Behçet's Disease (BD) is diagnosed as continuing systematic inflammation, lacking clarified etiology and pertinent detection of clinical usage. We screened bulk-seq and single-cell seq data of BD from GEO database, then investigated immune infiltration landscape in BD and further explored main immune cells in a single-cell resolution. It was inferred that monocytes might be the trigger cell of BD occurrence and development. On the other hand, we performed differentially expression analysis and weighted correlation network analysis, and intersected the selected genes with immune-related genes from ImmPort database. After KEGG/GO-enrichment, Protein-Protein Interaction network and miRNA-mRNA regulatory network were constructed. Meanwhile, we utilized the Least Absolute Shrinkage and Selection Operator to filtrate 5 significant intersected genes for BD's diagnosis (NRTN, TRDJ1, IGLV4-69, PDIA2 and AVPR1A), then drew Operating Characteristic curve to analyze diagnosis value. For experimental validation, the BD mice models were constructed and qRT-PCR experiment was performed to validate the expression level of selected genes. In conclusion, we investigated the immune landscape in BD and explored the role of monocyte in BD occurrence and development. Through gene selection and machine learning, five new potential diagnostic biomarkers were discovered and validated in both external dataset and experiment assay levels.

Characterized by glomerulonephritis, lupus nephritis (LN) worsens the prognosis of patients with systemic lupus erythematosus (SLE) and contributes to its increased mortality rate. Here, we investigated whether serum exosomal microRNAs (miRNAs) are promising new biomarkers of LN.

Serum exosomes were initially isolated using a reagent-based kit, and total RNA was extracted with the Trizol method. Small RNA sequencing was subsequently performed to identify differentially expressed exosomal miRNAs. Validation of these miRNAs was conducted via real-time quantitative polymerase chain reaction (RT-qPCR) on individual samples from both the training and validation cohorts, leading to the identification of candidate small RNAs specifically associated with LN. Receiver operating characteristic (ROC) curve analysis was employed to evaluate the diagnostic performance of the identified candidates. To further investigate the immune landscape, 12 types of cytokines were quantified using flow cytometry, and their correlations with the candidate miRNAs were analyzed via Spearman rank correlation. Additionally, the biological functions of these miRNAs were explored through enrichment analyses based on Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways.

The levels of hsa-miR-497-5p and hsa-miR-6515-5p were significantly higher in the LN group compared to the SLE without LN group, with a combined area under the ROC curve (AUC) of 0.798. Notably, these two miRNAs demonstrated exceptional discriminative performance in identifying LN patients with mild proteinuria, achieving an AUC of 0.844. Furthermore, flow cytometry analysis revealed markedly elevated serum levels of interferon (IFN)-γ, interleukin (IL)-8, and IL-17 in the LN group compared to healthy controls (HCs). Additionally, IL-6 and IL-17 levels were significantly higher in the LN group compared to the SLE without LN group. Hsa-miR-6515-5p exhibited a strong positive correlation with IL-8 and IFN-γ. Bioinformatic analyses indicated that these exosomal miRNAs may contribute to the progression of LN by regulating key signaling pathways, with the MAPK signaling pathway being prominently implicated.

Our study demonstrated that serum exosomal miRNAs, specifically hsa-miR-497-5p and hsa-miR-6515-5p, show significant potential as biomarkers for the early detection and prognosis of LN.

Circulating monocytes largely contribute to macrophage buildup in atheromata, which is crucial for clearing subendothelial LDLs (low-density lipoproteins) and dead cells; however, the transitional trajectory from monocytes to macrophages in atherosclerotic plaques and the underlying regulatory mechanism remain unclear. Moreover, the role of alternative polyadenylation, a posttranscriptional regulator of cell fate, in monocyte/macrophage fate decisions during atherogenesis is not entirely understood.

To identify monocyte/macrophage subtypes in atherosclerotic lesions and the effect of alternative polyadenylation on these subtypes and atherogenesis, single-cell RNA sequencing, 3'-end sequencing, flow cytometric, and histopathologic analyses were performed on plaques obtained from Apoe-/- mouse arteries with or without myeloid deletion of Srsf3 (serine/arginine-rich splicing factor 3). Cell fractionation, polysome profiling, L-azidohomoalanine metabolic labeling assay, and metabolomic profiling were conducted to disclose the underlying mechanisms. Reprogramming of widespread alternative polyadenylation patterns was estimated in human plaques via bulk RNA sequencing.

We identified a subset of lesional cells in a monocyte-to-macrophage transitional state, which exhibited high expression of chemokines in mice. Srsf3 deletion caused a maturation delay of these transitional cells and phagocytic impairment of lesional macrophages, aggravating atherosclerosis. Mechanistically, Srsf3 deficiency shortened 3' untranslated regions of mitochondria-associated Aars2 (alanyl-tRNA synthetase 2), disrupting its translation. The resultant impairment of protein synthesis in mitochondria led to mitochondrial dysfunction with declined NAD+ (nicotinamide adenine dinucleotide, oxidized form) levels, activation of the integrated stress response, and metabolic reprogramming in macrophages. Administering an NAD+ precursor nicotinamide mononucleotide or the integrated stress response inhibitor partially restored Srsf3-deficient macrophage maturation, and nicotinamide mononucleotide treatment mitigated the proatherosclerotic effects of Srsf3 deficiency. Consistently, Srsf3 downregulation, global 3' untranslated region shortening, and accumulation of these transitional macrophages were associated with atherosclerosis progression in humans.

Our study reveals that Srsf3-dependent generation of long 3' untranslated region is required for efficient mitochondrial translation, which promotes mature phagocytic macrophage formation, thereby playing a protective role in atherosclerosis.

MicroRNA-seq data is produced by aligning small RNA sequencing reads of different microRNA transcript isoforms, called isomiRs, to known microRNAs. Aggregation to microRNA-level counts discards information and violates core assumptions of differential expression methods developed for mRNA-seq data. We establish miRglmm, a differential expression method for microRNA-seq data, that uses a generalized linear mixed model of isomiR-level counts, facilitating detection of miRNA with differential expression or differential isomiR usage. We demonstrate that miRglmm outperforms current differential expression methods in estimating differential expression for miRNA, whether or not there is differential isomiR usage, and simultaneously provides estimates of isomiR-level differential expression.

