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Nature Mar 2015The first step in the biogenesis of microRNAs is the processing of primary microRNAs (pri-miRNAs) by the microprocessor complex, composed of the RNA-binding protein...
The first step in the biogenesis of microRNAs is the processing of primary microRNAs (pri-miRNAs) by the microprocessor complex, composed of the RNA-binding protein DGCR8 and the type III RNase DROSHA. This initial event requires recognition of the junction between the stem and the flanking single-stranded RNA of the pri-miRNA hairpin by DGCR8 followed by recruitment of DROSHA, which cleaves the RNA duplex to yield the pre-miRNA product. While the mechanisms underlying pri-miRNA processing have been determined, the mechanism by which DGCR8 recognizes and binds pri-miRNAs, as opposed to other secondary structures present in transcripts, is not understood. Here we find in mammalian cells that methyltransferase-like 3 (METTL3) methylates pri-miRNAs, marking them for recognition and processing by DGCR8. Consistent with this, METTL3 depletion reduced the binding of DGCR8 to pri-miRNAs and resulted in the global reduction of mature miRNAs and concomitant accumulation of unprocessed pri-miRNAs. In vitro processing reactions confirmed the sufficiency of the N(6)-methyladenosine (m(6)A) mark in promoting pri-miRNA processing. Finally, gain-of-function experiments revealed that METTL3 is sufficient to enhance miRNA maturation in a global and non-cell-type-specific manner. Our findings reveal that the m(6)A mark acts as a key post-transcriptional modification that promotes the initiation of miRNA biogenesis.
Topics: Adenosine; Base Sequence; Cell Line; Gene Expression Regulation; Humans; Methylation; Methyltransferases; MicroRNAs; Molecular Sequence Data; Nucleic Acid Conformation; RNA Processing, Post-Transcriptional; RNA-Binding Proteins; Substrate Specificity
PubMed: 25799998
DOI: 10.1038/nature14281 -
ELife Aug 2015MicroRNA targets are often recognized through pairing between the miRNA seed region and complementary sites within target mRNAs, but not all of these canonical sites are...
MicroRNA targets are often recognized through pairing between the miRNA seed region and complementary sites within target mRNAs, but not all of these canonical sites are equally effective, and both computational and in vivo UV-crosslinking approaches suggest that many mRNAs are targeted through non-canonical interactions. Here, we show that recently reported non-canonical sites do not mediate repression despite binding the miRNA, which indicates that the vast majority of functional sites are canonical. Accordingly, we developed an improved quantitative model of canonical targeting, using a compendium of experimental datasets that we pre-processed to minimize confounding biases. This model, which considers site type and another 14 features to predict the most effectively targeted mRNAs, performed significantly better than existing models and was as informative as the best high-throughput in vivo crosslinking approaches. It drives the latest version of TargetScan (v7.0; targetscan.org), thereby providing a valuable resource for placing miRNAs into gene-regulatory networks.
Topics: Animals; Binding Sites; Computational Biology; Gene Expression Regulation; Mammals; MicroRNAs; Molecular Biology; RNA, Messenger
PubMed: 26267216
DOI: 10.7554/eLife.05005 -
International Journal of Molecular... Dec 2022MicroRNAs (miRNAs) act as master regulators of gene expression in homeostasis and disease. Despite the rapidly growing body of evidence on the theranostic potential of... (Review)
Review
MicroRNAs (miRNAs) act as master regulators of gene expression in homeostasis and disease. Despite the rapidly growing body of evidence on the theranostic potential of restoring miRNA levels in pre-clinical models, the translation into clinics remains limited. Here, we review the current knowledge of miRNAs as T-cell targeting immunotherapeutic tools, and we offer an overview of the recent advances in miRNA delivery strategies, clinical trials and future perspectives in RNA interference technologies.
Topics: MicroRNAs; T-Lymphocytes; RNA Interference; Precision Medicine; Immunotherapy
PubMed: 36613706
DOI: 10.3390/ijms24010250 -
RNA (New York, N.Y.) Jul 2022Hairpin-containing pre-miRNAs, produced from pri-miRNAs, are precursors of miRNAs (microRNAs) that play essential roles in gene expression and various human diseases....
Hairpin-containing pre-miRNAs, produced from pri-miRNAs, are precursors of miRNAs (microRNAs) that play essential roles in gene expression and various human diseases. Current qPCR-based methods used to quantify pre-miRNAs are not effective to discriminate between pre-miRNAs and their parental pri-miRNAs. Here, we developed the intramolecular ligation method (iLIME) to quantify and sequence pre-miRNAs specifically. This method utilizes T4 RNA ligase 1 to convert pre-miRNAs into circularized RNAs, allowing us to design PCR primers to quantify pre-miRNAs, but not their parental pri-miRNAs. In addition, the iLIME also enables us to sequence the ends of pre-miRNAs using next-generation sequencing. Therefore, this method offers a simple and effective way to quantify and sequence pre-miRNAs, so it will be highly beneficial for investigating pre-miRNAs when addressing research questions and medical applications.
