Aptamers Targeted to an RNA Hairpin Show Improved Specificity Compared to that of Complementary Oligonucleotides

Abstract

Aptamers interacting with RNA hairpins through loop-loop (so-called kissing) interactions have been described as an alternative to antisense oligomers for the recognition of RNA hairpins. R06, an RNA aptamer, was previously shown to form a kissing complex with the TAR (trans-activating responsive) hairpin of HIV-1 RNA (Ducongé and Toulmé (1999) RNA 5, 1605). We derived a chimeric locked nucleic acid (LNA)/DNA aptamer from R06 that retains the binding properties of the originally selected R06 aptamer. We demonstrated that this LNA/DNA aptamer competes with a peptide of the retroviral protein Tat for binding to TAR, even though the binding sites of the two ligands do not overlap each other. This suggests that upon binding, the aptamer TAR adopts a conformation that is no longer appropriate for Tat association. In contrast, a LNA/DNA antisense oligomer, which exhibits the same binding constant and displays the same base-pairing potential as the chimeric aptamer, does not compete with Tat. Moreover, we showed that the LNA/DNA aptamer is a more specific TAR binder than the LNA/DNA antisense sequence. These results demonstrate the benefit of reading the three-dimensional shape of an RNA target rather than its primary sequence for the design of highly specific oligonucleotides.

Liquid-crystal NMR structure of HIV TAR RNA bound to its SELEX RNA aptamer reveals the origins of the high stability of the complex

“…Several of these ligands have been shown to regulate TAR-dependent transcription in vitro or in cultured cells (18,21). Chemically modified RNA aptamers, which strongly interact with the apical loop of the TAR hairpin could then be potential candidates for the development of anti-HIV drugs able to disrupt the ternary TAR-Tat-CycT1 complex, leading to abortive RNA synthesis.…”

Inhibition of hepatitis C virus replication and internal ribosome entry site-dependent translation by an RNA molecule

“…Therefore, it seems likely that the appearance of resistant viral variants would be reduced with the use of chimeric inhibitors. Moreover, it has been described that inhibitors able to read the 3D folding of their target RNA exhibit increased specificity compared with those that detect only the primary structure (Darfeuille et al, 2006). HH363-50 is composed of a hammerhead ribozyme that cleaves the HCV genome at nt 363, attached to an aptamer targeting domain IV of HCV-IRES.…”

In vitro selection of BNA (LNA) aptamers

“…Successful examples of 2',4'-BNA/LNA aptamers by post-SELEX modification have been reported. [10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25] Schmidt et al developed an LNA aptamer, TTA1, specific for human tenascin-C (TN-C). 11 The original form TN-9 was selected from a 2'-fluoropyrimidine RNA library (Fig.…”

“…Earlier, use of post-SELEX modification methods were mainly reported in studies on BNA aptamer development. [10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25] However, in recent years, in vitro selection of BNA aptamers is preferred owing to the discovery of polymerases available for BNA-containing oligonucleotide syntheses and genetic engineering of these polymerases. Last year, Pinheiro et al developed Tgo DNA polymerase variants, 26 which enable transcription and reverse transcription of six different XNAs (xenonucleic acids): HNA (1,5-anhydrohexitol nucleic acid), CeNA (cyclohexenyl nucleic acid), ANA (arabinonucleic acid), FANA (2'-fluoroarabinonucleic acid), TNA (α-L-threofuranosyl nucleic acid), and 2',4'-BNA/LNA (Fig.…”