In mammalian cells,
microRNAs (miRNAs) are incorporated into miRNA-induced silencing complexes (miRISCs), which regulate
protein expression post-transcriptionally through binding to 3'-untranslated regions of target
mRNAs. Argonaute2 (Ago2), a key component of the miRISC, recruits GW182, a component of the processing body (GW/P-body), to the target
mRNAs. To elucidate the function of GW182 in an miRNA-mediated translational repression, we analyzed Argonaute-binding sites in GW182. We found that human GW182 contains three
binding sites for Ago2, within the
amino-terminal glycine tryptophan (GW/WG)-repeated region that is characteristic of the GW182 family proteins. We also found that the first and second Ago2-binding site is conserved within the
amino-terminal half of
TNRC6B, which is a
paralog of GW182. Each of the Ago-binding sites is alone sufficient to bind Ago2. Furthermore, we demonstrated that multiple
Argonaute proteins were connected via the GW182 protein. A GW182 fragment containing the Ago2-binding region partially relieved let-7-mediated repression of
protein synthesis in a mammalian
cell-free system. Coincidentally, let-7-directed target
mRNA deadenylation was delayed. Together, these results strongly suggested that the interactions of GW182 with
Argonautes may induce the formation of large complexes containing miRNA target
mRNAs, and may be critical for miRNA-mediated translational repression.
