Paired-end small RNA sequencing reveals a possible overestimation in isomiR sequence repertoire previously reported from conventional single read data analysis
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE155370 | GEO | 2021/03/24
REPOSITORIES: GEO
Similar Datasets
Project description:We have sequenced miRNA libraries from human embryonic, neural and foetal mesenchymal stem cells. We report that the majority of miRNA genes encode mature isomers that vary in size by one or more bases at the 3’ and/or 5’ end of the miRNA. Northern blotting for individual miRNAs showed that the proportions of isomiRs expressed by a single miRNA gene often differ between cell and tissue types. IsomiRs were readily co-immunoprecipitated with Argonaute proteins in vivo and were active in luciferase assays, indicating that they are functional. Bioinformatics analysis predicts substantial differences in targeting between miRNAs with minor 5’ differences and in support of this we report that a 5’ isomiR-9-1 gained the ability to inhibit the expression of DNMT3B and NCAM2 but lost the ability to inhibit CDH1 in vitro. This result was confirmed by the use of isomiR-specific sponges. Our analysis of the miRGator database indicates that a small percentage of human miRNA genes express isomiRs as the dominant transcript in certain cell types and analysis of miRBase shows that 5’ isomiRs have replaced canonical miRNAs many times during evolution. This strongly indicates that isomiRs are of functional importance and have contributed to the evolution of miRNA genes
2014-08-06 | GSE58734 | GEO
Project description:MicroRNAs (miRNA) have alternative forms known as isomiRs, which differ from each other by a few nucleotides. Next generation sequencing platforms facilitate identification of these isomiRs and recent discoveries regarding their functional importance have increased our understandings of the regulatory complexities of the microRNAome. Observed changes in the miRNA profiles in mosquitoes infected with flaviviruses have implicated small RNAs in the interactions between viruses and their vectors. Here we analysed the isomiR profiles of both uninfected and infected blood fed Aedes aegypti mosquitoes with a major human pathogen, Dengue virus at two time points post-infection. We found noticeable changes to the isomiR expression profile in response to infection and aging. Data analysis revealed a distinct bias towards isomiR production in the mature miRNA as opposed to the star strand. Furthermore, we noticed that only in 40% of Ae. aegypti miRNAs, the most abundant reads for each particular miRNA match the exact sequence reported in the miRbase. The isomiR expression variations between an Ae. aegypti embryonic cell line (Aag2) and whole mosquitoes demonstrated a tissue-specific pattern of isomiR production. Our results illustrated a bias for certain types of isomiRs for each miRNA. The findings presented in this study also provide evidence that isomiR production is not a random phenomenon and may be important in DENV colonisation of its vector. Examination of isomiR production rate in DENV infected and non infected mosquitoes
2015-11-05 | E-GEOD-59516 | biostudies-arrayexpress
Project description:We have sequenced miRNA libraries from human embryonic, neural and foetal mesenchymal stem cells. We report that the majority of miRNA genes encode mature isomers that vary in size by one or more bases at the 3’ and/or 5’ end of the miRNA. Northern blotting for individual miRNAs showed that the proportions of isomiRs expressed by a single miRNA gene often differ between cell and tissue types. IsomiRs were readily co-immunoprecipitated with Argonaute proteins in vivo and were active in luciferase assays, indicating that they are functional. Bioinformatics analysis predicts substantial differences in targeting between miRNAs with minor 5’ differences and in support of this we report that a 5’ isomiR-9-1 gained the ability to inhibit the expression of DNMT3B and NCAM2 but lost the ability to inhibit CDH1 in vitro. This result was confirmed by the use of isomiR-specific sponges. Our analysis of the miRGator database indicates that a small percentage of human miRNA genes express isomiRs as the dominant transcript in certain cell types and analysis of miRBase shows that 5’ isomiRs have replaced canonical miRNAs many times during evolution. This strongly indicates that isomiRs are of functional importance and have contributed to the evolution of miRNA genes Sequence library of miRNAs from a single sample of human foetal mesenchymal stem cells. Results tested and confirmed by northern blotting. Please note that only raw data files are available for the embryonic and neual samples and thus, directly submitted to SRA (SRX547311, SRX548700, respectively under SRP042115/PRJNA247767)
2014-08-06 | E-GEOD-58734 | biostudies-arrayexpress
Project description:Purpose: We carried out small RNA sequencing to determine miRNA expression patterns of hippocampus and frontal cortex in six inbred mouse strains. We analyzed miRNA expression levels, isomiR distribution and miRNA editing events. Methods: We constructed small RNA libraries starting from the total RNA extracted from the hippocampus and frontal cortex of three animals per strain. The sequencing was carried out using Illumina HiSeq2000 (single-end). The sequence reads that passed quality filters were mapped to the mouse genome (mm10) and to known miRNAs (miRBase v21) using miRDeep2, SeqBuster, and sRNAbench, to obtain miRNA and isomiR expression levels and to predict novel miRNAs. Genome alignments were analysed for miRNA editing using FreeBayes. Results: We detected significant miRNA and isomiR expression level differences between the strains and brain regions. Most of the miRNA loci produced a number of isomiRs. Some miRNAs were consistently edited with a pattern that matches the activity of known RNA editing enzymes.
