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Gene Apr 2013To search for genetic regulators influencing miRNA transcript abundance, we performed a genome-wide association study (GWAS) to identify quantitative trait loci...
To search for genetic regulators influencing miRNA transcript abundance, we performed a genome-wide association study (GWAS) to identify quantitative trait loci associated with primary miRNA transcript abundance (pri-miQTL) using genotype data from HapMap CEU phased data. We detected robust expression for 150 pri-miRNAs out of 1523 interrogated using RNA-seq. We have identified some pri-miRNAs that showed significant evidence for cis- (34%) and trans-pri-miQTLs (3%). Furthermore, we observed that multiple cis-pri-miQTLs, showed allele-specific expression associated with single pri-miRNA expression. Interestingly, a cis-regulatory variation influenced the expression levels of two pri-miRNAs that expressed identical mature sequences. We also observed that a single trans-regulatory variation was associated with multiple unrelated pri-miRNAs: rs292253 was associated with the expression of hsa-mir-3181, hsa-mir-3665 and hsa-mir-762. These findings revealed that the expression of pri-miRNA detected by RNA-seq can be used to identify pri-miQTLs, as an alternative method to dissect the genetic control mechanisms governing pri-miRNA expression.
Topics: Cell Line, Tumor; Genome-Wide Association Study; HapMap Project; Humans; MicroRNAs; Polymorphism, Single Nucleotide; Quantitative Trait Loci; Sequence Analysis, RNA
PubMed: 23291411
DOI: 10.1016/j.gene.2012.08.015 -
Adipocyte Dec 2021Intramuscular fat, as one of the most important palatability attribute of beef carcase, is the primary determinant of beef quality. The research of adipogenesis...
Intramuscular fat, as one of the most important palatability attribute of beef carcase, is the primary determinant of beef quality. The research of adipogenesis mechanism would provide new insight into intramuscular fatty deposition. Here, the role of microRNA-378 was investigated during bovine adipogenic differentiation. It was revealed that miR-378 expression exists variably in bovine major tissue and organs by RT-qPCR. It was predicted that miR-378 targets CaMKK2, as an AMPKα kinase, by DIANA Tools. For better research, primary preadipocytes with stable transfection for up-/down-regulated expression of miR-378 were constructed by lentiviral vectors with GFP gene. The analyses of qPCR showed that and mRNA levels increased, but and mRNA levels decreased during adipogenic differentiation. When miR-378 was overexpressed, preadipocytes proliferation became slower, there are more cellular lipid droplets, and and mRNA levels were higher, but and were lower than control groups. Luciferase assay and western blot analysis validated that miR-378 binds the nucleotide sites of the 3'- untranslated region of , which inhibits the mRNA and protein expression of . These findings suggest that miR-378 promotes adipogenic differentiation in bovine intramuscular preadipocytes by targeting via AMPK signalling pathway.
Topics: Adipocytes; Adipogenesis; Animals; Cattle; Cell Differentiation; MicroRNAs; PPAR gamma
PubMed: 34693860
DOI: 10.1080/21623945.2021.1982526 -
Methods in Molecular Biology (Clifton,... 2018In the genome, primary microRNAs (pri-miRNAs) are encoded either as independent transcriptional units with their own promoters (intergenic miRNAs) or within the introns...
In the genome, primary microRNAs (pri-miRNAs) are encoded either as independent transcriptional units with their own promoters (intergenic miRNAs) or within the introns of other genes (intronic miRNAs). Here, we report two methods, one that we established for coupled RNAP II transcription and pri-miRNA processing and the other that is a three-way system for RNAP II transcription, pri-miRNA processing, and pre-mRNA splicing. In these systems, CMV-DNA constructs encoding the processing substrates are incubated in HeLa cell nuclear extracts in the presence of P-UTP to generate the nascent RNAP II transcripts, which are processed efficiently by the endogenous RNA processing machineries in nuclear extracts.