Activated hepatic stellate cells (HSCs) induce alternative (M2) polarization of macrophages and contribute to the progression of fibrosis and hepatocellular carcinoma (HCC). However, the effects of small extracellular vesicles released by HSCs (HSC-sEVs) during activation remain largely unknown.

The aim of this study was to investigate the role of extracellular vesicles released by HSCs (HSC-sEVs) at different stages of activation in macrophage polarization. The effects of sEVs from short-term activated and long-term activated HSCs on liver macrophages was studied. Small RNA sequencing analyses were performed to obtain differential miRNAs transported by the short-term and long-term activated HSC- sEVs. The in vivo effects of short-term activated HSC-sEV-specific miRNA on liver macrophage and liver fibrosis were confirmed in a CCl4-induced liver injury mouse model. To study the tumor suppressive effects of the macrophages educated by short-term activated HSC-sEV-specific miRNA, human hepatoma cells were mixed and subcutaneously cotransplanted with miR-99a-5p mimic-pretreated macrophages.

We found that consistent with activated HSCs, long-term activated HSC-sEVs (14dHSC-sEVs) induce bone marrow-derived monocytes (MOs) toward an M2 phenotype, but short-term activated HSC-sEVs (3dHSC-sEVs) induce the resident macrophages (Kupffer cells, KCs) toward a classically activated (M1) phenotype. We identified five 3dHSC-sEV-specific miRNAs, including miR-99a-5p. In vitro and in vivo experiments support that miR-99a-5p negatively regulates alternative polarization of macrophages, decreases collagen deposition in chronic liver injury model, and suppresses the progression of hepatoma in a xenograft model partially by targeting CD93.

Collectively, our work reveals an unexpected proinflammatory role of 3dHSC-sEVs, preliminarily explores the underlying mechanism, and evaluates the therapeutic potential of 3dHSC-sEV-specific miR-99a-5p for liver fibrosis and tumorigenesis.

20-Hydroxyecdysone (20E) is an important hormone that regulates insect development and metamorphosis. The fat body of insects plays a crucial role in nutrient storage and energy metabolism and is considered the exchange center for regulating insect development. The fat body undergoes remarkable transformation during insect metamorphosis and is primarily regulated by 20E. microRNAs (miRNAs) have been identified in different insects and have multiple functions in various physiological processes. However, the interaction of 20E and miRNAs in fat body regulation remains unclear.

We constructed six small RNA libraries using Bombyx mori fat body treated with 20E. Expression and functional analyses were conducted to identify 20E-responsive miRNAs. In total, 431 miRNAs were identified, including 389 known and 42 novel miRNAs. Differential expression analysis revealed significant expression changes in the expression of 40, 9, and 18 miRNAs at 2 h, 6 h, and 12 h after 20E treatment, respectively. The expression of 10 miRNAs was validated using quantitative real-time PCR. miR-34-5p is a highly conserved miRNA among the 10 validated miRNAs, and autophagy-related gene 1 (Atg1) was considered a target gene of miR-34-5p. The expression analysis of miR-34-5p and Atg1 exhibited an opposite expression pattern in the fat body after the 20E treatment. Dual-luciferase assay indicated that miR-34-5p could inhibit Atg1 expression by targeting a binding site in CDS region of Atg1. In larval fat body, overexpressing miR-34-5p by injecting miR-34-5p agomir suppressed the expression of Atg1 and autophagy, whereas knocking down miR-34-5p by injecting miR-34-5p antagomir induced the expression of Atg1 and autophagy. Meanwhile, Atg1 silencing by RNAi also inhibited autophagy. These results indicate that miR-34-5p participates in 20E-induced autophagy in B. mori fat body by interacting with Atg1.

We systematically identified and functionally characterized miRNAs associated with 20E regulation in the fat body of B. mori. miR-34-5p is involved in 20E-induced autophagy in B. mori by regulating its target gene Atg1. These results provide insight into the role of sophisticated interactions between miRNAs, 20E regulation, and autophagy in fat body remodeling and insect metamorphosis.

Cigarette smoke (CS) creates a "cancer field" in the lung that promotes malignant transformation. The molecular changes within this field are not fully characterized. We examined the significance of microRNA-1 (miR-1) downregulation as one of these changes. We found that tumor miR-1 levels in three non-small cell lung cancer cohorts show inverse correlations with the smoking burden. Lung MiR-1 levels follow a spatial gradient, have prognostic significance, and correlate inversely with the molecular markers of injury. In CS-exposed lungs, miR-1 is specifically downregulated in the endothelium. Exposure to CS induces angiogenesis by selectively degrading mature miR-1 via a vascular endothelial growth factor-driven pathway. Applying a multi-step molecular screen, we identified angiogenic genes regulated by miR-1 in the lungs of smokers. Knockdown of one of these genes, Notch homolog protein 3, simulates the anti-angiogenic effects of miR-1. These findings suggest that miR-1 can be used as an indicator of malignant transformation.

The occurrence of diabetic kidney disease (DKD), a critical microvascular issue in diabetes, is progressively on the rise. In recent years, long noncoding RNAs (lncRNAs) have garnered considerable attention as a novel and critical layer of biological regulation. Our knowledge regarding the roles and underlying mechanisms of lncRNAs in various diseases, including DKD, continues to evolve. Similarly, microRNAs (miRNAs), which are small noncoding RNAs, have been recognized as crucial contributors to cellular processes and disease pathogenesis. Emerging studies have highlighted the complex interactions between lncRNAs and miRNAs, particularly in the context of DKD, underscoring their importance in complex human diseases. Renal intrinsic cell damage is an important cause of inducing DKD. Persistent high glucose stimulation leads to remodeling of renal intrinsic cells and a cascade of pathological changes. This article aims to review recent literature on the lncRNAs-mediated regulation of miRNAs affecting renal intrinsic cells in DKD and to propose novel molecular-level therapeutic strategies for DKD. Through in-depth investigation of this dynamic molecular interaction, we can gain a profound understanding of the potential mechanisms underlying diabetic nephropathy, potentially identifying new targets for therapeutic intervention and paving the way for personalized and effective treatments.

