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At least 55 records · Page 3Linked to original sources
PubMed2004

miRNP:mRNA association in polyribosomes in a human neuronal cell line.

MicroRNAs (miRNAs) are small regulatory RNAs that control gene expression by base-pairing with their mRNA targets. miRNAs assemble into ribonucleoprotein complexes termed miRNPs. Animal miRNAs recognize their mRNA targets via partial antisense complementarity and repress mRNA translation at a step after translation initiation. How animal miRNAs recognize their mRNA targets and how they control their translation is unknown. Here we describe that in a human neuronal cell line, the miRNP proteins eIF2C2 (a member of the Argonaute family of proteins), Gemin3, and Gemin4 along with miRNAs cosediment with polyribosomes. Furthermore, we describe a physical association between a let-7b (miRNA)-containing miRNP and its putative human mRNA target in polyribosome-containing fractions. These findings suggest that miRNP proteins may play important roles in target mRNA recognition and translational repression.

Binding Sites↗
PubMed2006

An LNA-based loss-of-function assay for micro-RNAs.

MicroRNAs (miRNAs) have recently emerged as being essential for development and for the control of cell proliferation/differentiation in various organisms. However, little is known about miRNA function and mode of action at the cellular level. We have designed a miRNA loss-of-function assay, based on chemically modified locked nucleic acids (LNA) antisense oligonucleotides and usable in tissue culture cells. We show that LNA/DNA mixed oligonucleotides form highly stable duplexes with miRNAs in vitro. Ex vivo, the target miRNA becomes undetectable in cells transfected with the antisense oligonucleotide. The effect is dose-dependent, long-lasting, and specific. Moreover, using a reporter assay, we show that antisense LNA/DNA oligonucleotides inhibit short non-coding RNAs at the functional level. Thus LNA/DNA mixmers represent powerful tools for functional analysis of miRNAs.

Cells, Cultured↗
PubMed2006

Computational identification of microRNAs and their targets.

MicroRNAs (miRNAs) are one class of newly identified riboregulators of gene expression in many eukaryotic organisms. They play important roles in multiple biological and metabolic processes, including developmental timing, signal transduction, cell maintenance and differentiation, diseases and cancers. miRNAs regulate gene expression at the posttranscriptional level by directly cleaving targeted mRNAs or repressing translation. Although the founding members of miRNAs were discovered by genetic screening approaches, experimental approaches were limited by their low efficiency, time consuming, and high cost. As an alternative, computational approaches were developed. Computational approaches for identifying miRNAs are based on the following major characteristics of miRNAs: hairpin-shaped secondary structures, high conservation for some miRNAs, and high minimal folding free energy index (MFEI). Computational approaches also play an important role in identifying miRNA targets. A majority of known miRNAs and their targets were identified by computational approaches. Several web-based or non-web-based computer software programs are publicly available for predicting miRNAs and their targets.

Animals↗
PubMed2005

Current efforts in the analysis of RNAi and RNAi target genes.

RNAi is RNA interference by short RNAs. It influences gene-expression by down-regulation of mRNAs, typically by complementarity to the 3' UTR (untranslated region) of the mRNA. microRNAs (miRNAs) are short RNAs acting as natural RNAi. miRNAs mediate down-regulation of many mRNAs from developmental genes and transcription factor genes. Natural examples for this additional level of post-transcriptional control are increasing. Suitable computer-based search strategies for new miRNA candidates include precursor folding as well as different compositional search strategies. Example programs for this are presented. New own and other data are provided for an overview on such strategies. A strategy feasible in plants for miRNA target identification is direct base pairing of miRNAs to potential mRNA target 3' UTRs. Correct identification in animals usually requires comparative genomics and conserved UTR regions pairing to conserved miRNA substructures. A number of example programs and target examples for these tasks are examined. Finally, strategies and programs for artificial gene silencing by designed RNAi are explained.