Topics: Cell-Free System; HeLa Cells; Humans; MicroRNAs; RNA Polymerase II; RNA Precursors; RNA Splicing; Transcription, Genetic
PubMed: 29959672
DOI: 10.1007/978-1-4939-8624-8_4 -
Nucleic Acids Research Jan 2011miRBase is the primary online repository for all microRNA sequences and annotation. The current release (miRBase 16) contains over 15,000 microRNA gene loci in over 140...
miRBase is the primary online repository for all microRNA sequences and annotation. The current release (miRBase 16) contains over 15,000 microRNA gene loci in over 140 species, and over 17,000 distinct mature microRNA sequences. Deep-sequencing technologies have delivered a sharp rise in the rate of novel microRNA discovery. We have mapped reads from short RNA deep-sequencing experiments to microRNAs in miRBase and developed web interfaces to view these mappings. The user can view all read data associated with a given microRNA annotation, filter reads by experiment and count, and search for microRNAs by tissue- and stage-specific expression. These data can be used as a proxy for relative expression levels of microRNA sequences, provide detailed evidence for microRNA annotations and alternative isoforms of mature microRNAs, and allow us to revisit previous annotations. miRBase is available online at: http://www.mirbase.org/.
Topics: Databases, Nucleic Acid; MicroRNAs; Sequence Analysis, RNA; Systems Integration
PubMed: 21037258
DOI: 10.1093/nar/gkq1027 -
Molecular Therapy : the Journal of the... Dec 2015Heart failure (HF) is the end result of a diverse set of causes such as genetic cardiomyopathies, coronary artery disease, and hypertension and represents the primary... (Review)
Review
Heart failure (HF) is the end result of a diverse set of causes such as genetic cardiomyopathies, coronary artery disease, and hypertension and represents the primary cause of hospitalization in Europe. This serious clinical disorder is mostly associated with pathological remodeling of the myocardium, pump failure, and sudden death. While the survival of HF patients can be prolonged with conventional pharmacological therapies, the prognosis remains poor. New therapeutic modalities are thus needed that will target the underlying causes and not only the symptoms of the disease. Under chronic cardiac stress, small noncoding RNAs, in particular microRNAs, act as critical regulators of cardiac tissue remodeling and represent a new class of therapeutic targets in patients suffering from HF. Here, we focus on the potential use of microRNA inhibitors as a new treatment paradigm for HF.
Topics: Animals; Antisense Elements (Genetics); Disease Models, Animal; Gene Expression Regulation; Heart Failure; Humans; MicroRNAs; Myocardium
PubMed: 26216517
DOI: 10.1038/mt.2015.133 -
Experimental Hematology Nov 2007Expression profiling of microRNA (miRNA) was performed in granulocytes isolated from patients with primary myelofibrosis (PMF), with the aim of identifying abnormally...
OBJECTIVE
Expression profiling of microRNA (miRNA) was performed in granulocytes isolated from patients with primary myelofibrosis (PMF), with the aim of identifying abnormally expressed miRNAs in comparison with normal subjects or patients with polycythemia vera (PV) or essential thrombocythemia (ET).
PATIENTS AND METHODS
Using stem loop-primed reverse transcription and TaqMan quantitative real-time polymerase chain reaction, the expression of 156 mature miRNAs was evaluated using pooled granulocytes from PMF patients, either wild-type or JAK2(617V>F) mutant with >51% allele burden, and control subjects. Differentially expressed miRNAs were then validated on additional control and PMF samples, and also on PV or ET granulocytes.
RESULTS
There was a global downregulation of miRNA expression in PMF granulocytes; 60 miRNAs, of 128 called present, displayed differential expression compared to normal samples. Twelve miRNAs, which had been selected based on statistically different expression level, were finally validated. In PMF granulocytes, levels of miR-31, -150, and -95 were significantly lower, while those of miR-190 significantly greater, than control and PV or ET samples; on the other hand, miR-34a, -342, -326, -105, -149, and -147 were similarly reduced in patients with PMF, PV, or ET compared to controls. Increased expression of miR-182 and -183 correlated with JAK2(617V>F) allele burden. Three in silico-predicted putative target genes (DTR, HMGA2, and MYB), showed deregulated expression in PMF granulocytes that correlated with expression level of regulatory miRNA.