2018-01-18 | GSE84408 | GEO
Project description:MicroRNAs (miRNA) have alternative forms known as isomiRs, which differ from each other by a few nucleotides. Next generation sequencing platforms facilitate identification of these isomiRs and recent discoveries regarding their functional importance have increased our understandings of the regulatory complexities of the microRNAome. Observed changes in the miRNA profiles in mosquitoes infected with flaviviruses have implicated small RNAs in the interactions between viruses and their vectors. Here we analysed the isomiR profiles of both uninfected and infected blood fed Aedes aegypti mosquitoes with a major human pathogen, Dengue virus at two time points post-infection. We found noticeable changes to the isomiR expression profile in response to infection and aging. Data analysis revealed a distinct bias towards isomiR production in the mature miRNA as opposed to the star strand. Furthermore, we noticed that only in 40% of Ae. aegypti miRNAs, the most abundant reads for each particular miRNA match the exact sequence reported in the miRbase. The isomiR expression variations between an Ae. aegypti embryonic cell line (Aag2) and whole mosquitoes demonstrated a tissue-specific pattern of isomiR production. Our results illustrated a bias for certain types of isomiRs for each miRNA. The findings presented in this study also provide evidence that isomiR production is not a random phenomenon and may be important in DENV colonisation of its vector.
2015-11-05 | GSE59516 | GEO
Project description:miRNAs are small noncoding RNA molecules that play an important role in post-transcriptional regulation of gene expression. Length and/or sequence variants of the same miRNA are termed isomiRs. While most isomiRs are functionally redundant compared to their canonical counterparts, so-called 5âisomiRs exhibit a shifted 5â end and therefore a shifted seed sequence resulting in a different target spectrum. However, not much is known about the functional relevance of these isoforms. Analysis of miRNA-seq data from breast cancer cell lines identified six pairs of highly expressed miRNAs and associated 5âisomiRs. Among them, hsa-miR-140-3p was of particular interest because its 5âisomiR showed higher expression compared to the canonical miRNA annotated in miRbase. This miRNA has previously been shown to control stemness of breast cancer cells. MiRNAseq data of breast cancer patients (TCGA dataset) showed that both the canonical hsa-miR-140-3p and its 5âisomiR-140-3p were highly expressed in patients compared to normal breast tissue. In the current work, we present the functional characterization of 5âisomiR-140-3p and the cellular phenotypes associated with its overexpression in MCF10A and MDA-MB-231 cell lines in comparison to the canonical hsa-miR-140-3p. Contrary to the effect of the canonical hsa miR 140-3p, overexpression of the 5âisomiR-140-3p led to a decrease in cell viability. The latter observation was supported by cell cycle analysis, where the 5âisomiR-140-3p but not the hsa-miR-140-3p caused cell cycle arrest in G0/G1-phase. Additionally, 5âismoiR-140-3p overexpression was found to cause a decrease in cell migration in MCF10A cells. We identified three novel direct target genes of the 5â isomiR-140-3p; COL4A1, ITGA6 and MARCKSL1. Finally, we have shown that knocking down these genes partially phenocopied the effects of the 5âisomiR-140-4p overexpression, where COL4A1 and ITGA6 knockdown led to reduced cell viability and cell cycle arrest, while MARCKSL1 knockdown resulted in a decrease in the migratory potential of cells. In summary, this work presents evidence that there is a functional synergy between the canonical hsa-miR-140-3p and the newly identified 5âisomiR-140-3p in suppressing growth and progression of breast cancer by simultaneously targeting genes related to differentiation, proliferation, and migration. With this array, we aimed to address the question which genes are regulated by either of the two forms of the miRNA. Samples were measured in one biological replicate of cells transfected with mimic-ctrl1 and mimic-ctrl2 (Dharmacon) as control samples and two biological replicates of cells transfected with hsa-miR-140-3p and 5'isomiR-140-3p (Exiqon) in 30nM concentration using Lipofectamin 2000 as transfection reagent.
2015-10-31 | E-GEOD-74539 | biostudies-arrayexpress
Project description:TUT4 and TUT7 are terminal uridyl transferases that adds uridine bases to the 3′ end of RNAs such as miRNAs and mRNAs. The overall aim of this experiment was to identify differentially regulated mRNAs and miRNAs in catalytic knockouts of TUT4/7 relative to the wildtype. Catalytic mutants of TUT4/7 was generated in the prostate cancer cell line DU145 and the ovarian cancer cell line IGROV1. As the miRNA regulation by TUT4/7 depends on the oncoprotein LIN28A, LIN28A was lentivirally integrated in the LIN28A- DU145-derived lines and control. IGROV1 endogenously expresses LIN28A. We report that excluding the let-7 family, miRNAs do not depend on LIN28A for TUT4/7-mediated regulation. As the entire miRNA is captured in a read, we provide the isomiR landscape in the presence and absence of TUT4/7. Loss of uridylated isomiRs results in a simultaneous gain in adenylated isomiRs. Finally, we report cell line specific miRNA-mRNA regulation.