Animals↗
PubMed2006

A mutation creating a potential illegitimate microRNA target site in the myostatin gene affects muscularity in sheep.

Texel sheep are renowned for their exceptional meatiness. To identify the genes underlying this economically important feature, we performed a whole-genome scan in a Romanov x Texel F2 population. We mapped a quantitative trait locus with a major effect on muscle mass to chromosome 2 and subsequently fine-mapped it to a chromosome interval encompassing the myostatin (GDF8) gene. We herein demonstrate that the GDF8 allele of Texel sheep is characterized by a G to A transition in the 3' UTR that creates a target site for mir1 and mir206, microRNAs (miRNAs) that are highly expressed in skeletal muscle. This causes translational inhibition of the myostatin gene and hence contributes to the muscular hypertrophy of Texel sheep. Analysis of SNP databases for humans and mice demonstrates that mutations creating or destroying putative miRNA target sites are abundant and might be important effectors of phenotypic variation.

Animals↗
PubMed2006

LNA-modified oligonucleotides mediate specific inhibition of microRNA function.

microRNAs are short, endogenous non-coding RNAs that act as post-transcriptional modulators of gene expression. Important functions for microRNAs have been found in the regulation of development, cellular proliferation and differentiation, while perturbed miRNA expression patterns have been observed in many human cancers. Here we present a method for specific inhibition of miRNA function through interaction with LNA-modified antisense oligonucleotides and report the specificity of this application. We show that LNA-modified oligonucleotides can inhibit exogenously introduced miRNAs with high specificity using a heterologous reporter assay, and furthermore demonstrate their ability to inhibit an endogenous miRNA in Drosophila melanogaster cells, leading to up-regulation of the cognate target protein. The method shows stoichiometric and reliable inhibition of the targeted miRNA and can thus be applied to studies of miRNA functions and validation of putative target genes.

Animals↗
PubMed2004

MicroRNA regulation of NAC-domain targets is required for proper formation and separation of adjacent embryonic, vegetative, and floral organs.

BACKGROUND: MicroRNAs (miRNAs) are approximately 21 nucleotide (nt) RNAs that regulate gene expression in plants and animals. Most known plant miRNAs target transcription factors that influence cell fate determination, and biological functions of miRNA-directed regulation have been reported for four of 15 known microRNA gene families: miR172, miR159, miR165, and miR168. Here, we identify a developmental role for miR164-directed regulation of NAC-domain genes, which encode a family of transcription factors that includes CUP-SHAPED COTYLEDON1 (CUC1) and CUC2. RESULTS: Expression of a miR164-resistant version of CUC1 mRNA from the CUC1 promoter causes alterations in Arabidopsis embryonic, vegetative, and floral development, including cotyledon orientation defects, reduction of rosette leaf petioles, dramatically misshapen rosette leaves, one to four extra petals, and one or two missing sepals. Reciprocally, constitutive overexpression of miR164 recapitulates cuc1 cuc2 double mutant phenotypes, including cotyledon and floral organ fusions. miR164 overexpression also leads to phenotypes not previously observed in cuc1 cuc2 mutants, including leaf and stem fusions. These likely reflect the misregulation of other NAC-domain mRNAs, including NAC1, At5g07680, and At5g61430, for which miR164-directed cleavage products were detected. CONCLUSIONS: These results demonstrate that miR164-directed regulation of CUC1 is necessary for normal embryonic, vegetative, and floral development. They also show that proper miR164 dosage or localization is required for separation of adjacent embryonic, vegetative, and floral organs, thus implicating miR164 as a common regulatory component of the molecular circuitry that controls the separation of different developing organs and thereby exposes a posttranscriptional layer of NAC-domain gene regulation during plant development.

Amino Acid Sequence↗
PubMed2024

RNA-seq reveals differentially expressed lncRNAs and circRNAs and their associated functional network in HTR-8/Svneo cells under hypoxic conditions.