Topics: High-Throughput Nucleotide Sequencing; Humans; MicroRNAs; Real-Time Polymerase Chain Reaction
PubMed: 35487691
DOI: 10.1261/rna.079101.122 -
Journal of Integrative Bioinformatics Dec 2016Editorial The term MicroRNA or its contraction miRNA currently appears in 21,215 titles of abstracts, published between 1997 and now, available on Pubmed...
Editorial The term MicroRNA or its contraction miRNA currently appears in 21,215 titles of abstracts, published between 1997 and now, available on Pubmed (2016-21-22:12:59 EET). 4,108 of these were published in 2016 alone which signifies the importance of miRNA-related research. MicroRNAs can be detected experimentally using various techniques like directional cloning of endogenous small RNAs but they are time consuming [1]. Additionally, it is necessary for the miRNA and its mRNA target(s) to be co-expressed to infer a functional relationship which is difficult, if not impossible, to achieve [2]. Since experimental approaches are facing such difficulties, they have been complemented by computational approaches [3] thereby defining the field of computational miRNomics. Due to the rapid development in the discipline, it is important to assess the state-of-the-art. In this special issue, several areas of the field are investigated ranging from pre-miRNA detection via machine learning to application of differential expression analysis in plants. First, Saçar Demirci et al. discuss an approach to virus pre-miRNA detection using machine learning [4]. Such approaches are based on parameterization of miRNAs and Yousef et al. discuss how to select among such features [5]. A different computational perspective is provided by Kotipalli et al. who model the kinetics of miRNA genesis and targeting [6]. To fuel more refined future models for genesis and targeting, it is important to establish miRNA and target expression under varying conditions. Zhang et al. [7] and Kanke et al. [8] discuss two approaches to quantify miRNAs and other non-coding short RNAs. Diler et al., finally, discuss actual biological implications of differentially expressed miRNAs [9]. This special issue on computational miRNomics, thus, provides a trajectory from detection of pre-miRNAs to biological implications of differentially expressed miRNAs. Additional topics will be covered in the upcoming second volume of the special issue on computational miRNomics.
Topics: Computational Biology; Machine Learning; MicroRNAs
PubMed: 29216003
DOI: 10.1515/jib-2016-302 -
Annals of Medicine Dec 2022MicroRNAs (miRNAs) are a class of small non-coding, single-stranded RNAs (ribonucleic acids) that play important roles in many vital processes through their impact on... (Review)
Review
MicroRNAs (miRNAs) are a class of small non-coding, single-stranded RNAs (ribonucleic acids) that play important roles in many vital processes through their impact on gene expression. One such miRNA, miR210, represents a hypoxia-induced cellular miRNA group that hold a variety of functions. This review article highlights the importance of miR-210 in the development of pre-eclampsia.KEY MESSAGEmiR-210 is a promising biomarker for monitoring pregnancy with pre-eclampsia. Overexpression of miR-210 had a negative impact on the process of cell migration and trophoblast invasion.
Topics: Cell Movement; Female; Humans; MicroRNAs; Placenta; Pre-Eclampsia; Pregnancy; Trophoblasts
PubMed: 35543206
DOI: 10.1080/07853890.2022.2071459 -
Methods in Molecular Biology (Clifton,... 2021Research on RNA function and therapeutic potential is dominated by the use of chemoengineered RNA mimics. Recent efforts have led to the establishment of novel...
Research on RNA function and therapeutic potential is dominated by the use of chemoengineered RNA mimics. Recent efforts have led to the establishment of novel technologies for the production of recombinant or bioengineered RNA molecules, which should better recapitulate the structures, functions and safety profiles of natural RNAs because both are produced and folded in living cells. Herein, we describe a robust approach for reproducible fermentation production of bioengineered RNA agents (BERAs) carrying warhead miRNAs, siRNAs, aptamers, or other forms of small RNAs, based upon an optimal hybrid tRNA/pre-miRNA carrier. Target BERA/sRNAs are readily purified by fast protein liquid chromatography (FPLC) to a high degree of homogeneity (>97%). This approach offers a consistent high-level expression (>30% of total bacterial RNAs) and large-scale production of ready-to-use BERAs (multiple to tens milligrams from 1 L bacterial culture).
