MicroRNAs (miRNAs) are a class of small non-coding RNA molecules that can play critical roles as regulators of numerous pathways and biological processes including the immune response. Emerging as ...one of the most important miRNAs to orchestrate immune and inflammatory signaling, often through its recognized target genes, IRAK1 and TRAF6, is microRNA-146a (miR-146a). MiR-146a is one, of a small number of miRNAs, whose expression is strongly induced following challenge of cells with bacterial endotoxin, and prolonged expression has been linked to immune tolerance, implying that it acts as a fine-tuning mechanism to prevent an overstimulation of the inflammatory response. In other cells, miR-146a has been shown to play a role in the control of the differentiation of megakaryocytic and monocytic lineages, adaptive immunity, and cancer. In this review, we discuss the central role prescribed to miR-146a in innate immunity. We particularly focus on the role played by miR-146a in the regulation and signaling mediated by one of the main pattern recognition receptors, toll/IL-1 receptors (TLRs). Additionally, we also discuss the role of miR-146a in several classes of autoimmune pathologies where this miRNA has been shown to be dysregulated, as well as its potential role in the pathobiology of neurodegenerative diseases.
Increasing evidence supports the involvement of microRNAs (miRNAs) in inflammatory and immune processes in prion neuropathogenesis. MiRNAs are small, non-coding RNA molecules which are emerging as ...key regulators of numerous cellular processes. We established miR-146a over-expression in prion-infected mouse brain tissues concurrent with the onset of prion deposition and appearance of activated microglia. Expression profiling of a variety of central nervous system derived cell-lines revealed that miR-146a is preferentially expressed in cells of microglial lineage. Prominent up-regulation of miR-146a was evident in the microglial cell lines BV-2 following TLR2 or TLR4 activation and also EOC 13.31 via TLR2 that reached a maximum 24-48 hours post-stimulation, concomitant with the return to basal levels of transcription of induced cytokines. Gain- and loss-of-function studies with miR-146a revealed a substantial deregulation of inflammatory response pathways in response to TLR2 stimulation. Significant transcriptional alterations in response to miR-146a perturbation included downstream mediators of the pro-inflammatory transcription factor, nuclear factor-kappa B (NF-κB) and the JAK-STAT signaling pathway. Microarray analysis also predicts a role for miR-146a regulation of morphological changes in microglial activation states as well as phagocytic mediators of the oxidative burst such as CYBA and NOS3. Based on our results, we propose a role for miR-146a as a potent modulator of microglial function by regulating the activation state during prion induced neurodegeneration.
The mechanisms that lead to neuronal death in neurodegenerative diseases are poorly understood. Prion diseases, like many more common disorders such as Alzheimer's and Parkinson's diseases, are ...characterized by the progressive accumulation of misfolded disease-specific proteins. The earliest changes observed in brain tissue include a reduction in synaptic number and retraction of dendritic spines, followed by reduced length and branching of neurites. These pathologies are observable during presymptomatic stages of disease and are accompanied by altered expression of transcripts that include miRNAs. Here we report that miR-16 localized within hippocampal CA1 neurons is increased during early prion disease. Modulating miR-16 expression in mature murine hippocampal neurons by expression from a lentivirus, thus mimicking the modest increase seen in vivo, was found to induce neurodegeneration. This was characterized by retraction of neurites and reduced branching. We performed immunoprecipitation of the miR-16 enriched RISC complex, and identified associated transcripts from the co-immunoprecipitated RNA (Ago2 RIP-Chip). These transcripts were enriched with predicted binding sites for miR-16, including the validated miR-16 targets APP and BCL2, as well as numerous novel targets. In particular, genes within the neurotrophin receptor mediated MAPK/ERK pathway were potentially regulated by miR-16; including TrkB (NTRK2), MEK1 (MAP2K1) and c-Raf (RAF). Increased miR-16 expression in neurons during presymptomatic prion disease and reduction in proteins involved in MAPK/ERK signaling represents a possible mechanism by which neurite length and branching are decreased during early stages of disease.
