The RNA-induced silencing complex: a versatile gene-silencing machine

Abstract

RNA interference is a powerful mechanism of gene silencing that underlies many aspects of eukaryotic biology. On the molecular level, RNA interference is mediated by a family of ribonucleoprotein complexes called RNA-induced silencing complexes (RISCs), which can be programmed to target virtually any nucleic acid sequence for silencing. The ability of RISC to locate target RNAs has been co-opted by evolution many times to generate a broad spectrum of gene-silencing pathways. Here, we review the fundamental biochemical and biophysical properties of RISC that facilitate gene targeting and describe the various mechanisms of gene silencing known to exploit RISC activity.

FIGURE 1.

Formation of RISCs and other silencing complexes. Silencing RNA can be derived from exogenous or intracellular origins, depending on the organism and cell type. RNA can also be introduced artificially using siRNA or plasmid-based short hairpin RNA (shRNA) systems. RNAs transcribed from the genome may be retained in the nucleus (as with piRNAs) to carry out silencing or may be exported (as with miRNAs). In the cytoplasm, dsRNA is processed by the endonuclease Dicer and loaded onto an Argonaute protein, and after the strand selection process, the newly formed RISC is equipped to silence target genes by one of several mechanisms.

FIGURE 2.

Structure of a prokaryotic Argonaute with bound guide and target. a, crystal structures of Thermus thermophilus Argonaute bound to a 10-mer DNA (left; DNA not shown to depict “apo”-Argonaute), a 21-mer guide DNA (center), or a 21-mer guide DNA with a 20-mer target RNA (right) illustrate Argonaute fold and function. The 5′-end of the guide DNA contacts the middle (Mid; dark green) and PIWI (light green) domains, and the 3′-end binds the N-terminal (blue) and PAZ (purple) domains. b, shown is a schematic representation of Argonaute domains and regions of interaction between the protein and guide strand (dashed lines). The asterisk denotes the location of slicer cleavage.

FIGURE 3.

RISC effector processes. Once bound to its target RNA, RISC may down-regulate gene expression by one of several mechanisms, depending on the type of Argonaute and cellular context. In the cytoplasm, mRNA targets can be cleaved via RISC slicer activity or translationally repressed. In the nucleus, RISC can take the form of an RITS complex, which interacts with RNA polymerase II (PolII) and nascent RNA transcripts and directs chromatin remodeling to achieve epigenetic silencing.