Akt-Mediated Phosphorylation of Argonaute 2 Downregulates Cleavage and Upregulates Translational Repression of MicroRNA Targets

Abstract

SUMMARY A high-throughput RNA interference (RNAi) screen targeting 542 genes of the human kinome was used to discover regulators of RNAi. Here we report that the proto-oncogene Akt-3/PKBγ (Akt3) phosphorylates Argonaute 2 (Ago2) at Ser387 which down-regulates cleavage and up-regulates translational repression of endogenous microRNA (miRNA)-targeted mRNAs. We further demonstrate that Akt3 co-immunoprecipitates with Ago2 and that phosphorylation of Ago2 at Ser387 facilitates its interaction with GW182 and localization to cytoplasmic P-bodies, where miRNA-targeted mRNAs are thought to be stored and degraded. Therefore, Akt3-mediated phosphorylation of Ago2 is a molecular switch between target mRNA cleavage and translational repression activities of Ago2.

NMDAR ‐dependent Argonaute 2 phosphorylation regulates mi RNA activity and dendritic spine plasticity

“…In agreement with our results, a previous study demonstrated that S387 phosphorylation in HeLa cells increased Ago2‐GW182 binding and had no effect on miRNA loading (Horman et al , 2013). While S387 is situated between the PAZ and MID domains of Ago2, GW182 associates with the PIWI domain towards the C‐terminus of Ago2 (Pfaff et al , 2013), suggesting that phosphorylated S387 is unlikely to contribute to the GW182 binding site directly.…”

“…Previous reports suggest the Ago2‐GW182 interaction is regulated in HeLa cells by phosphorylation of Ago2 at S387 by the kinase Akt (Horman et al , 2013; Bridge et al , 2017). We therefore hypothesised that the NMDAR‐stimulated increase in binding would be mediated by a similar mechanism.…”

“…Importantly, Ago2 can be phosphorylated at a number of residues, some of which have been suggested to regulate RISC activity in non‐neuronal cell lines by controlling Ago2‐RNA or Ago2‐protein interactions (Jee & Lai, 2014). Phosphorylation of Ago2 at serine 387 (S387) enhances its interaction with GW182 and increases miRNA‐mediated translational repression in HeLa cells (Horman et al , 2013). The regulation of RISC protein–protein interactions or RISC activity by Ago2 phosphorylation remains completely unexplored in neurons, and we hypothesised that regulating critical RISC protein–protein interactions via Ago2 phosphorylation is a key mechanism to mediate the rapid modulation of miRNA‐mediated gene silencing in dendrites in response to neuronal stimulation.…”

Genomically amplified Akt3 activates DNA repair pathway and promotes glioma progression

“…This study highlighted Akt1/3-mediated phosphorylation of an RNA processing regulator, which led to alternative splicing of FGFR2. Additional studies are reporting potential nuclear functions of Akt3, including Ago2-mediated translational repressive activities (28), transcriptional control of IGFBP3 in lung cancer cells (29), and ERBB2, ERBB3, and ERα in breast tumor cells from a mouse transgenic model (30). Although these effects are likely mediated by multiple complex systems, Akt3 has been reported to control the stability of the major nuclear export protein, chromosome maintenance region-1 (CRM-1), leading to altered mitochondrial biogenesis and autophagy (31).…”

The complexity of miRNA-mediated repression

“…This is likely due to enhanced interaction of phosphorylated Ago2 with GW182 and P-bodies. 116,117 Two recent studies investigated the role of Ago2 in stress response. One showed that EGFR induces phosphorylation of Y393 of Ago2 in hypoxia, which in turn inhibits miRNA maturation, possibly contributing to tumour cell survival.…”