•Upregulation of miR-16 during preclinical prion disease accompanies reduction in synapses, neurite length and branching•Overexpression of miR-16 in hippocampal neurons results in enrichment of miR-16 target transcripts in the RISC complex•Network analysis suggests miR-16 targets are involved in the neurotrophin receptor-mediated MAPK/ERK pathway•Overexpression of miR-16 leads to retraction of neurites and reduced branching in cultures of hippocampal neurons
MicroRNAs (miRNAs) are small, non-coding RNA molecules which are emerging as key regulators of numerous cellular processes. Compelling evidence links miRNAs to the control of neuronal development and ...differentiation, however, little is known about their role in neurodegeneration. We used microarrays and RT-PCR to profile miRNA expression changes in the brains of mice infected with mouse-adapted scrapie. We determined 15 miRNAs were de-regulated during the disease processes; miR-342-3p, miR-320, let-7b, miR-328, miR-128, miR-139-5p and miR-146a were over 2.5 fold up-regulated and miR-338-3p and miR-337-3p over 2.5 fold down-regulated. Only one of these miRNAs, miR-128, has previously been shown to be de-regulated in neurodegenerative disease. De-regulation of a unique subset of miRNAs suggests a conserved, disease-specific pattern of differentially expressed miRNAs is associated with prion-induced neurodegeneration. Computational analysis predicted numerous potential gene targets of these miRNAs, including 119 genes previously determined to be also de-regulated in mouse scrapie. We used a co-ordinated approach to integrate miRNA and mRNA profiling, bioinformatic predictions and biochemical validation to determine miRNA regulated processes and genes potentially involved in disease progression. In particular, a correlation between miRNA expression and putative gene targets involved in intracellular protein-degradation pathways and signaling pathways related to cell death, synapse function and neurogenesis was identified.
Highlights ► miRNAs are particularly plentiful in neurons, where they fulfill important roles in the development and plasticity of neural circuits. ► A subset of synaptic microRNAs controls the ...morphogenesis of dendrites and spines by regulating the local synthesis of important synaptic proteins. ► microRNA expression and function itself is subject to a plethora of activity-dependent regulatory mechanisms. ► First examples of a physiological role of microRNAs in learning and memory in both invertebrate and vertebrate model organisms are emerging. ► Deregulated microRNA expression might contribute to neurodevelopmental and neuropsychiatric disorders, thereby representing a novel target for therapeutic intervention.
MicroRNAs (miRNA) are a novel class of small, non-coding, gene regulatory RNA molecules that have diverse roles in a variety of eukaryotic biological processes. High-throughput detection and ...differential expression analysis of these molecules, by microarray technology, may contribute to a greater understanding of the many biological events regulated by these molecules. In this investigation we compared two different methodologies for the preparation of labelled miRNAs from mouse CNS tissue for microarray analysis. Labelled miRNAs were prepared either by a procedure involving linear amplification of miRNAs (labelled-aRNA) or using a direct labelling strategy (labelled-cDNA) and analysed using a custom miRNA microarray platform. Our aim was to develop a rapid, sensitive methodology to profile miRNAs that could be adapted for use on limited amounts of tissue.
We demonstrate the detection of an equivalent set of miRNAs from mouse CNS tissues using both amplified and non-amplified labelled miRNAs. Validation of the expression of these miRNAs in the CNS by multiplex real-time PCR confirmed the reliability of our microarray platform. We found that although the amplification step increased the sensitivity of detection of miRNAs, we observed a concomitant decrease in specificity for closely related probes, as well as increased variation introduced by dye bias.
The data presented in this investigation identifies several important sources of systematic bias that must be considered upon linear amplification of miRNA for microarray analysis in comparison to directly labelled miRNA.