In an obesogenic environment, such as the one we have been experiencing in recent decades, epigenetics provides answers to the relationship between hereditary and environmentally acquired patterns that have significantly contributed to the global rise in obesity prevalence. MicroRNA (miRNA) constitutes a diminutive non-coding small RNA molecule, 20 to 24 nucleotides in length, that functions as a regulator of gene regulation at the post-translational level. Circulating miRNAs (c-miRNAs) have been detected in multiple body fluids, including blood, plasma, serum, saliva, milk from breastfeeding mothers, and urine. These molecules hold significant therapeutic value and serve as extracellular biomarkers in metabolic diseases. They aid in the diagnosis and tracking of therapy responses, as well as dietary and physical habit modifications. Researchers have studied c-miRNAs as potential biomarkers for diagnosing and characterizing systemic diseases in people of all ages and backgrounds since then. These conditions encompass dyslipidemia, type 2 diabetes mellitus (T2DM), cardiovascular risk, metabolic syndrome, cardiovascular diseases, and obesity. This review therefore analyzes the usefulness of c-miRNAs as therapeutic markers over the past decades. It also provides an update on c-miRNAs associated with general obesity and overweight, as well as with the most prevalent pathologies in the adult population. It also examines the effect of different nutritional approaches and physical activity regarding the activity of miRNAs in circulation in adults with overweight or general obesity. All of this is done with the aim of evaluating their potential use as biomarkers in various research contexts related to overweight and obesity in adults.

Supplementing choline and DHA to pregnant gilts modified fetal pig hepatic global DNA methylation induced by gestational malnutrition, suggesting that gene expression and regulation and its associated metabolic pathways are affected in the liver of offspring during growth and development.

This study aimed to investigate the effect of maternal supplementation of choline, DHA, and their interaction on hepatic mRNA expression, miRNA regulation, and metabolic pathways in the fetal pigs born to malnourished mothers.

The abundance of mRNA and miRNA was profiled in fetal liver from sows with undernutrition supplemented with choline and DHA in a 2 × 2 factorial design. The effects of choline, DHA, and their interaction on mRNA and miRNA expression were evaluated. Identification of the Biological Processes from the Gene Ontology database and miRNA Target Prediction Analysis were performed using the DAVID Functional Annotation Tool and Ingenuity Pathway Analysis. The identified miRNA-mRNA pairings were validated using RT-qPCR.

In total, 144 mRNA and 1 miRNA were altered by supplementation of choline, and the alterations were associated with the inhibitions of cardiac hypertrophy signaling, IL-6 signaling, IL-3 signaling, the Th1 pathway, and the acute phase response signaling pathway. Further, 151 mRNAs and 6 miRNAs were altered by maternal supplementation DHA and were associated with inhibition of 5 inositol-related pathways, 5 immune-related pathways, and 7 other pathways and the stimulation of peroxisome proliferator-activated receptor signaling and RhoGDI signaling pathways. In addition, 383 mRNAs and 25 miRNAs displayed choline × DHA interactions including synergistic effects on acute phase response signaling, and antagonistic effects on tRNA splicing, peroxisome proliferator-activated receptor α/retinoid X receptor α activation, and sirtuin signaling, NAD signaling, and RNA polymerase I transcription pathways. Ten of the identified 20 miRNA-mRNA pairings were validated using RT-qPCR.

The supplementation of choline, DHA, or choline plus DHA to pregnant gilts modifies liver mRNA, miRNA, and pathways in fetal pigs during gestational undernutrition.

microRNAs (miRNAs) are important post-transcriptional regulators that activate silencing mechanisms by annealing to mRNA transcripts. While plant miRNAs match their targets with nearly-full complementarity leading to mRNA cleavage, miRNAs in most animals require only a short sequence called 'seed' to inhibit target translation. Recent findings showed that miRNAs in cnidarians, early-branching metazoans, act similarly to plant miRNAs, by exhibiting full complementarity and target cleavage; however, it remained unknown if seed-based regulation was possible in cnidarians. Here, we investigate the miRNA-target complementarity requirements for miRNA activity in the cnidarian Nematostella vectensis. We show that bilaterian-like complementarity of seed-only or seed and supplementary 3' matches are insufficient for miRNA-mediated knockdown. Furthermore, miRNA-target mismatches in the cleavage site decrease knockdown efficiency. Finally, miRNA silencing of a target with three seed binding sites in the 3' untranslated region that mimics typical miRNA targeting was repressed in zebrafish but not in Nematostella and another cnidarian, Hydractinia symbiolongicarpus. Altogether, these results unravel striking similarities between plant and cnidarian miRNAs supporting a possible common evolutionary origin of miRNAs in plants and animals.

Over the past decade, a scientific field has been developed demonstrating microRNAs (miRNAs) to be actively sorted into extracellular vesicles via specific nucleotide motifs that interact with discrete RNA-binding proteins. These miRNAs are proposed to be transported into recipient cells in which they can regulate specific cellular pathways. This mechanism could have enormous potential in explaining how cells signal and regulate other cells nearby or at a distance. Tens of studies have built this theme of a regulated transport of miRNAs. However, some concerns exist about this field. Taken together, there are concerns of a lack of a consistent motif, RNA-binding protein, or preferential miRNA involved in this process. In this study, we provide an expert and extensive analysis of the field that makes the cases for and against an active sorting mechanism. We provide potential explanations on why there is a lack of agreement. Most importantly, we provide ideas on how to move this field forward with more rigor and reproducibility. It is hoped that by engaging in a scientific debate of the pros and cons of this field, more rigorous experiments can be performed to conclusively demonstrate this biological activity.

