The extinction of conditioned fear is a fundamental aspect of adaptive behavior, orchestrated by synaptic activity in the infralimbic prefrontal cortex (ILPFC). The intricate process of fear extinction memory formation involves coordinated changes in gene expression, with long noncoding RNAs (lncRNAs) emerging as pivotal regulatory molecules. While significant strides have been made in understanding the mechanisms governing fear extinction, a comprehensive comprehension of the molecular code remains elusive.
The Role of lncRNAs in Fear Extinction
Long noncoding RNAs (lncRNAs) have garnered attention for their multifaceted roles in crucial biological processes, such as gene regulation, translation, and RNA trafficking. These molecules exhibit cell-type-specific and spatiotemporal expression, with a notable enrichment in the brain.
The synaptic compartment, a hub for neural activity, has been identified as a site where certain lncRNAs accumulate in response to stimuli. A prime example is the lncRNA ADEPTR, which localizes in dendrites and modulates activity-dependent changes in synaptic plasticity, suggesting a potential role in memory regulation.
Previous Discoveries in Activity-Dependent Epigenetic Regulation
Prior research has uncovered activity-dependent epigenetic regulation associated with fear-related learning, with nuclear lncRNAs playing a modulatory role. This prompted an exploration into whether synapse-enriched lncRNAs are integral to the localized regulation of cellular processes governing fear extinction.
To address this question, researchers utilized lncRNA capture-sequencing to map the expression of synapse-enriched lncRNAs in the ILPFC of adult male C57/bl6 mice. Subsequent single-molecule tracking in live cortical neurons and a CRISPR-inspired cell-type- and synapse-specific, state-dependent RNA knockdown approach were employed to investigate the involvement of specific lncRNAs in fear extinction.
The study uncovered a critical involvement of a variant of the lncRNA Gas5 in the regulation of fear extinction memory. Gas5 was found to play a pivotal role in the trafficking of RNA granules at the synapse, influencing intrinsic neuronal excitability and contributing to the formation of fear extinction memory. This highlights the intricate and nuanced molecular mechanisms that govern the synaptic processes underlying fear extinction.
Synaptic Regulation by Gas5 in Fear Extinction Memory
The present study unveils a substantial population of synapse-associated long noncoding RNAs (lncRNAs), with a specific focus on a variant of Gas5 as a crucial regulator of fear extinction memory. This synaptic Gas5 isoform interacts with key proteins, CAPRIN1 and G3BP2, involved in translation, RNA trafficking, and RNA granule assembly at the synapse.
The study provides compelling evidence of Gas5’s involvement in intrinsic excitability and the localized disassembly of RNA granules, establishing its pivotal role in synaptic mechanisms governing fear extinction memory.
Mechanisms of lncRNA-Mediated Regulation
The observed effects suggest a mechanism by which lncRNA activity, exemplified by Gas5, influences the behavior of RNA condensates through interactions with granule proteins. The potential coordination of RNA granule trafficking and clustering by Gas5 offers insights into its role in organizing learning-induced activities of key proteins crucial for local protein translation and subsequent memory formation (see Fig. A).
Association with Ribonucleoproteins (RNPs) and RNA Granule Dynamics
Our findings align with previous research indicating that lncRNAs can associate with ribonucleoproteins (RNPs) to form RNA condensates. Specifically, the interaction between Gas5 and CAPRIN1 and G3BP2 sheds light on the involvement of these proteins in stable complexes that regulate condensate localization and dynamics. The impact of Gas5 knockdown on the mobility and trafficking of G3BP2-containing RNA granules underscores Gas5’s role as a scaffold for coordinating the clustering of RNA granules in an activity-dependent manner.
Alternative Splicing and Functional Specificity
The Gas5 lncRNA undergoes significant alternative splicing, leading to various isoforms. Our study emphasizes the functional involvement of the synaptic Gas5 variant in fear extinction memory, with intriguing brain-region-specific roles suggested by a different splice variant in the nucleus accumbens. The identification of snoRNAs within Gas5 introns and the enrichment of intron-retained Gas5 transcripts in the nucleus suggest complex regulatory mechanisms that warrant further exploration.
Pseudogenes and Unannotated lncRNAs
The study sheds light on the presence of unannotated or pseudogene-labeled synapse-enriched lncRNAs, challenging conventional notions about pseudogene functionality. The association of pseudogenic lncRNAs with LINE and SINE elements hints at potential functional roles, raising questions about the structural modules of lncRNAs and their relationship to neuronal function.
Limitations and Future Directions
While the lncRNA capture-seq approach has facilitated the identification of low-abundance lncRNAs, inherent limitations such as coverage bias and reliance on incomplete annotations pose challenges. Technical obstacles and incomplete annotations may result in oversight, emphasizing the need for increased sample input and refined probe targeting. Further research is warranted to explore the functional and mechanistic roles of individual synaptic lncRNAs, addressing potential off-target effects associated with CRISPR-based knockdowns.
Implications for Fear Extinction Mechanisms
In summary, this study significantly advances our understanding of the molecular mechanisms underlying fear extinction memory. The identification of synapse-enriched lncRNAs, particularly the Gas5 variant, offers new insights into the intricate interplay between lncRNA activity, RNA granule dynamics, and the coordination of synaptic processes critical for fear extinction. These findings provide a foundation for future studies exploring the broader landscape of synaptic lncRNAs and their roles in various learning paradigms and activity-dependent conditions.
Fig. A : Model of the proposed mechanism by which Gas5 influences synaptic activity and the formation of fear extinction memory.
Extinction learning leads to the accumulation of the Gas5 variant in the synaptic compartment, which then sequesters CAPRIN1 and G3BP2 containing RNA granules away from clustering, leading to an increase in local protein synthesis and tighter control over synaptic plasticity.
reference link : https://www.nature.com/articles/s41467-023-43535-1