title: Properties of N(4)-methylated cytidines in miRNA mimics.
abstract: Experiments conducted with micro RNA (miRNA) mimics often result in subtle phenotypic changes and hence require careful controls. A commonly used type of control reagent in the antisense/RNA interference fields is the mismatched sequence. However, it is difficult to use mismatch controls for miRNAs, mainly because base permutation in the seed region may generate a new miRNA seed with its own associated target transcripts. We incorporated N(4)-methylcytidine and N(4),N(4)-dimethylcytidine into a series of RNAs using the convertible nucleoside approach and measured their effects on hybridization affinity with complementary RNAs, and on miRNA-mediated and small interfering RNA (SiRNA)-mediated silencing. We report here that incorporation of a single N(4),N(4)-dimethylcytidine into the seed region of miRNAs can be used as a new class of negative miRNA control which (1) does not constitute a new seed sequence; (2) is accepted by the RNA-induced silencing complex (RISC); (3) causes a significant loss of binding affinity to target RNAs; and (4) is synthesized conveniently into oligoribonucleotides.
authors.affiliation: Institute of Pharmaceutical Sciences, Department of Chemistry and Applied Biosciences, ETH Zurich, Zurich, Switzerland.
authors.forename: Boris
authors.initials: B
authors.lastname: Guennewig
authors.forename: Moritz
authors.initials: M
authors.lastname: Stoltz
authors.forename: Mirjam
authors.initials: M
authors.lastname: Menzi
authors.forename: Afzal M
authors.initials: AM
authors.lastname: Dogar
authors.forename: Jonathan
authors.initials: J
authors.lastname: Hall
mesh_terms: D020413:3' Untranslated Regions; D020029:Base Pairing; D001483:Base Sequence; D003562:Cytidine / Q000031:analogs & derivatives* / Q000737:chemistry*; D055785:Gene Knockdown Techniques; D017930:Genes, Reporter; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D035683:MicroRNAs / Q000737:chemistry / Q000235:genetics*; D009843:Oligoribonucleotides / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics; D034622:RNA Interference; D034743:RNA-Induced Silencing Complex; D044366:Transition Temperature
chemical_list: D020413:3' Untranslated Regions; C542541:MIRN106 microRNA, human; C522425:MIRN34 microRNA, human; D035683:MicroRNAs; D009843:Oligoribonucleotides; D034743:RNA-Induced Silencing Complex; D003562:Cytidine; D008156:Luciferases