Chronic heart failure (CHF) combined with hyperuricemia (HUA) is a comorbidity that is hard to diagnose by a single biomarker. Exosomal miRNAs are differentially expressed in cardiovascular diseases and are closely associated with regulating most biological functions. This study aimed to provide evidence for miRNA as a new molecular marker for precise diagnosis of the comorbidity of CHF with HUA and further analyze the potential targets of differentially expressed miRNA. This controlled study included 30 CHF patients combined with HUA (Group T) and 30 healthy volunteers (Group C). 6 peripheral blood samples from Group T and Group C were analyzed for exosomal miRNAs by high-throughput sequencing and then validated in the remaining 24 peripheral blood samples from Group T and Group C by applying real-time PCR (RT-PCR). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were performed using R software to predict the differential miRNAs' action targets. 42 differentially expressed miRNAs were detected (18 upregulated and 24 downregulated), in which miR-27a-5p was significantly upregulated (P<0.01), and miR-139-3p was significantly downregulated (P<0.01) in Group T. The combination of miR-27a-5p and miR-139-3p predicted the development of CHF combined with HUA with a maximum area under the curve (AUC) of 0.899 (95 % CI: 0.812-0.987, SEN=79.2 %, SPE=91.7 %, J value = 0.709). GO and KEGG enrichment analysis revealed that the differentially expressed miRNAs had a role in activating the AMPK-mTOR signaling pathway to activate the autophagic response. Collectively, our findings suggest that upregulated exosomal miR-27a-5p combined with downregulated exosomal miR-139-3p can be used as a novel molecular marker for precise diagnosis of CHF combined with HUA and enhanced autophagy by AMPK-mTOR signaling pathway may be one pathogenesis of the differentially expressed miRNAs.

Balanced mating type polymorphisms offer a distinct window into the forces shaping sexual reproduction strategies. Multiple hermaphroditic genera in Juglandaceae, including walnuts (Juglans) and hickories (Carya), show a 1:1 genetic dimorphism for male versus female flowering order (heterodichogamy). We map two distinct Mendelian inheritance mechanisms to ancient (>37 million years old) genus-wide structural DNA polymorphisms. The dominant haplotype for female-first flowering in Juglans contains tandem repeats of the 3' untranslated region of a gene putatively involved in trehalose-6-phosphate metabolism and is associated with increased cis gene expression in developing male flowers, possibly mediated by small RNAs. The Carya locus contains ~20 syntenic genes and shows molecular signatures of sex chromosome-like evolution. Inheritance mechanisms for heterodichogamy are deeply conserved, yet may occasionally turn over, as in sex determination.

Low back pain (LBP) is generally caused by lumbar degeneration without effective treatment. Lumbar degeneration is influenced by aberrant axial mechanical stress (MS), with facet joint osteoarthritis (FJOA) representing one of its primary pathological manifestations. MicroRNA (miRNA), functioning as an early intermediate in the transcription process, has frequently been demonstrated to serve as a critical mediator linking mechanical stress perception with cellular processes such as growth, development, aging, and apoptosis. We hypothesized that miR-3085-3p regulates chondrocyte apoptosis under mechanical stress, influencing FJOA and serving as a key regulator.

The severity of cartilage degeneration in bipedal standing models (BSM) was established and validated through micro-CT and histopathology. Cyclic tensile strain experiments (CTS) were conducted on the ATDC5 cell line to simulate MS. In situ hybridization was utilized to assess the expression levels of miR-3085-3p in degraded facet articular cartilage. The role of miR-3085-3p and its interaction with the downstream mRNA target Hspb6 were investigated through a combination of bioinformatic analysis, quantitative real-time polymerase chain reaction, western blotting, immunofluorescence, and luciferase assay. In vivo experiments on BSM, the functional impact of miR-3085-3p was further examined through transfection with adeno-associated virus (AAV).

It was observed that miR-3085-3p induced endoplasmic reticulum (ER) stress and apoptosis in chondrocytes and cartilage tissues under MS. The detrimental impact of miR-3085-3p was associated with the downregulation of Hspb6 expression, resulting in disruption of endoplasmic reticulum folding function. Additionally, intra-articular transfection of AAV miR-3085-3p mimics in mice facet joints led to spontaneous cartilage loss, while AAV miRNA-3085-3p sponge administration mitigated FJOA in the murine BSM model.

Mechanical stress-regulated miR-3085-3p up regulation induced the ER stress and aggravates FJOA development through targeting HSPB6, suggesting miR-3085-3p may be a novel therapeutic target for FJOA.Translational potential of this article: Our study confirmed the elevated expression of miR-3085-3p in lumbar facet joints following mechanical stress loading, suggesting that miR-3085-3p may serve as a biomarker for the clinical management of FJOA. Additionally, we demonstrated that the knockdown of miR-3085-3p in animal facet joints mitigated facet joint degeneration, thereby identifying a potential therapeutic target for FJOA.

Plant microRNAs (miRNAs) are small non-coding RNA molecules that usually negatively regulate gene expression at the post-transcriptional level. Recent data reveal that plant miRNAs are not limited to individual plants but can transfer across different species, allowing for communication with the plant, animal, and microbial worlds in a cross-kingdom approach. This review discusses the differences in miRNA biosynthesis between plants and animals and summarizes the current research on the cross-species regulatory effects of plant miRNAs on nearby plants, pathogenic fungi, and insects, which can be applied to crop disease and pest resistance. In particular, this review highlights the latest findings regarding the function of plant miRNAs in the transboundary regulation of human gene expression, which may greatly expand the clinical applicability of plant miRNAs as intriguing tools in natural plant-based medicinal products in the future.

Engineered extracellular vesicles (EVs) have been recognized as important therapeutics for gene and cell therapy. To achieve clinically desired therapy, technologies for EV engineering have high demands on the efficacy in producing EVs and their qualities, which, however, remain challenging to conventional routes due to their limited control on therapeutic payload delivery, EV secretion, and extracellular microenvironments. Here, we report a nanoplatform (denoted as PURE) that enables efficient electro-transfection while stimulating cells to produce high-quality EVs carrying functional RNAs. PURE further employs an ammonium removal zone to maintain the physiological conditions of the extracellular microenvironment and an EV uptake zone that efficiently (87.1%) captures EVs in situ with porous hydrogels. The platform achieved about a 12-fold higher yield of engineered EVs and a 146-fold abundance of desired therapeutics compared to those naturally secreted from cells. The PURE-engineered miR-130a-EVs were validated for effectively upregulating the mTOR signaling pathway in both in vitro and in vivo. Their therapeutic capability was then verified by enhancing the in vitro activation of primordial follicles. In vivo applications further highlighted the therapeutic effects of miR-130a-EVs in restoring ovary function in aged mice. The PURE platform represents a strategy for the clinical translation of EV-mediated therapy.