CONCLUSIONS
A defined miRNA profile distinguishes PMF granulocytes from those of normal subjects and, partially, also from PV or ET patients.
Topics: Case-Control Studies; Gene Expression Profiling; Gene Expression Regulation; Granulocytes; Humans; Janus Kinase 2; MicroRNAs; Primary Myelofibrosis
PubMed: 17976522
DOI: 10.1016/j.exphem.2007.08.020 -
Molecules and Cells Dec 2013MicroRNAs are short 21-22 nucleotide single strand RNAs that are involved in post-transcriptional regulation of gene expression. Most microRNAs are first transcribed as...
MicroRNAs are short 21-22 nucleotide single strand RNAs that are involved in post-transcriptional regulation of gene expression. Most microRNAs are first transcribed as long primary microRNAs and then undergo a two step-wise sequential processing to yield single-stranded mature microRNAs. It has been suggested that the loop region of primary microRNAs plays an important role in regulating microRNA biogenesis and target recognition. However, despite the fact that several single nucleotide polymorphisms have been identified in mature microRNA sequences and are related to human diseases, it remains unclear whether and how the single nucleotide polymorphisms in the loop regions of primary microRNAs would affect the biogenesis and function of microRNAs. Herein, we provide evidence that primary microRNAs loop nucleotides control the accuracy and efficiency of microRNA processing. Accordingly, we identified 32 single nucleotide polymorphisms in the loop regions of human primary microRNAs using bioinformatics, and further validated three loss-of-function and one gain-of-function single nucleotide polymorphisms using dual-luciferase assays. Thus, these results reveal a critical regulatory role encoded in the loop nucleotides of primary microRNAs for microRNA processing and function.
Topics: Cell Line; Gene Expression Regulation; Genetic Variation; Humans; MicroRNAs; Nucleic Acid Conformation; Polymorphism, Single Nucleotide; RNA Processing, Post-Transcriptional; RNA, Double-Stranded
PubMed: 24241682
DOI: 10.1007/s10059-013-0171-1 -
PloS One 2012MicroRNAs are known to be generated from primary transcripts mainly through the sequential cleavages by two enzymes, Drosha and Dicer. The sequence of a mature microRNA,...
BACKGROUND
MicroRNAs are known to be generated from primary transcripts mainly through the sequential cleavages by two enzymes, Drosha and Dicer. The sequence of a mature microRNA, especially the 'seeding sequence', largely determines its binding ability and specificity to target mRNAs. Therefore, methods that predict mature microRNA sequences with high accuracy will benefit the identification and characterization of novel microRNAs and their targets, and contribute to inferring the post-transcriptional regulation network at a genome scale.
METHODOLOGY/PRINCIPAL FINDINGS
We have developed a method, MiRmat, to predict the mature microRNA sequence. MiRmat is essentially composed of two parts: the prediction of Drosha processing site and the identification of Dicer processing site. Based on the analysis of microRNAs from 12 species, we found that the patterns of free energy profiles are conserved among vertebrate microRNA hairpins. Therefore, we introduced in our method the free energy distribution pattern of the downstream part of pri-microRNA secondary structure and Random Forest algorithm to predict the mature microRNA sequence. Based on the evaluation on an independent test dataset from 10 vertebrates, MiRmat was shown to identify 77.8% of the Drosha processing sites and 92.8% of the Dicer sites within a deviation of 2 nt. In a more stringent evaluation by excluding the microRNAs sharing the same family between the training set and test set, MiRmat kept a rather well performance of 71.9% and 87.2% of the identification rate on the Drosha and Dicer site respectively, which represents the ability to deal with the novel microRNA family. MiRmat outperforms other state-of-the-art methods and has a high degree of efficacy for the prediction of mature microRNA sequences of vertebrates.
