Book of Abstracts: Albany 2003
June 17-21 2003
Small Interfering RNAs and MicroRNAs in Animal Systems
RNA interference (RNAi) represents an evolutionary conserved cellular defense mechanism for controlling the expression of alien genes in protists, filamentous fungi, plants, animals and humans. RNAi is triggered by double-stranded RNA (dsRNA) and causes sequence-specific degradation of homologous mRNAs. The mediators of target RNA cleavage are duplexes of 21-nt small interfering RNAs (siRNAs) produced by Dicer RNAse III cleavage of long dsRNAs or RNA hairpins (1). Duplexes of 21-nt siRNAs with symmetric 2-nt 3' overhangs or RNA hairpins with a single 2-nt 3'-overhang are efficient triggers of RNAi in mammalian cells siRNAs and hairpin RNAs of less than 30 base pairs do not trigger sequence-unspecific effects in mammalian cells (interferon response) and produce ?knockdown? cells with characteristic ?knockdown? phenotypes (2). We are characterizing the human ribonucleoprotein complexes involved in RNAi and recently showed the participation of the human Argonaute family members, eIF2C1 and eIF2C2, in the formation of the target RNA-cleaving RNA-induced silencing complex (RISC) (3). Active RISC contains a single-stranded antisense RNA that guides the target RNA cleavage.
Another class of small regulatory RNAs has been identified from cloning of small expressed RNAs from total RNA preparations. These microRNAs represent an extensive class of evolutionary conserved noncoding RNAs. Mature miRNAs are about 22 nucleotide in length and excised from stem-loop precursors of about 60 nucleotide in length. In contrast to siRNAs that are complementary to their target mRNAs, animal miRNAs appear to be only partially complementary and are believed to act predominantly as translational regulators. The characterization of miRNAs from Drosophila melanogaster indicated that there is a third species of small RNAs of about 25 nucleotide in length derived from double-stranded RNA of retrotransposable elements.
1Laboratory for RNA Molecular Biology