In mammalian cells, primary miRNAs are cleaved at their hairpin structures by the Microprocessor complex, whose core is composed of DROSHA and DGCR8. Here, we show that 5' flanking regions, resulting from Microprocessor cleavage, are targeted by the RNA exosome in mouse embryonic stem cells (mESCs). This is facilitated by a physical link between DGCR8 and the nuclear exosome targeting (NEXT) component ZCCHC8. Surprisingly, however, both biochemical and mutagenesis studies demonstrate that a variant NEXT complex, containing the RNA helicase MTR4 but devoid of the RNA-binding protein RBM7, is the active entity. This Microprocessor-NEXT variant also targets stem-loop-containing RNAs expressed from other genomic regions, such as enhancers. By contrast, Microprocessor does not contribute to the turnover of less structured NEXT substrates. Our results therefore demonstrate that MTR4-ZCCHC8 can link to either RBM7 or DGCR8/DROSHA to target different RNA substrates depending on their structural context.

The prevalence of type 2 diabetes is increasing worldwide. Various molecular mechanisms have been proposed to interfere with the insulin signaling pathway. Recent advances in proteomics and genomics indicate that one such mechanism involves the post-transcriptional regulation of insulin signaling by microRNA (miRNA). These noncoding RNAs typically induce messenger RNA (mRNA) degradation or translational repression by interacting with the 3' untranslated region (3'UTR) of target mRNA. Dietary components and patterns, which can either enhance or impair the insulin signaling pathway, have been found to regulate miRNA expression in both in vitro and in vivo studies. This review provides an overview of the current knowledge of how dietary components influence the expression of miRNAs related to the control of the insulin signaling pathway and discusses the potential application of these findings in precision nutrition.

DNA demethylases TET2 and TET3 play a fundamental role in thymic invariant natural killer T (iNKT) cell differentiation by mediating DNA demethylation of genes encoding for lineage specifying factors. Paradoxically, differential gene expression analysis revealed that significant number of genes were upregulated upon TET2 and TET3 loss in iNKT cells. This unexpected finding could be potentially explained if loss of TET proteins was reducing the expression of proteins that suppress gene expression. In this study, we discover that TET2 and TET3 synergistically regulate Drosha expression, by generating 5hmC across the gene body and by impacting chromatin accessibility. As DROSHA is involved in microRNA biogenesis, we proceed to investigate the impact of TET2/3 loss on microRNAs in iNKT cells. We report that among the downregulated microRNAs are members of the Let-7 family that downregulate in vivo the expression of the iNKT cell lineage specifying factor PLZF. Our data link TET proteins with microRNA expression and reveal an additional layer of TET mediated regulation of gene expression.

MicroRNAs (miRNAs) are small, non-coding RNAs that are expressed in a tissue- and temporal-specific manner during development. They have been found to be highly conserved during the evolution of different species. miRNAs regulate the expression of several genes in various organisms, with some regulating the expression of multiple genes with similar or completely unrelated functions. Frequent disease and insect pest infestations severely limit agricultural development. Thus, cultivating resistant crops via miRNA-directed gene regulation in plants, insects, and pathogens is an important aspect of modern breeding practices. To strengthen the application of miRNAs in sustainable agriculture, plant endogenous or exogenous miRNAs have been used for plant breeding. Consequently, the development of biological pesticides based on miRNAs has become an important avenue for future pest control methods. However, selecting the appropriate miRNA according to the desired target traits in the target organism is key to successfully using this technology for pest control. This review summarizes the progress in research on miRNAs in plants and other species involved in regulating plant disease and pest resistance pathways. We also discuss the molecular mechanisms of relevant target genes to provide new ideas for future research on pest and disease resistance and breeding in plants.

microRNAs (miRNAs) were recognized as a promising source of diagnostic biomarker. Herein, we aim to evaluate the performance of an ultrasensitive method for detecting serum miRNAs using single molecule arrays (Simoa).

In this study, candidate miRNAs were trained and tested by RT-qPCR in a cohort of PTB patients. Besides that, ultrasensitive serum miRNA detection were developed using the Single Molecule Array (Simoa) platform. In this ultra-sensitive sandwich assay, two target-specific LNA-modified oligonucleotide probes can be simply designed to be complementary to the half-sequence of the target miRNA respectively. We characterized its analytical performance and measured miRNAs in the serum of patients with pulmonary tuberculosis and healthy individuals.

We identified a five signature including three upregulated (miR-101, miR-196b, miR-29a) and two downregulated (miR-320b, miR-99b) miRNAs for distinguishing PTB patients from HCs, and validated in our 104 PTB patients. On the basis of Simoa technology, we developed a novel, fully automated digital analyser, which can be used to directly detect miRNAs in serum samples without pre-amplification. We successfully detected miRNAs at femtomolar concentrations (with limits of detection [LODs] ranging from 0.449 to 1.889 fM). Simoa-determined serum miR-29a and miR-99b concentrations in patients with PTB ((median 6.06 fM [range 0.00-75.22]), (median 2.53 fM [range 0.00-24.95]), respectively) were significantly higher than those in HCs ((median 2.42 fM [range 0.00-28.64]) (P < 0.05), (median 0.54 fM [range 0.00-9.12] (P < 0.0001), respectively). Serum levels of miR-320b were significantly reduced in patients with PTB (median 2.11 fM [range 0.00-39.30]) compared with those in the HCs (median 4.76 fM [range 0.00-25.10]) (P < 0.001). A combination of three miRNAs (miR-29a, miR-99b, and miR-320b) exhibited a good capacity to distinguish PTB from HCs, with an area under the curve (AUC) of 0.818 (sensitivity: 83.9%; specificity: 79.7%).

