miR-137 and sleep

The evolutionarily conserved miRNA-137 targets the neuropeptide hypocretin/orexin and modulates the wake to sleep ratio.

Holm et al. PNAS 2022

Overview

- This study investigates the intracellular mechanisms regulating the neuropeptide hypocretin (Hcrt), also known as orexin, which is essential for sleep–wake stability in vertebrates.

- The research aims to identify microRNAs (miRNAs), particularly miR-137, that target Hcrt mRNA and modulate its expression, influencing sleep and wakefulness.

- The significance lies in understanding posttranscriptional regulation of Hcrt, which has implications for sleep disorders such as narcolepsy and insomnia, and potential therapeutic avenues.

Key Concepts

Hypocretin/Orexin System: A hypothalamic neuropeptide signaling pathway critical for maintaining wakefulness; deficits cause narcolepsy.

MicroRNAs (miRNAs): Small noncoding RNAs (~20–25 nucleotides) that regulate gene expression by binding to target mRNAs, leading to degradation or translational repression.

Evolutionary Conservation: The study emphasizes that miR-137's interaction with Hcrt is conserved across species (mice, humans, zebrafish), indicating a fundamental biological role.

Conclusions

- The study uncovers a conserved posttranscriptional regulatory mechanism where miR-137 modulates Hcrt expression, thereby influencing sleep–wake states.

- Manipulating miR-137 levels affects wakefulness and sleep architecture across species, highlighting its potential as a therapeutic target.

- Human genetic data link MIR137 variants to sleep duration, suggesting relevance for individual differences in sleep patterns.

- These findings deepen understanding of the molecular regulation of sleep and open avenues for novel treatments targeting miRNA pathways.

Methodology

In Silico Prediction: Used algorithms (MicroCosm, TargetScan, DIANA, PITA) to identify miRNAs targeting HCRT mRNA, predicting miR-137, miR-637, and miR-654-5p as candidates.

In Vitro Experiments: Transfected human neuroblastoma cell lines (SK-N-MC, SK-N-DZ) with precursor miRNAs; measured HCRT mRNA levels via qRT-PCR; employed luciferase reporter assays to confirm direct targeting.

In Vivo Studies: Injected miRNA mimics or anti-miRNAs intracerebroventricularly into mouse pups; used viral vectors to inhibit miR-137 specifically in Hcrt neurons; measured Hcrt expression and sleep/wake behavior.

Genetic Association Analysis: Analyzed GWAS data from UK Biobank linking MIR137 locus variants to sleep duration.

Cross-Species Validation: Used zebrafish embryos injected with anti-miR-137 morpholino to assess effects on activity and sleep patterns.

Cytokine Stimulation: Treated human neuroblastoma cells with cytokines (IL-13, TNF, IFNs) to observe effects on miR-137 levels and Hcrt expression.

Key Findings

miR-137 targets HCRT: Confirmed through computational prediction, luciferase assays, and mutational analysis.

Conservation Across Species: The interaction between miR-137 and Hcrt is conserved in mice, humans, and zebrafish.

Regulatory Role of miR-137:

- Endogenous miR-137 levels vary diurnally in mice hypothalamus.

- Downregulation of miR-137 increases Hcrt expression and promotes wakefulness.

- Conversely, overexpression reduces Hcrt levels.

Behavioral Effects:

- Inhibiting miR-137 in Hcrt neurons results in increased wakefulness, longer wake bouts, and reduced NREM sleep during early dark hours.

- Zebrafish with inhibited miR-137 show increased activity and decreased sleep.

Cytokine Influence: IL-13 induces miR-137 upregulation, leading to decreased Hcrt expression.

Genetic Association: SNPs in MIR137 locus are significantly associated with sleep duration in humans.

Limitations

- The extent of Hcrt regulation by miR-137 is partial (~20% change), unlikely to cause severe narcolepsy but may influence subtler sleep phenotypes.

- Overexpression or inhibition experiments may not fully replicate physiological conditions; compensatory mechanisms may occur over time.

- The study primarily establishes correlation and mechanistic potential but does not definitively prove causality in complex human sleep disorders.

- Genetic association data suggest correlation but do not confirm direct functional effects of MIR137 variants on sleep regulation.

- Cross-species findings (zebrafish) support conservation but require further validation in mammals.