title: An LNA-based loss-of-function assay for micro-RNAs.
abstract: MicroRNAs (miRNAs) have recently emerged as being essential for development and for the control of cell proliferation/differentiation in various organisms. However, little is known about miRNA function and mode of action at the cellular level. We have designed a miRNA loss-of-function assay, based on chemically modified locked nucleic acids (LNA) antisense oligonucleotides and usable in tissue culture cells. We show that LNA/DNA mixed oligonucleotides form highly stable duplexes with miRNAs in vitro. Ex vivo, the target miRNA becomes undetectable in cells transfected with the antisense oligonucleotide. The effect is dose-dependent, long-lasting, and specific. Moreover, using a reporter assay, we show that antisense LNA/DNA oligonucleotides inhibit short non-coding RNAs at the functional level. Thus LNA/DNA mixmers represent powerful tools for functional analysis of miRNAs.
authors.affiliation: CNRS FRE 2944, Ligue nationale contre le cancer, 7, rue Guy Moquet, 94801 Villejuif cedex, France. nirina@vjf.cnrs.fr
authors.forename: I
authors.initials: I
authors.lastname: Naguibneva
authors.forename: M
authors.initials: M
authors.lastname: Ameyar-Zazoua
authors.forename: N
authors.initials: N
authors.lastname: Nonne
authors.forename: A
authors.initials: A
authors.lastname: Polesskaya
authors.forename: S
authors.initials: S
authors.lastname: Ait-Si-Ali
authors.forename: R
authors.initials: R
authors.lastname: Groisman
authors.forename: M
authors.initials: M
authors.lastname: Souidi
authors.forename: L L
authors.initials: LL
authors.lastname: Pritchard
authors.forename: A
authors.initials: A
authors.lastname: Harel-Bellan
mesh_terms: D002478:Cells, Cultured; D006801:Humans; D035683:MicroRNAs / Q000037:antagonists & inhibitors / Q000502:physiology*; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*
chemical_list: D035683:MicroRNAs; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense; C477371:locked nucleic acid