This study benchmarks the role of Simoa as a promising tool for monitoring miRNAs in serum and offers considerable potential as a non-invasive platform for the early diagnosis of PTB.

The advent of high-throughput sequencing uncovered that our genome is pervasively transcribed into RNAs that are seemingly not translated into proteins. It was also found that non-coding RNA transcripts outnumber canonical protein-coding genes. This mindboggling discovery prompted a surge in non-coding RNA research that started unraveling the functional relevance of these new genetic units, shaking the classic definition of "gene". While the non-coding RNA revolution was still taking place, polysome/ribosome profiling and mass spectrometry analyses revealed that peptides can be translated from non-canonical open reading frames. Therefore, it is becoming evident that the coding vs non-coding dichotomy is way blurrier than anticipated. In this review, we focus on several examples in which the binary classification of coding vs non-coding genes is outdated, since the same bifunctional gene expresses both coding and non-coding products. We discuss the implications of this intricate usage of transcripts in terms of molecular mechanisms of gene expression and biological outputs, which are often concordant, but can also surprisingly be discordant. Finally, we discuss the methodological caveats that are associated with the study of bifunctional genes, and we highlight the opportunities and challenges of therapeutic exploitation of this intricacy towards the development of anticancer therapies.

Periodontitis, with its persistent nature, causes significant distress for most sufferers. Current treatments, such as mechanical cleaning and surgery, often fail to fully address the underlying overactivation of fibroblasts that drives this degradation. Targeting the post-transcriptional regulation of fibroblasts, particularly at the 3'-untranslated regions (3'UTR) of pathogenic genes, offers a therapeutic strategy for periodontitis. Herein, we developed a DNA nanorobot for this purpose. This system uses a dynamic DNA nanoframework to incorporate therapeutic microRNAs through molecular recognition and covalent bonds, facilitated by DNA monomers modified with disulfide bonds. The assembled-DNA nanoframework is encapsulated in a cell membrane embedded with a fibroblast-targeting peptide. By analyzing the 3'UTR regions of pathogenic fibroblast genes FOSB and JUND, we identified the therapeutic microRNA as miR-1-3p and integrated it into this system. As expected, the DNA nanorobot delivered the internal components to fibroblasts by the targeting peptide and outer membrane that responsively releases miR-1-3p under intracellular glutathione. It resulted in a precise reduction of mRNA and suppression of protein function in pathogenic genes, effectively reprogramming fibroblast behavior. Our results confirm that this approach not only mitigates the inflammation but also promotes tissue regeneration in periodontal models, offering a promising therapeutic avenue for periodontitis.

Genitourinary cancer (GUC) represents more than one fifth of all human cancers. This makes the development of approaches to its early diagnosis an important task of modern biomedicine. Circulating microRNAs, short (17-25 nucleotides) non-coding RNA molecules found in human biological fluids and performing a regulatory role in the cell, are considered as promising diagnostic and prognostic biomarkers of cancers, including GUC. In this review we have considered the current state of research aimed at assessing microRNAs as biomarkers of such human GUC types as malignant tumors of the bladder, kidney, prostate, testicles, ovaries, and cervix. A special attention has been paid to studies devoted to the identification of microRNAs in urine as a surrogate "liquid biopsy" that may provide the simplest and cheapest approach to mass non-invasive screening of human GUC. The use of microRNA panels instead of single types of microRNA generally leads to higher sensitivity and specificity of the developed diagnostic tests. However, to date, work on the microRNAs assessment as biomarkers of human GUC is still of a research nature, and the further introduction of diagnostic tests based on microRNAs into practice requires successful clinical trials.

Extracellular vesicles (EVs) are abundant in body fluids, contributing to intercellular signalling by transferring cargo that includes microRNAs (miRs)-themselves implicated in pathobiology. For the first time we evaluated the potential of EV miRs to contribute diagnostic information in early RA, predict methotrexate (MTX) efficacy or shed light on the drug's mechanism of action.

Seven hundred and ninety-eight miRs isolated from serum-derived EVs of 46 patients with untreated RA, 23 with untreated polymyalgia rheumatica (PMR; inflammatory disease control group) and 12 in whom significant inflammatory disease had been excluded (non-inflammatory controls; NICs) were profiled (NanoString); the same measurements were made for RA patients after 6 months' MTX treatment. Analyses took multiple testing into account.

Twenty-eight EV miRs were robustly differentially expressed between early RA (but not PMR) patients and NICs after correction for age and sex, suggesting discriminatory value. Cross-validated partial least squares-discriminant analysis also indicated the predictive potential of a distinct baseline EV miR signature with respect to MTX-induced remission at 6 months. The change in expression of 13 miRs over the course of MTX treatment differed significantly between responders and non-responders, and four of those exhibiting increased relative abundance amongst responders have known roles in regulating the pathogenic potential of synovial fibroblasts, namely miR-212-3p, miR-338-5p, miR-410-3p and miR-537.

Our data highlight the potential of serum EV miRs as diagnostic and therapeutic biomarkers, highlighting a novel potential mechanism by which MTX may exert its therapeutic effect in early RA that warrants further investigation.

MicroRNAs (miRNAs/ miRs) are short, noncoding RNAs, usually consisting of 18 to 24 nucleotides, that control gene expression after the process of transcription and have crucial roles in several clinical processes. This article seeks to provide an in-depth review and evaluation of the many activities of miR-128, accentuating its potential as a versatile biomarker and target for therapy; The circulating miR-128 has garnered interest because of its substantial influence on gene regulation and its simplicity in extraction. Several miRNAs, such as miR-128, have been extracted from circulating blood cells, cerebrospinal fluid, and plasma/serum. The miR-128 molecule can specifically target a diverse range of genes, enabling it to have intricate physiological impacts by concurrently regulating many interrelated pathways. It has a vital function in several biological processes, such as modulating the immune system, regulating brain plasticity, organizing the cytoskeleton, and inducing neuronal death. In addition, miR-128 modulates genes associated with cell proliferation, the cell cycle, apoptosis, plasma LDL levels, and gene expression regulation in cardiac development. The dysregulation of miR-128 expression and activity is associated with the development of immunological responses, changes in neural plasticity, programmed cell death, cholesterol metabolism, and heightened vulnerability to autoimmune illnesses, neuroimmune disorders, cancer, and cardiac problems; The paper highlights the importance of studying the consequences of miR-128 dysregulation in these specific locations. By examining the implications of miRNA-128 dysregulation in these areas, the article underscores its significance in diagnosis and treatment, providing a foundation for research and clinical applications.

