Corticosterone-mediated regulation and functions of miR-218-5p in rat brain

Abstract

Chronic stress is one of the key precipitating factors in major depressive disorder (MDD). Stress associated studies have underscored the mechanistic role of epigenetic master players like microRNAs (miRNAs) in depression pathophysiology at both preclinical and clinical levels. Previously, we had reported changes in miR-218-5p expression in response to corticosterone (CORT) induced chronic stress. MiR-218-5p was one of the most significantly induced miRNAs in the prefrontal cortex (PFC) of rats under chronic stress. In the present report, we have investigated how chronic CORT exposure mechanistically affected miR-218-5p expression in the rat brain and how miR-218 could trigger molecular changes on its downstream regulatory pathways. Elevated expression of miR-218-5p was found in the PFC of CORT-treated rats. A glucocorticoid receptor (GR) targeted Chromatin-Immunoprecipitation (ChIP) assay revealed high GR occupancy on the promoter region of Slit3 gene hosting miR-218-2 in its 3rd intron. RNA-sequencing data based on RNA Induced silencing Complex Immunoprecipitation (RISC-IP) with AGO2 in SH-SY5Y cells detected six consistent target genes of miR-218-5p (APOL4, DTWD1, BNIP1, METTL22, SNAPC1, and HDAC6). The expression of all five genes, except APOL4, was successfully validated with qPCR in CORT-treated rat PFC. Further, Hdac6-based ChIP-seq experiment helped in mapping major genomic loci enriched for intergenic regions in the PFC of CORT-treated rat. A proximity-based gene ontology (GO) analysis revealed a majority of the intergenic sites to be part of key genes implicated in central nervous system functions, notably synapse organization, neuron projection morphogenesis, and axonogenesis. Our results suggest that the upregulation of miR-218-5p in PFC of CORT-treated rats possibly resulted from GR biding in the promoter region of Slit3 gene. Interestingly, Hdac6 was one of the consistent target genes potentially found to regulate CNS related genes by chromatin modification. Collectively, these findings establish the role of miR-218-5p in chronic stress and the epigenetic function it plays to induce chromatin-based transcriptional changes of several CNS genes in triggering stress-induced disorders, including depression. This also opens up the scope to understand the role of miR-218-5p as a potential target for noncoding RNA therapeutics in clinical depression.

miR-218: A Stress-Responsive Epigenetic Modifier

“…It has earlier been shown that the other member of the SLIT gene family, i.e., SLIT2, harbors intronic miR-218-1, which encodes the other isoform of the miR-218 family (miR-218-1-3p). It is interesting to highlight that both isoforms are rooted in the Slit-Robo signaling axis with a functional role in depression-/anxiety-like behaviors as previously reported in adult mice [ 33 ]. The secreted Slit ligands and their Robo receptors constitute a signaling pathway that controls the directed migration of neurons and vascular endothelial cells during embryonic development.…”

“…The characteristic stem-loop structure of precursor miR-218 has been shown on Figure 2 a. The transcriptional origin of miR-218 has been mapped on two separate loci, miR-218-1 and miR-218-2 and found to be located at the introns of SLIT2 and SLIT3 genes, respectively [ 33 ]. More specifically, miR-218-1 is located within intron 15 of the SLIT2 gene, and mir-218-2 is present within intron 14 of the SLIT3 gene.…”

“…Multiple studies have explored the role of miR-218 in regulating stress response pertaining to depression [ 28 , 33 , 89 ]. A very recent study by our lab has made a significant contribution to understanding the role of this miRNA in tethering hundreds of key genes ( Figure 3 ) implicated in central nervous system functions, notably synapse organization, neuron-projection morphogenesis, and axonogenesis ( Figure 4 a,b) [ 33 ]. The study highlighted the role that miR-218-5p plays in chronic stress and how it can contribute to acquiring maladaptive changes in the MDD brain through epigenetic dysfunctions.…”

“…These genes have earlier been associated with stress and depression pathophysiology [ 104 ]. A follow-up study published in 2022 further explored how CORT-treatment mechanistically affected miR-218-5p expression and how miR-218 could trigger molecular changes in its downstream regulatory pathways [ 33 ]. A GR-targeted chromatin immunoprecipitation (ChIP) assay revealed high GR occupancy on the promoter region of the SLIT3 gene hosting miR-218-2 in its third intron.…”

“…Increasing focus on miR-218 has been associated with various brain functions, including synaptic plasticity, cell survival, and behavior [ 29 , 30 ]. Our group and other investigators have shown that this miRNA is involved in stress and susceptibility to developing depression and can be used as a biomarker for diagnosing stress-related disorders [ 31 , 32 , 33 ]. In this review, we have provided an overview of the importance of studying miRNAs in brain functions and how they are epigenetically connected with various neuropsychiatric abnormalities, especially depression and other stress-related mental disorders.…”