Topics: Base Sequence; Bioengineering; Chromatography, Ion Exchange; Cloning, Molecular; Drug Contamination; Electrophoresis, Polyacrylamide Gel; Endotoxins; Escherichia coli; Fermentation; MicroRNAs; Nucleic Acid Denaturation; Plasmids; Polymerase Chain Reaction; RNA; RNA, Bacterial; RNA, Transfer; RNA, Untranslated
PubMed: 34086286
DOI: 10.1007/978-1-0716-1499-0_18 -
International Journal of Molecular... Sep 2022Medicinal plant microRNAs (miRNAs) are an endogenous class of small RNA central to the posttranscriptional regulation of gene expression. Biosynthetic research has shown... (Review)
Review
Medicinal plant microRNAs (miRNAs) are an endogenous class of small RNA central to the posttranscriptional regulation of gene expression. Biosynthetic research has shown that the mature miRNAs in medicinal plants can be produced from either the standard messenger RNA splicing mechanism or the pre-ribosomal RNA splicing process. The medicinal plant miRNA function is separated into two levels: (1) the cross-kingdom level, which is the regulation of disease-related genes in animal cells by oral intake, and (2) the intra-kingdom level, which is the participation of metabolism, development, and stress adaptation in homologous or heterologous plants. Increasing research continues to enrich the biosynthesis and function of medicinal plant miRNAs. In this review, peer-reviewed papers on medicinal plant miRNAs published on the Web of Science were discussed, covering a total of 78 species. The feasibility of the emerging role of medicinal plant miRNAs in regulating animal gene function was critically evaluated. Staged progress in intra-kingdom miRNA research has only been found in a few medicinal plants, which may be mainly inhibited by their long growth cycle, high demand for growth environment, immature genetic transformation, and difficult RNA extraction. The present review clarifies the research significance, opportunities, and challenges of medicinal plant miRNAs in drug development and agricultural production. The discussion of the latest results furthers the understanding of medicinal plant miRNAs and helps the rational design of the corresponding miRNA/target genes functional modules.
Topics: Animals; Gene Expression Regulation, Plant; MicroRNAs; Plants, Medicinal; RNA, Messenger; RNA, Plant; RNA, Ribosomal
PubMed: 36142389
DOI: 10.3390/ijms231810477 -
Journal of Pharmaceutical and... Jun 2022Let-7e precursor microRNA has the potential to adopt a G-quadruplex (rG4) structure and recently, its roles in oncology have been the focus of much attention, as it is...
Let-7e precursor microRNA has the potential to adopt a G-quadruplex (rG4) structure and recently, its roles in oncology have been the focus of much attention, as it is now known that let-7e pre-miRNA is frequently dysregulated in cancers. Therefore, it is crucial to unveil and fully characterize its ability to adopt a rG4 structure, which could be stabilized or destabilized by small molecules and proteins such as nucleolin, a protein that is deeply associated with miRNA biogenesis. Herein, by combining a set of different methods such as circular dichroism (CD), nuclear magnetic resonance (NMR), UV spectroscopy (thermal difference spectra (TDS) and isothermal difference spectra (IDS)) and polyacrylamide gel electrophoresis (PAGE), we demonstrate the formation of the rG4 structure found in let-7e pre-miRNA sequence in the presence of K (5'-GGGCUGAGGUAGGAGG-3'). The ability of eight small molecules (or ligands) to bind to and stabilize this rG4 structure was also fully assessed. The dissociation constants for each RNA G-quadruplex/ligand complex, determined by surface plasmon resonance (SPR), ranged in the 10 to 10 M range. Lastly, the binding of the rG4 structure to nucleolin in the presence and absence of ligands was evaluated via CD, SPR, PAGE and confocal microscopy. The small molecules 360 A and PDS demonstrated attractive properties to targetthe rG4 structure of let-7e pre-miRNA and control its biology. Our findings also highlighted that the interaction of TMPyP4 with the G-quadruplex of let-7e precursor miRNA could block the formation of the complex between the rG4 and nucleolin. Overall, this study introduces an approach to target the rG4 found in let-7e pre-miRNA which opens up a new opportunity to control the microRNA biogenesis.
Topics: G-Quadruplexes; Ligands; MicroRNAs; Phosphoproteins; RNA-Binding Proteins; Nucleolin
PubMed: 35462282
DOI: 10.1016/j.jpba.2022.114757 -
Cancer Research Mar 2015miRNAs are important regulatory elements for gene expression that are involved in diverse physiologic and pathologic processes. Canonical miRNA biogenesis consists of a... (Review)
Review
miRNAs are important regulatory elements for gene expression that are involved in diverse physiologic and pathologic processes. Canonical miRNA biogenesis consists of a two-step processing, from primary transcripts (pri-miRNA) to precursor miRNAs (pre-miRNA) mediated by Drosha in the nucleus and from pre-miRNAs to mature miRNAs mediated by Dicer in the cytoplasm. Various routes of miRNA maturation that are tightly regulated by signaling cascades and specific to an individual or a subclass of miRNAs have been recently identified. Here, we review the current findings in signaling-mediated miRNA processing as well as their potential clinical relevance in cancer.
Topics: Animals; Gene Expression; Humans; MicroRNAs; Neoplasms; Signal Transduction
PubMed: 25660948
DOI: 10.1158/0008-5472.CAN-14-2568