MicroRNA-seq data is produced by aligning small RNA sequencing reads of different miRNA transcript isoforms, called isomiRs, to known microRNAs. Aggregation to microRNA-level counts discards information and violates core assumptions of differential expression (DE) methods developed for mRNA-seq data. We establish miRglmm, a DE method for microRNA-seq data, that uses a generalized linear mixed model of isomiR-level counts, facilitating detection of miRNA with differential expression or differential isomiR usage. We demonstrate that miRglmm outperforms current DE methods in estimating DE for miRNA, whether or not there is significant isomiR variability, and simultaneously provides estimates of isomiR-level DE.

The function of exosomal miRNAs (miRs) in retinal degeneration is largely unclear. We were aimed to investigate the functions of exosomes as well as their miRs derived from retinal pigment epithelial (RPE) cells following exposure to oxidative stress (OS). After the OS by lipopolysaccharide and rotenone on RPE cells, interleukin-1 beta (IL-1β), Interleukin-6 (IL-6), Tumor Necrosis Factor-alpha (TNF-α) were upregulated, along with the decreased mitochondrial membrane potential and upregulated oxidative damage marker 8-OH-dG in RPE cells. RPE-derived exosomes were then isolated, identified, injected into the subretinal space in mice. After subretinal injection, RPE-exosomes after OS not only induced higher ROS level and apoptotic retinal cells, but also elevated IL-1β, IL-6 alongside TNF-α expressions among retina/RPE/choroidal complex. Next, miRs inside the exosomes were sequenced by the next generation sequencing (NGS) technology. NGS revealed that certain miRs were abundant in exosomes, while others were selectively kept by RPE cells. Further, downregulated miRs, like miR-125b-5p, miR-125a-5p, alongside miR-128-3p, and upregulated miR, such as miR-7-5p were validated byRT-qPCR. Finally, Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were used to find the possible target genes of those selective exosomal miRs. Our results proved that the RPE-derived exosomes after OS selectively express certain miRs, providing novel insights into the pathogenesis of age-related macular degeneration (AMD) in future.

The plant long noncoding (lnc)RNA field is on the brink of transitioning from large-scale identification of lncRNAs to their functional characterization. Due to the cross-kingdom conservation of interaction types and molecular functions, there is much to be learned from mammalian lncRNA research. Here, we discuss the different molecular processes involving lncRNAs from the regulation of chromatin to splicing. Furthermore, we discuss the lncRNA interactome, which includes proteins, other RNAs, and DNA. We explore and discuss how mammalian lncRNA functionalities could be reflected in similar pathways in plants and hypothesize that several breakthroughs in mammalian research could lead to the discovery of novel plant lncRNA molecular functions. Expanding our knowledge of the biological role of lncRNAs and their multiple applications paves the way for future agricultural applications.

Globally, osteoporosis is the most common systemic skeletal disease. There are many factors that influence osteoporosis' development and progression. During the pathogenesis of this disease, bone turnover is imbalanced between resorption and the formation of bone tissue. A growing interest has been devoted to the role that microRNA (miRNA) plays in osteoporosis regulation. A microRNA (miRNA) is a group of small single-stranded RNA molecules involved in regulating gene expression in eukaryotic organisms. As microRNAs (miRNAs) are key regulators of gene expression and can modulate processes related to bone metabolism, they have become increasingly important for studying osteoporosis pathogenesis. The available research suggests that miRNAs play an important role in regulating processes associated with bone metabolism, especially by influencing bone resorption and synthesis. Furthermore, microRNAs can also serve as potential therapeutic targets for osteoporosis, besides being a rapid and specific biomarker.

Nonsense-mediated mRNA decay (NMD) stands out as a prominent RNA surveillance mechanism within eukaryotes, meticulously overseeing both RNA abundance and integrity by eliminating aberrant transcripts. These defective transcripts are discerned through the concerted efforts of translating ribosomes, eukaryotic release factors (eRFs), and trans-acting NMD factors, with Up-Frameshift 3 (UPF3) serving as a noteworthy component. Remarkably, in humans, UPF3 exists in two paralogous forms, UPF3A (UPF3) and UPF3B (UPF3X). Beyond its role in quality control, UPF3 wields significant influence over critical cellular processes, including neural development, synaptic plasticity, and axon guidance. However, the precise regulatory mechanisms governing UPF3 remain elusive. MicroRNAs (miRNAs) emerge as pivotal post-transcriptional gene regulators, exerting substantial impact on diverse pathological and physiological pathways. This comprehensive review encapsulates our current understanding of the intricate regulatory nexus between NMD and miRNAs, with particular emphasis on the essential role played by UPF3B in neurodevelopment. Additionally, we bring out the significance of the 3'-untranslated region (3'-UTR) as the molecular bridge connecting NMD and miRNA-mediated gene regulation. Furthermore, we provide an in-depth exploration of diverse computational tools tailored for the prediction of potential miRNA targets. To complement these computational approaches, we delineate experimental techniques designed to validate predicted miRNA-mRNA interactions, empowering readers with the knowledge necessary to select the most appropriate methodology for their specific research objectives.

