharmacochimie Moléculaire, CNRS UMR5063, 38400 St Martin d'Hères, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Peyrin
authors.affiliation: University Grenoble Alpes, Département de Pharmacochimie Moléculaire, CNRS UMR5063, 38400 St Martin d'Hères, France.
authors.forename: Corinne
authors.initials: C
authors.lastname: Ravelet
authors.affiliation: University of Bordeaux, ARNA Laboratory, 146 rue Léo Saignat, 33076 Bordeaux, France Inserm U1212, 146 rue Léo Saignat, 33076 Bordeaux, France CNRS UMR5320, 146 rue Léo Saignat, 33076 Bordeaux, France jean-jacques.toulme@inserm.fr.
authors.forename: Jean-Jacques
authors.initials: JJ
authors.lastname: Toulmé
mesh_terms: D000241:Adenosine / Q000737:chemistry; D052157:Aptamers, Nucleotide / Q000737:chemistry*; D020029:Base Pairing; D005454:Fluorescence Polarization; D006160:Guanosine Triphosphate / Q000737:chemistry; D055029:Inverted Repeat Sequences; D007700:Kinetics; D008024:Ligands; D059372:Nucleotide Motifs; D012313:RNA / Q000737:chemistry*; D020349:Surface Plasmon Resonance
chemical_list: D052157:Aptamers, Nucleotide; D008024:Ligands; D012313:RNA; D006160:Guanosine Triphosphate; D000241:Adenosine