NMDA receptor-dependent regulation of miRNA expression and association with Argonaute during LTP in vivo

Abstract

microRNAs (miRNAs) are major regulators of protein synthesis in the brain. A major goal is to identify changes in miRNA expression underlying protein synthesis-dependent forms of synaptic plasticity such as long-term potentiation (LTP). Previous analyses focused on changes in miRNA levels in total lysate samples. Here, we asked whether changes in total miRNA accurately reflect changes in the amount of miRNA bound to Argonaute protein within the miRNA-induced silencing complex (miRISC). Ago2 immunoprecipitation was used to isolate RISC-associated miRNAs following high-frequency stimulation (HFS)-induced LTP in the dentate gyrus of anesthetized rats. Using locked-nucleic acid-based PCR cards for high-throughput screening and independent validation by quantitative TaqMan RT-PCR, we identified differential regulation of Ago2-associated and total miRNA expression. The ratio of Ago2/total miRNA expression was regulated bidirectionally in a miRNA-specific manner and was largely dependent on N-methyl-D-aspartate receptor (NMDA) activation during LTP induction. The present results identify miRNA association with Ago2 as a potential control point in activity-dependent synaptic plasticity in the adult brain. Finally, novel computational analysis for targets of the Ago2-associated miRNAs identifies 21 pathways that are enriched and differentially targeted by the miRNAs including axon guidance, mTOR, MAPK, Ras, and LTP.

NMDA receptor–dependent dephosphorylation of serine 387 in Argonaute 2 increases its degradation and affects dendritic spine density and maturation

“…Reducing Ago2 phosphorylation at Ser-387 has been shown to decrease miRNA-induced repression of mRNAs (46). Furthermore, several research groups reported that miRNA-dependent translational regulation of local protein expression is associated with synaptogenesis and synaptic plasticity (22)(23)(24)(25)(26)(27). Hence, we hypothesized that the NMDA-R-dependent dephosphorylation of Ago2 Ser-387 could be involved in regulating the release of mRNA repression and thereby allowing the local translation of mRNAs in dendrites and spines.…”

“…The regulation of mRNA translation by miRNAs occurs in the miRNA-induced silencing complex (miRISC), which enables miRNAs to target their mRNAs (12,14,20,21). Several studies have shown that the association of miRISC with different scaffold or RNA-binding proteins underlies the translational regulation associated with synaptogenesis and synaptic plasticity (22)(23)(24)(25)(26)(27).…”

MicroRNA-30c-5p modulates neuropathic pain in rodents

“…The recruitment of miR-30c-5p to the miRNA-induced silencing complexes is enhanced in the spinal cord after NI in rats We analyzed whether spinal overexpression of miR-30c-5p after NI is associated with a parallel increase in miR-30c-5p bound to Argonaute2 (Ago2) protein in the miRNA-induced silencing complexes (miRISCs), which would reflect the functional impact of miR-30c-5p overexpression on the translation of its targeted mRNAs (7). To this end, Ago2 was immunoprecipitated with a specific antibody (Ab) in SDH lysates from NI and sham rats (32). As shown in fig fig.…”

Expression of NMDA receptor and microRNA-219 in rats submitted to cerebral ischemia associated with alcoholism

“…Among them, Pai et al, who, after silencing Ago2 protein and RISC complex in rat long-term potentiation found by real-time PCR, discovered that the regulation of expression levels of several miRNAs, including the miR-219, was totally dependent on the NMDA receptor activation during long-term potentiation 13 . In another study, Kocerba et al analyzed the expression of miR-219, a brain-specific miRNA, after pharmacological intervention (administration of dizocilpine) or genetic disruption (NR1 hypomorphism).…”