Epigenetic silencing through methylation is one of the major mechanisms for downregulation of tumor suppressor miRNAs in various malignancies. The aim of this study was to identify novel tumor suppressor miRNAs which are silenced by DNA hypermethylation and investigate the role of at least one of these in oral squamous cell carcinoma (OSCC) pathogenesis. We treated cells from an OSCC cell line SCC131 with 5-Azacytidine, a DNA methyltransferase inhibitor, to reactivate tumor suppressor miRNA genes silenced/downregulated due to DNA methylation. At 5-day post-treatment, total RNA was isolated from the 5-Azacytidine and vehicle control-treated cells. The expression of 2,459 mature miRNAs was analysed between 5-Azacytidine and control-treated OSCC cells by the microRNA microarray analysis. Of the 50 miRNAs which were found to be upregulated following 5-Azacytidine treatment, we decided to work with miR-6741-3p in details for further analysis, as it showed a mean fold expression of >4.0. The results of qRT-PCR, Western blotting, and dual-luciferase reporter assay indicated that miR-6741-3p directly targets the oncogene SRSF3 at the translational level only. The tumor-suppressive role of miR-6741-3p was established by various in vitro assays and in vivo study in NU/J athymic nude mice. Our results revealed that miR-6741-3p plays a tumor-suppressive role in OSCC pathogenesis, in part, by directly regulating SRSF3. Based on our observations, we propose that miR-6741-3p may serve as a potential biological target in tumor diagnostics, prognostic evaluation, and treatment of OSCC and perhaps other malignancies.

Circadian rhythms integrate a finely tuned network of biological processes recurring every 24 h, intricately coordinating the machinery of all cells. This self-regulating system plays a pivotal role in synchronizing physiological and behavioral responses, ensuring an adaptive metabolism within the environmental milieu, including dietary and physical activity habits. The systemic integration of circadian homeostasis involves a balance of biological rhythms, each synchronically linked to the central circadian clock. Central to this orchestration is the temporal dimension of nutrient and food intake, an aspect closely interwoven with the neuroendocrine circuit, gut physiology, and resident microbiota. Indeed, the timing of meals exerts a profound influence on cell cycle regulation through genomic and epigenetic processes, particularly those involving gene expression, DNA methylation and repair, and non-coding RNA activity. These (epi)genomic interactions involve a dynamic interface between circadian rhythms, nutrition, and the gut microbiota, shaping the metabolic and immune landscape of the host. This research endeavors to illustrate the intricate (epi)genetic interplay that modulates the synchronization of circadian rhythms, nutritional signaling, and the gut microbiota, unravelling the repercussions on metabolic health while suggesting the potential benefits of feed circadian realignment as a non-invasive therapeutic strategy for systemic metabolic modulation via gut microbiota. This exploration delves into the interconnections that underscore the significance of temporal eating patterns, offering insights regarding circadian rhythms, gut microbiota, and chrono-nutrition interactions with (epi)genomic phenomena, thereby influencing diverse aspects of metabolic, well-being, and quality of life outcomes.

MicroRNAs (miRNAs) are important and ubiquitous regulators of gene expression in both plants and animals. They are thought to have evolved convergently in these lineages and hypothesized to have played a role in the evolution of multicellularity. In line with this hypothesis, miRNAs have so far only been described in few unicellular eukaryotes. Here, we investigate the presence and evolution of miRNAs in Amoebozoa, focusing on species belonging to Acanthamoeba, Physarum and dictyostelid taxonomic groups, representing a range of unicellular and multicellular lifestyles. miRNAs that adhere to both the stringent plant and animal miRNA criteria were identified in all examined amoebae, expanding the total number of protists harbouring miRNAs from 7 to 15. We found conserved miRNAs between closely related species, but the majority of species feature only unique miRNAs. This shows rapid gain and/or loss of miRNAs in Amoebozoa, further illustrated by a detailed comparison between two evolutionary closely related dictyostelids. Additionally, loss of miRNAs in the Dictyostelium discoideum drnB mutant did not seem to affect multicellular development and, hence, demonstrates that the presence of miRNAs does not appear to be a strict requirement for the transition from uni- to multicellular life.

A subset of circular RNAs (circRNAs) and linear RNAs have been proposed to 'sponge' or block microRNA activity. Additionally, certain RNAs induce microRNA destruction through the process of Target RNA-Directed MicroRNA Degradation (TDMD), but whether both linear and circular transcripts are equivalent in driving TDMD is unknown. Here, we studied whether circular/linear topology of endogenous and artificial RNA targets affects TDMD. Consistent with previous knowledge that Cdr1as (ciRS-7) circular RNA protects miR-7 from Cyrano-mediated TDMD, we demonstrate that depletion of Cdr1as reduces miR-7 abundance. In contrast, overexpression of an artificial linear version of Cdr1as drives miR-7 degradation. Using plasmids that express a circRNA with minimal co-expressed cognate linear RNA, we show differential effects on TDMD that cannot be attributed to the nucleotide sequence, as the TDMD properties of a sequence often differ when in a circular versus linear form. By analysing RNA sequencing data of a neuron differentiation system, we further detect potential effects of circRNAs on microRNA stability. Our results support the view that RNA circularity influences TDMD, either enhancing or inhibiting it on specific microRNAs.

Human and viral microRNAs (miRNAs) are involved in the regulation of gene transcription, and the establishment of their profiles in acute (AHI) and chronic (CHI) HIV infections may shed light on the pathogenetic events related to different phases of HIV disease. Next-generation sequencing (NGS) of miRNA libraries was performed, and the reads were used to analyze miRNA differential expression in the plasma with AHI and CHI. Functional analysis was then undertaken to investigate the biological processes characterizing the two phases of HIV infection. Except for hsa-miR-122-5p, which was found in 3.39% AHI vs. 0.18% CHI, the most represented human miRNAs were similarly represented in AHI and CHI. However, when considering the overall detected miRNAs in AHI and CHI, 15 displayed differential expression (FDR p < 0.05). Functional analysis identified 163 target mRNAs involved in promoting angiogenesis activation in AHI versus CHI through the action of hsa-miR10b-5p, hsa-miR1290, hsa-miR1-3p, and hsa-miR296-5p. The viral miRNAs detected, all belonging to herpesviruses, accounted for only 0.014% of total reads. The present data suggest that AHI patients exhibit strong innate immune activation through the upregulation of hsa-miR-122-5p and early activation of angiogenesis. More specific investigations are needed to study the role of viral miRNAs in HIV pathogenesis.