Abstract
MicroRNAs (miRNAs) are important regulators of localized mRNA translation in neuronal dendrites. The presence of RNA-induced silencing complex proteins in these compartments and the dynamic miRNA expression changes that occur in response to neuronal stimulation highlight their importance in synaptic plasticity. Previously, we demonstrated a novel interaction between the major RNA-induced silencing complex component Argounaute-2 (Ago2) and the BAR (bin/amphiphysin/rvs) domain protein PICK1. PICK1 recruits Ago2 to recycling endosomes in dendrites, where it inhibits miRNA-mediated translational repression. Chemical induction of long-term depression via NMDA receptor activation causes the dissociation of Ago2 from PICK1 and a consequent increase in dendritic miRNA-mediated gene silencing. The mechanism that underlies the regulation of PICK1-Ago2 binding is unknown. In this study, we demonstrate that the PICK1-Ago2 interaction is directly sensitive to Ca2+ ions so that high [Ca2+]free reduces PICK1 binding to Ago2. Mutating a stretch of C-terminal Ca2+-binding residues in PICK1 results in a complete block of NMDA-induced PICK1-Ago2 disassociation in cortical neurons. Furthermore, the same mutant also blocks NMDA-stimulated miRNA-mediated gene silencing. This study defines a novel mechanism whereby elevated [Ca2+] induced by NMDA receptor activation modulates Ago2 and miRNA activity via PICK1. Our work suggests a Ca2+-dependent process to regulate miRNA activity in neurons in response to the induction of long-term depression.
Highlights
MicroRNAs are important regulators of localized mRNA translation in neuronal dendrites
Because the PICK1-Ago2 interaction is reduced upon NMDA receptor (NMDAR)-mediated chemically induced LTD (cLTD) [16], we investigated whether Ca2ϩ modulates PICK1-Ago2 binding
We showed previously that PICK1 inhibits Ago2 function in neuronal dendrites and that, during cLTD, Ago2 is released from PICK1, causing an increase in miRNA-mediated translational repression [16]
Summary
PICK1 has been identified previously as a Ca2ϩ sensor that interacts with the AMPAR subunit GluA2 in a Ca2ϩ-dependent manner [14]. CLTD caused a similar increase in GluA2 interaction with both PICK1 WT and the Aϫ10 mutant (Fig. 5, A and C) These data indicate that the Ca2ϩ-sensing property of the PICK1 C-terminal acidic region underlies the release of Ago from PICK1 following cLTD induction but that it is not involved in regulating the interaction with AMPARs. PICK1 localizes to endosomal compartments [20], and we showed previously that Ago is present with PICK1 on recycling endosomes and that Ago dissociates from PICK1-containing compartments in response to cLTD induction [16]. With GFP-PICK1 A10 did not alter Let activity, as measured using the LIN41 luciferase reporter under basal or cLTD conditions (Fig. 9, E and F) Taken together, these experiments suggest that the C-terminal Ca2ϩ-binding region of PICK1 regulates dendritic miRNA-mediated gene silencing events in response to cLTD induction
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