Placental hypoxia is hazardous to maternal health as well as fetal growth and development. Preeclampsia and intrauterine growth restriction are common pregnancy problems, and one of the causes is placental hypoxia. Placental hypoxia is linked to a number of pregnancy illnessesv. To investigate their potential function in anoxic circumstances, we mimicked the anoxic environment of HTR-8/Svneo cells and performed lncRNA and circRNA studies on anoxic HTR-8/Svneo cells using high-throughput RNA sequencing. The miRNA target genes were predicted by integrating the aberrant expression of miRNAs in the placenta of preeclampsia and intrauterine growth restriction, and a ceRNA network map was developed to conduct a complete transcriptomic and bioinformatics investigation of circRNAs and lncRNAs. The signaling pathways in which the genes were primarily engaged were predicted using GO and KEGG analyses. To propose a novel explanation for trophoblastic organism failure caused by lncRNAs and circRNAs in an anoxic environment.

Humans↗
PubMed2005

The developmental miRNA profiles of zebrafish as determined by small RNA cloning.

MicroRNAs (miRNAs) represent a family of small, regulatory, noncoding RNAs that are found in plants and animals. Here, we describe the miRNA profile of the zebrafish Danio rerio resolved in a developmental and cell-type-specific manner. The profiles were obtained from larger-scale sequencing of small RNA libraries prepared from developmentally staged zebrafish, and two adult fibroblast cell lines derived from the caudal fin (ZFL) and the liver epithelium (SJD). We identified a total of 154 distinct miRNAs expressed from 343 miRNA genes. Other experimental/computational sources support an additional 10 miRNAs encoded by 19 genes. The miRNAs can be classified into 87 distinct families. Cross-species comparison indicates that 81 families are conserved in mammals, 17 of which also have at least one member conserved in an invertebrate. Our analysis reveals that the zygotes are essentially devoid of miRNAs and that their expression begins during the blastula period with a zebrafish-specific family of miRNAs encoded by closely spaced multicopy genes. Computational predictions of zebrafish miRNA targets are provided that take into account the depth of evolutionary conservation. Besides miRNAs, we identified a prominent class of repeat-associated small interfering RNAs (rasiRNAs).

Animals↗
PubMed2026

Semaglutide treatment in MOSH is associated with altered DNA methylation patterns of genes related to glycolipid metabolism.

Male obesity-associated secondary hypogonadism(MOSH) is a common disease among severely obese male patients. Although surgical interventions have demonstrated clinical benefits, a subset of patients continue to experience MOSH following surgery. Therefore, this study aims to investigate epigenetic changes associated with the use of the weight-loss drug Semaglutide in MOSH, focusing on DNA methylation and miRNA expression. In this exploratory study, samples were classified into three groups: a control group (n = 2), a MOSH group (n = 7), and a follow-up group (n = 4). DNA methylation analysis was performed on all samples, while miRNA sequencing was conducted on a subset of the samples: 2 from the control group, 7 from the MOSH group, and 2 from the follow-up group. Differentially expressed miRNAs (DEMs) were analyzed through the R package "limma", and the methylation level of CpG sites was analyzed based on the methylation β value, obtaining differentially methylated genes (DMGs). The functional enrichment analysis of miRNA target genes and methylation change genes was conducted using the R package "clusterProfiler". Finally, the regulatory networks of miRNA and methylation genes as well as the protein-protein interaction (PPI) network were analyzed. A total of 6 DEMs were screened out. The target genes of these DEMs were mainly enriched in pathways such as ATP binding, phosphorylation, cell adhesion, and Glycosphingolipid biosynthesis. Eighty DMGs were identified, and the largest number of DMGs were found in the X chromosome. In the regulatory network of DMGs and DEMs, hsa-miR-423-5p regulates most of these DMGs. Moreover, the PPI network shows that DPP6, DPP10, CACNA1C, and CNTNAP2 are the proteins with the strongest connectivity. Notably, differential CpG methylation changes were observed on chromosome 7, indicating a potential region of epigenetic alteration in MOSH; however, the biological and functional relevance of these changes remains unclear. Collectively, these findings suggest that Semaglutide treatment in MOSH may be associated with concurrent alterations in DNA methylation and miRNA expression, implicating genes related to energy and glycolipid metabolism, including DPP6, DPP10, CACNA1C, and CNTNAP2. These results are exploratory and hypothesis-generating, providing preliminary observations to inform future validation studies.