CONCLUSION
MiRmat was developed for identifying microRNA mature sequence(s) by introducing the free energy distribution of RNA stem-loop structure and the Random Forest algorithm. We prove that MiRmat has better performance than the existing tools and is applicable among vertebrates. MiRmat is freely available at http://mcube.nju.edu.cn/jwang/lab/soft/MiRmat/.
Topics: Algorithms; Animals; DEAD-box RNA Helicases; Gene Expression Regulation; Humans; MicroRNAs; RNA, Messenger; Ribonuclease III; Species Specificity
PubMed: 23300555
DOI: 10.1371/journal.pone.0051673 -
Molecular Medicine Reports Sep 2015Alzheimer's disease (AD) is a progressive neurodegenerative disorder and is the most common form of dementia among the aging population. Although the incidence of the...
A single nucleotide polymorphism in primary-microRNA-146a reduces the expression of mature microRNA-146a in patients with Alzheimer's disease and is associated with the pathogenesis of Alzheimer's disease.
Alzheimer's disease (AD) is a progressive neurodegenerative disorder and is the most common form of dementia among the aging population. Although the incidence of the disease continues to increase, no cure has been developed. Effective treatment is restricted not only due to the lack of curative medicine, but also due to limited understanding of the underlying mechanisms and the difficulties in accurately diagnosing AD in its earliest stages prior to clinical symptoms. Micro (mi) RNAs (miR) have gained increasing attention in the investigation of neurodegenerative diseases. Previous reports have demonstrated that deregulation of miR‑146a‑5p is associated with the pathogenesis of human AD. In the present study, the coding region of primary (pri)‑miR‑146a in patients with AD was scanned and the rare C allele of rs2910164 was found to be associated with AD. Using reverse transcription quantitative polymerase chain reaction, it was demonstrated that site variation reduced the expression of mature miR‑146a‑5p. Notably, a reduction in the expression of miR‑146a‑5p led to less efficient inhibition of target genes, including Toll‑like receptor (TLR)2, which is important in the pathogenesis of AD. Biological function investigations in RAW264.7 cells indicated that, compared with the G allele, the rare C allele upregulated the expression of tumor necrosis factor‑α following stimulation with β‑amyloid. These findings suggested that one common polymorphism in pri‑miR‑146a may contribute to the genetic predisposition to AD by disrupting the production of miR‑146a‑5p and affecting the expression and function of TLR2.
Topics: 3' Untranslated Regions; Aged; Aged, 80 and over; Alleles; Alzheimer Disease; Animals; Base Sequence; Case-Control Studies; Cell Line; Female; Genes, Reporter; Genetic Predisposition to Disease; Genotype; HEK293 Cells; Humans; Male; Mice; MicroRNAs; Middle Aged; Polymorphism, Single Nucleotide; Sequence Alignment; Toll-Like Receptor 2; Tumor Necrosis Factor-alpha
PubMed: 26095531
DOI: 10.3892/mmr.2015.3968 -
Biochemical and Biophysical Research... Mar 2011The RNAse III Drosha is responsible for the first step of microRNA maturation, the cleavage of primary miRNA to produce the precursor miRNA. Processing by Drosha is...
The RNAse III Drosha is responsible for the first step of microRNA maturation, the cleavage of primary miRNA to produce the precursor miRNA. Processing by Drosha is finely regulated and influences the amount of mature microRNA in a cell. We describe in the present work a method to quantify Drosha processing activity in-vivo, which is applicable to any microRNA. With respect to other methods for measuring Drosha activity, our system is faster and scalable, can be used with any cellular system and does not require cell sorting or use of radioactive isotopes. This system is useful to study regulation of Drosha activity in physiological and pathological conditions.
Topics: Genes, Reporter; HEK293 Cells; Humans; Luciferases; Methods; MicroRNAs; RNA Processing, Post-Transcriptional; Ribonuclease III
PubMed: 21352811
DOI: 10.1016/j.bbrc.2011.02.055