Developmental defects by antisense-mediated inactivation of micro-RNAs 2 and 13 in Drosophila and the identification of putative target genes.
Micro-RNAs are a class of small non-coding regulatory RNAs that impair translation by imperfect base pairing to mRNAs. For analysis of their cellular function we injected different miRNA-specific DNA antisense oligonucleotides in Drosophila embryos. In four cases we observed severe interference with normal development, one had a moderate impact and six oligonucleotides did not cause detectable phenotypes. We further used the miR-13a DNA antisense oligonucleotide as a PCR primer on a cDNA library template. In this experimental way we identified nine Drosophila genes, which are characterised by 3' untranslated region motifs that allow imperfect duplex formation with miR-13 or related miRNAs. These genes, which include Sos and Myd88, represent putative targets for miRNA regulation. Mutagenesis of the target motif of two genes followed by transfection in Drosophila Schneider 2 (S2) cells and subsequent reporter gene analysis confirmed the hypothesis that the binding potential of miR-13 is inversely correlated with gene expression.
Boutla
Alexandra
A
Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology, Hellas, PO Box 1527, GR-71110 Heraklion/Crete, Greece.
Delidakis
Christos
C
Tabler
Martin
M
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D016373:DNA, Antisense / Q000008:administration & dosage / Q000235:genetics*; D004330:Drosophila / Q000166:cytology / Q000196:embryology / Q000235:genetics*; D004625:Embryo, Nonmammalian / Q000002:abnormalities / Q000187:drug effects / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D017344:Genes, Insect / Q000235:genetics*; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics*; D009154:Mutation; D016376:Oligonucleotides, Antisense / Q000008:administration & dosage / Q000235:genetics; D014176:Protein Biosynthesis; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D016373:DNA, Antisense; D035683:MicroRNAs; D016376:Oligonucleotides, Antisense; D011993:Recombinant Fusion Proteins; D008156:Luciferases