Semaglutide↗
PubMed2006

The diverse functions of microRNAs in animal development and disease.

MicroRNAs (miRNAs) control gene expression by translational inhibition and destabilization of mRNAs. While hundreds of miRNAs have been found, only a few have been studied in detail. miRNAs have been implicated in tissue morphogenesis, cellular processes like apoptosis, and major signaling pathways. Emerging evidence suggests a direct link between miRNAs and disease, and miRNA expression signatures are associated with various types of cancer. In addition, the gain and loss of miRNA target sites appears to be causal to some genetic disorders. Here, we discuss the current literature on the role of miRNAs in animal development and disease.

Animal Diseases↗
PubMed2006

AGO1 homeostasis entails coexpression of MIR168 and AGO1 and preferential stabilization of miR168 by AGO1.

Arabidopsis ARGONAUTE1 (AGO1) encodes the RNA slicer enzyme of the microRNA (miRNA) pathway and is regulated by miR168-programmed, AGO1-catalyzed mRNA cleavage. Here, we describe two additional regulatory processes required for AGO1 homeostasis: transcriptional coregulation of MIR168 and AGO1 genes, and posttranscriptional stabilization of miR168 by AGO1. Disrupting any of these regulatory processes by using mutations or transgenes disturbs a proper functioning of the miRNA pathway. In contrast, minor perturbation leads to fine-tuned posttranscriptional adjustment of miR168 and AGO1 levels, thereby maintaining a proper balance of other miRNAs, which, together with AGO1, control the mRNA levels of miRNA targets. We suggest that miR168 stabilization occurs at the level of silencing-complex assembly and that modulating the efficiency of assembling miRNA-programmed silencing complexes will also be important in other contexts.

Arabidopsis↗
PubMed2005

MicroRNA-dependent trans-acting siRNA production.

Less than a year has elapsed between the discovery of trans-acting small interfering RNAs (tasiRNAs) in plants and the elucidation of the major steps of the corresponding pathway. During tasiRNA biogenesis, polyadenylated RNAs transcribed from non-protein-coding TAS genes are cleaved by a microRNA (miRNA)-programmed RNA-induced silencing complex. In contrast to classical miRNA targets, RDR6 and SGS3 convert one of the TAS RNA cleavage products into double-stranded RNA, which is subsequently processed, in a phase determined by the initial miRNA cleavage site, by DICER-LIKE 4 to generate a 21-nucleotide tasiRNA population. tasiRNAs guide endogenous mRNA cleavage through the action of AGO1 or, perhaps in some cases, AGO7. Some of the tasiRNA targets probably regulate the juvenile-to-adult phase transition, but the roles of other tasiRNA targets remain to be determined.

Arabidopsis↗
PubMed2006

miTarget: microRNA target gene prediction using a support vector machine.

BACKGROUND: MicroRNAs (miRNAs) are small noncoding RNAs, which play significant roles as posttranscriptional regulators. The functions of animal miRNAs are generally based on complementarity for their 5' components. Although several computational miRNA target-gene prediction methods have been proposed, they still have limitations in revealing actual target genes. RESULTS: We implemented miTarget, a support vector machine (SVM) classifier for miRNA target gene prediction. It uses a radial basis function kernel as a similarity measure for SVM features, categorized by structural, thermodynamic, and position-based features. The latter features are introduced in this study for the first time and reflect the mechanism of miRNA binding. The SVM classifier produces high performance with a biologically relevant data set obtained from the literature, compared with previous tools. We predicted significant functions for human miR-1, miR-124a, and miR-373 using Gene Ontology (GO) analysis and revealed the importance of pairing at positions 4, 5, and 6 in the 5' region of a miRNA from a feature selection experiment. We also provide a web interface for the program. CONCLUSION: miTarget is a reliable miRNA target gene prediction tool and is a successful application of an SVM classifier. Compared with previous tools, its predictions are meaningful by GO analysis and its performance can be improved given more training examples.

