Human Argonaute2 to form minimal RISC
Pyrococcus furiosus (archaea)
RNA interference (RNAi) denotes a gene silencing mechanism by double stranded RNAs. If double stranded RNAs exist in a cell, they are processed to be small interfering RNAs (siRNAs), which are ~22-nucleotide double stranded RNAs, through the action of Dicer protein. The siRNAs enter the RNAi effector complex (RISC), and serve as a guide to selection of silencing targets based on sequence complementarity. RISC can silence its target by RNA cleavage, by preventing protein synthesis or by triggering chromatin remodeling and consequent transcriptional repression. The component of RISC is not well-known, however, a small RNA guide and an RNA slicer, Argonaute family, are considered to be the core element.
The protein described here is an Argonaute protein from Pyrococcus furiosus (PfAgo). Argonaute proteins have N-terminal, middle, PIWI and PAZ domains. The N-terminal, middle and PIWI domains form a crescent-shaped base which holds above a stalk-like the PAZ domain. An siRNA is accommodated within the groove of the PAZ domain and the base. The 5' end of the siRNA interacts with the PIWI domain, whereas its 3' end interacts with the PAZ domain. The PIWI domain is considered to have an RNase active site, because a divalent cation binding site, DDH motif conserved in the RNase H family, exists. The RNA cleavage of RNase H family results in the formation of a 5' product with 3' hydroxyl and a 3' product with 5' phosphate. Furthermore, PIWI domain has relatively conserved region where several basic amino acid residues (K or R) locate (known as PIWI box). PIWI box is considered as the binding site of 5' phosphate of the siRNA. This 5' phosphate-PIWI box interaction is very important because that enhances the fidelity of specific RNA cleavage of RISC. Although Argonaute protein itself lacks ATP-assisted rapid product release mechanism (this may be carried out by another factor), Argonaute-siRNA complex is able to cleave substrate RNAs accurately. Therefore, Argonaute-siRNA complex can be considered as a minimal RISC.
Protein Data Bank (PDB)
author: Daisuke Ino