2021-07-09 | GSE179741 | GEO
Project description:Increasing evidence suggest anti-senescence and geroprotective roles for the glucose-lowering drug metformin. In this framework, the ability of metformin to affect the biogenesis of selected microRNAs (miRNAs) was recently suggested. IsomiRs are distinct variations of miRNA sequences, such as addition or deletion of one or more nucleotides at the 5’ and/or 3’ ends of the canonical miRNA sequence. Since no study has comprehensively assessed the miRNA signatures, including isomiR variants, affected by metformin treatment during cellular senescence, we analysed miRNA and isomiR expression in human umbilical vein endothelial cells (HUVECs) undergoing replicative senescence in presence of metformin.
2020-12-03 | GSE149771 | GEO
Project description:miRNAs are small noncoding RNA molecules that play an important role in post-transcriptional regulation of gene expression. Length and/or sequence variants of the same miRNA are termed isomiRs. While most isomiRs are functionally redundant compared to their canonical counterparts, so-called 5’isomiRs exhibit a shifted 5’ end and therefore a shifted seed sequence resulting in a different target spectrum. However, not much is known about the functional relevance of these isoforms. Analysis of miRNA-seq data from breast cancer cell lines identified six pairs of highly expressed miRNAs and associated 5’isomiRs. Among them, hsa-miR-140-3p was of particular interest because its 5’isomiR showed higher expression compared to the canonical miRNA annotated in miRbase. This miRNA has previously been shown to control stemness of breast cancer cells. MiRNAseq data of breast cancer patients (TCGA dataset) showed that both the canonical hsa-miR-140-3p and its 5’isomiR-140-3p were highly expressed in patients compared to normal breast tissue. In the current work, we present the functional characterization of 5’isomiR-140-3p and the cellular phenotypes associated with its overexpression in MCF10A and MDA-MB-231 cell lines in comparison to the canonical hsa-miR-140-3p. Contrary to the effect of the canonical hsa miR 140-3p, overexpression of the 5’isomiR-140-3p led to a decrease in cell viability. The latter observation was supported by cell cycle analysis, where the 5’isomiR-140-3p but not the hsa-miR-140-3p caused cell cycle arrest in G0/G1-phase. Additionally, 5’ismoiR-140-3p overexpression was found to cause a decrease in cell migration in MCF10A cells. We identified three novel direct target genes of the 5’ isomiR-140-3p; COL4A1, ITGA6 and MARCKSL1. Finally, we have shown that knocking down these genes partially phenocopied the effects of the 5’isomiR-140-4p overexpression, where COL4A1 and ITGA6 knockdown led to reduced cell viability and cell cycle arrest, while MARCKSL1 knockdown resulted in a decrease in the migratory potential of cells. In summary, this work presents evidence that there is a functional synergy between the canonical hsa-miR-140-3p and the newly identified 5’isomiR-140-3p in suppressing growth and progression of breast cancer by simultaneously targeting genes related to differentiation, proliferation, and migration. With this array, we aimed to address the question which genes are regulated by either of the two forms of the miRNA.
2015-10-31 | GSE74539 | GEO
Project description:MicroRNAs (miRNAs) are small RNAs of ~22 nucleotides in length that are involved in the regulation of a variety of physiological and pathological processes. Advances in high-throughput small RNA sequencing (smRNA-seq), one of the next generation sequencing applications, have reshaped the miRNA research landscape. In this study, we established an integrative database containing analysis pipelines and analysis results of 609 human and mice smRNA-seq results, including public data from GEO and private ones. YM500 collects analysis results for miRNA quantification, for isomiR identification (incl. RNA editing), for arm switching discovery, and, more importantly, for novel miRNA predictions. Wetlab validation on >100 miRNAs confirmed high correlation between miRNA profiling and RT-qPCR results (R=0.84). This database allows researchers to search these 4 different types of analysis results via our interactive web interface. YM500 allows researchers to define the criteria of isomiRs, as well as integrates the information of dbSNP to help researchers to distinguish isomiRs from SNPs. A user-friendly interface is provided to integrate miRNA-related information and existing evidences from hundreds of sequencing datasets. The identified novel miRNAs and isomiRs hold the potentials for both basic research and biotech applications. YM500 is now available on http://ngs.ym.edu.tw/ym500/. There are 34 in-house datasets in YM500 . They are 9 human embryonal tumors, 9 human germ cell tumors, and 16 human gliomas.
2012-12-03 | E-GEOD-39841 | biostudies-arrayexpress