Algorithms↗
PubMed2003

Identification of Drosophila MicroRNA targets.

MicroRNAs (miRNAs) are short RNA molecules that regulate gene expression by binding to target messenger RNAs and by controlling protein production or causing RNA cleavage. To date, functions have been assigned to only a few of the hundreds of identified miRNAs, in part because of the difficulty in identifying their targets. The short length of miRNAs and the fact that their complementarity to target sequences is imperfect mean that target identification in animal genomes is not possible by standard sequence comparison methods. Here we screen conserved 3' UTR sequences from the Drosophila melanogaster genome for potential miRNA targets. The screening procedure combines a sequence search with an evaluation of the predicted miRNA-target heteroduplex structures and energies. We show that this approach successfully identifies the five previously validated let-7, lin-4, and bantam targets from a large database and predict new targets for Drosophila miRNAs. Our target predictions reveal striking clusters of functionally related targets among the top predictions for specific miRNAs. These include Notch target genes for miR-7, proapoptotic genes for the miR-2 family, and enzymes from a metabolic pathway for miR-277. We experimentally verified three predicted targets each for miR-7 and the miR-2 family, doubling the number of validated targets for animal miRNAs. Statistical analysis indicates that the best single predicted target sites are at the border of significance; thus, target predictions should be considered as tentative until experimentally validated. We identify features shared by all validated targets that can be used to evaluate target predictions for animal miRNAs. Our initial evaluation and experimental validation of target predictions suggest functions for two miRNAs. For others, the screen suggests plausible functions, such as a role for miR-277 as a metabolic switch controlling amino acid catabolism. Cross-genome comparison proved essential, as it allows reduction of the sequence search space. Improvements in genome annotation and increased availability of cDNA sequences from other genomes will allow more sensitive screens. An increase in the number of confirmed targets is expected to reveal general structural features that can be used to improve their detection. While the screen is likely to miss some targets, our study shows that valid targets can be identified from sequence alone.

3' Untranslated Regions↗
PubMed2026

miR-6388 regulates granulosa cell function in sheep by targeting GDF9 and modulating the TGF-β signaling pathway.

Litter size is an economically important trait in sheep and is closely associated with ovarian follicular development and granulosa cell (GC) function. This study investigated the association between GDF9 polymorphisms and litter size, and examined the post-transcriptional regulation of GDF9 by miR-6388 in ovine GCs. Variants were initially identified by Sanger sequencing in 20 ewes, and subsequently genotyped in 377 three-year-old ewes, including 231 Sonid (SN) sheep and 146 Ujimqin (UM) sheep for association analysis. Candidate miRNAs targeting litter size-associated variants in the GDF9 3'UTR were predicted, and the miR-6388-GDF9 interaction was evaluated using dual-luciferase reporter assays. RT-qPCR, Western blotting, EdU incorporation, and flow cytometry were used to assess endogenous GDF9 expression and GC function. Twelve single-nucleotide polymorphisms were identified, including the putatively novel variant g.42114076C > G. The linkage disequilibrium block comprising g.42116936C > T, g.42113821T > A, and g.42113962G > A polymorphisms of GDF9 was significantly associated with litter size in both SN and UM sheep, whereas the c.477G > A was associated with litter size only in UM sheep. Reporter assays showed that the GDF9 3'UTR region carrying the G allele of g.42113962G > A was more responsive to miR-6388-mediated repression than the region carrying the A allele. miR-6388 overexpression reduced GDF9 mRNA and GDF9 protein levels, inhibited GC proliferation, altered cell-cycle distribution, and promoted apoptosis, whereas miR-6388 knockdown increased GDF9 expression and GC proliferation and reduced apoptosis. These cellular changes were accompanied by altered expression of cell-cycle and apoptosis-related genes and TGF-β signaling-related components.

