A nuclear role for ARGONAUTE-2 in regulation of neuronal alternative polyadenylation

Abstract

Argonaute 2 (AGO2), the effector protein partner of microRNAs (miRNAs) in the cytoplasmic RNA induced silencing complex, is further involved in nuclear RNA processing. However, a role for AGO2 in regulation of alternative polyadenylation was not yet demonstrated. Here, we reveal unexpected abundance of AGO2 in mouse neuronal nuclei and characterize nuclear AGO2 interactors by mass spectrometry. We discover that AGO2 broadly regulated alternative polyadenylation (APA) in neuronal cells. Specifically, we demonstrate how two isoforms of Ret mRNA, which encodes a receptor tyrosine kinase are regulated by AGO2-depenent APA, affecting downstream GDNF signaling in primary motor neurons.

Widespread association of the Argonaute protein AGO2 with meiotic chromatin suggests a distinct nuclear function in mammalian male reproduction

“…We isolated nuclear and cytoplasmic fractions from each population and confirmed AGO2 protein localization in both the nucleus and cytoplasm of meiotic and postmeiotic male germ cells, in agreement with previous observations in other mammalian tissues ( Fig. 1 A; Kim et al 2011 ; Gagnon et al 2014 ; Ravid et al 2020 ; Chu et al 2021 ), suggesting that AGO2 can function in both the nucleus and cytoplasm at multiple steps of spermatogenic development.…”

“…2 D). In addition, 16 of the nuclear AGO2 protein partners we identify here were also found to interact with AGO2 in nuclei of mouse neurons ( Ravid et al 2020 ). Nuclear AGO2 may target a common set of genes across multiple cell types, with tissue-specific functions imposed by transcription factor partners or additional downstream regulatory mechanisms.…”

A nuclear role for the Argonaute protein AGO2 in mammalian gametogenesis

“…We collected purified meiotic (pachytene spermatocyte) and post-meiotic (round spermatid) male germ cells from adult mouse testes (Figure S1E) and isolated nuclear and cytoplasmic fractions. We confirmed AGO2 protein expression in both nucleus and cytoplasm of meiotic and post-meiotic male germ cells, in agreement with previous observations in other mammalian tissues (Gagnon et al, 2014;Kim et al, 2011;Ravid et al, 2020) (Figure 1F), suggesting that AGO2 can function in both nucleus and cytoplasm at multiple steps of spermatogenic development.…”

“…Supporting a more generalizable role for nuclear AGO2 is the fact that many of its gene and transcript targets are involved in somatic, rather than spermatogenic, development. In addition, sixteen of the nuclear AGO2 protein partners we identify here were recently found to interact with AGO2 in nuclei of mouse neurons (Ravid et al, 2020). AGO2 may therefore regulate a common set of genes across multiple cell types, with tissue-specific functions imposed by additional downstream regulatory mechanisms.…”