Discrepancy in synaptic structural plasticity is one of the earliest manifestations of the neurodegenerative state. In prion diseases, a reduction in synapses and dendritic spine densities is ...observed during preclinical disease in neurons of the cortex and hippocampus. The underlying molecular mechanisms of these alterations have not been identified but microRNAs (miRNAs), many of which are enriched at the synapse, likely regulate local protein synthesis in rapid response to stressors such as replicating prions. MiRNAs are therefore candidate regulators of these early neurodegenerative changes and may provide clues as to the molecular pathways involved. We therefore determined changes in mature miRNA abundance within synaptoneurosomes isolated from prion-infected, as compared to mock-infected animals, at asymptomatic and symptomatic stages of disease. During preclinical disease, miRNAs that are enriched in neurons including miR-124a-3p, miR-136-5p and miR-376a-3p were elevated. At later stages of disease we found increases in miRNAs that have previously been identified as deregulated in brain tissues of prion infected mice, as well as in Alzheimer's disease (AD) models. These include miR-146a-5p, miR-142-3p, miR-143-3p, miR-145a-5p, miR-451a, miR-let-7b, miR-320 and miR-150-5p. A number of miRNAs also decreased in abundance during clinical disease. These included almost all members of the related miR-200 family (miR-200a-3p, miR-200b-3p, miR-200c-3p, miR-141-3p, and miR-429-3p) and the 182 cluster (miR-182-5p and miR-183-5p).
•MicroRNAs have altered abundance in synaptoneurosomes during prion disease.•Temporally distinct changes accompany preclinical and clinical stages of disease.•Preclinical disease exhibits increases in miR-124, miR-136, and miR-376.•Altered miR-146a, miR-142 and miR-200 family members accompany clinical disease.•Correlation with the synaptoneurosome proteome suggests key synaptic targets.
Abstract Due to the complex architecture of the brain, the precise regulation of the numerous genes and signalling molecules involved is paramount. A recently identified class of master regulatory ...molecules, known as microRNAs (miRNAs), have the potential to assist in the countless regulatory mechanisms that occur in the brain during neuronal development and function. In the process, these molecules have gained the ability to provide a very pervasive and potent layer of genetic control. MiRNAs, in general, are genome encoded, short, non-protein coding RNA molecules that are involved in gene regulation by targeting for translational repression and/or degradation large numbers of mRNA molecules simultaneously. While the brain is replete with miRNAs, their particular role(s) in the developmental and functional programs of neurons is just emerging. Additionally, dysfunction of these molecules may also contribute to the etiology of several neurodegenerative conditions. Therefore, the central aim of this review is to highlight recent findings in the field of miRNAs in neuronal development, function and dysfunction.
ABSTRACT
The involvement of SNPs in miRNA target sites remains poorly investigated in neurodegenerative disease. In addition to associations with disease risk, such genetic variations can also ...provide novel insight into mechanistic pathways that may be responsible for disease etiology and/or pathobiology. To identify SNPs associated specifically with degenerating neurons, we restricted our analysis to genes that are dysregulated in CA1 hippocampal neurons of mice during early, preclinical phase of Prion disease. The 125 genes chosen are also implicated in other numerous degenerative and neurological diseases and disorders and are therefore likely to be of fundamental importance. We predicted those SNPs that could increase, decrease, or have neutral effects on miRNA binding. This group of genes was more likely to possess DNA variants than were genes chosen at random. Furthermore, many of the SNPs are common within the human population, and could contribute to the growing awareness that miRNAs and associated SNPs could account for detrimental neurological states. Interestingly, SNPs that overlapped miRNA‐binding sites in the 3′‐UTR of GABA‐receptor subunit coding genes were particularly enriched. Moreover, we demonstrated that SNP rs9291296 would strengthen miR‐26a‐5p binding to a highly conserved site in the 3′‐UTR of gamma‐aminobutyric acid receptor subunit alpha‐4.
We identified numerous SNPs overlapping miRNA‐binding sites within an assortment of genes implicated in the very early mechanisms of neurodegeneration characterized by Prion disease. Many of these SNPs are functional as they alter the binding potential of spatiotemporally coexpressed miRNAs. Additionally, we also show that these SNPs are common within the human population. Taken together, our data contribute to the emerging awareness that miRNAs and associated SNPs could account for detrimental neurological states that afflict humans.
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