Animals↗
PubMed2025

De novo transcriptome assembly and gene expression analysis of Cnidium officinale under high-temperature conditions.

BACKGROUND: The medicinal plant Cnidium officinale (CO) is widespread in Northeast Asia and vulnerable to heat stress. The naturally occurring composition of pharmacological ingredients of CO results in overall physiological consequences; therefore, it is crucial to have a comprehensive understanding of metabolic response to ambient heat in terms of acclimation to estimate how much CO is exposed to threatening environmental conditions. RESULTS: Transcriptome analysis is critical for understanding the consequences of long-term physiological adaptation of CO to abiotic stress. However, transcriptome analysis on this species, particularly under prolonged stress conditions, has remained limited. We employed a temperature gradient tunnel (TGT) to subject CO to high-temperature exposure for four months, enabling us to observe the cumulative effects of heat and assess its acclimation mechanisms. In the absence of genome sequencing data, we performed de novo transcriptome assembly and compared DEGs from temperature treatment plots of a TGT and a growth chamber (GC). Since interpreting transcriptomic data can be complex, we employed a sequential analytical approach, including DEG clustering, GO enrichment, KEGG pathway mapping, miRNA-target gene analysis, and multiple rounds of RNA sequencing validation. DEGs were classified into two categories: genes exhibiting significant fold changes and genes showing significant count changes rather than fold changes. Then, we analyzed the functional roles of DEGs to determine which pathways respond to ambient and stressful high temperatures and validated the findings through cross-comparison with GC. Additionally, we conducted miRNA analysis to investigate post-transcriptional regulation under high temperatures. CO grown under higher ambient temperatures exhibited slight upregulation of pathways related to protein stability and turnover, ABA biosynthesis, and energy production, such as photosynthesis and oxidative phosphorylation. However, under extreme heat stress, most metabolic pathways were downregulated except for those involved in transcription, translation, oxidative phosphorylation and the biosynthesis of cutin, suberin, and wax. CONCLUSION: This study demonstrated that proper clustering of genes based on expression levels and fold changes in two different experimental conditions, along with pathway mapping, may provide a comprehensive understanding of CO's response to heat stress. These insights could contribute to future research on heat tolerance and crop improvement.

Gene Expression Profiling↗
PubMed2004

HIV-1 encoded candidate micro-RNAs and their cellular targets.

MicroRNAs (miRNAs) are small RNAs of 21-25 nucleotides that specifically regulate cellular gene expression at the post-transcriptional level. miRNAs are derived from the maturation by cellular RNases III of imperfect stem loop structures of ~ 70 nucleotides. Evidence for hundreds of miRNAs and their corresponding targets has been reported in the literature for plants, insects, invertebrate animals, and mammals. While not all of these miRNA/target pairs have been functionally verified, some clearly serve roles in regulating normal development and physiology. Recently, it has been queried whether the genome of human viruses like their cellular counterpart also encode miRNA. To date, there has been only one report pertaining to this question. The Epstein-Barr virus (EBV) has been shown to encode five miRNAs. Here, we extend the analysis of miRNA-encoding potential to the human immunodeficiency virus (HIV). Using computer-directed analyses, we found that HIV putatively encodes five candidate pre-miRNAs. We then matched deduced mature miRNA sequences from these 5 pre-miRNAs against a database of 3' untranslated sequences (UTR) from the human genome. These searches revealed a large number of cellular transcripts that could potentially be targeted by these viral miRNA (vmiRNA) sequences. We propose that HIV has evolved to use vmiRNAs as a means to regulate cellular milieu for its benefit.

3' Untranslated Regions↗