Pre-microRNA-derived sequences in the supraspliceosome
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE100803 | GEO | 2018/07/01
REPOSITORIES: GEO
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Project description:During microRNA-biogenesis, stem-loop structured precursors are processed in two endonucleolytic steps, giving rise to mature microRNAs (miRNAs). This process is not only regulated at the level of transcription but also posttranscriptionally by RNA-binding proteins (RBPs). Here we have used a proteomics-based pull-down approach to map and characterize the interactome of 72 pre-miRNAs in eleven cell lines. We identify over 150 RBPs that interact specifically with distinct pre-miRNAs. To demonstrate their functional relevance, we used RNAi knockdown and CRISPR/Cas-mediated knockout experiments and analyzed changes in miRNA levels. Indeed, a large number of the investigated candidates have effects on miRNA-processing under our experimental conditions, including several C3H-zinc-finger proteins. Ultimately, we show that TRIM71/Lin41 is a potent regulator of miR-29a processing and its inactivation directly affects miR-29a targets. In summary, we provide an extended database of RBPs that interact with pre-miRNAs in different cell types helping to elucidate posttranscriptional regulation of miRNA biogenesis on a global scale.
2017-04-26 | PXD004193 | Pride
Project description:Hairpin-containing pre-miRNAs are precursors of microRNAs (miRNAs) that play important roles in cellular processes and various human diseases. Pre-miRNAs are produced from longer primary transcripts (pri-miRNAs). The falsity in cellular expression and sequences of pre-miRNAs might cause cellular defects or human diseases. However, the current pre-miRNA quantification methods by qPCR cannot discriminate between pre-miRNAs and their parental pri-miRNAs. In addition, the ligation of the sequencing adapter to 5’-end of pre-miRNAs is inefficient, therefore, pre-miRNA sequencing is highly impractical. Here, we developed a method, called the intramolecular ligation method (iLIME) for pre-miRNA quantification and sequencing. This method utilized T4 RNA ligase 1 to convert hairpin-pre-miRNAs into circularized RNAs that do not naturally exist in cells. The resulting circuliarized RNAs allow us to design unique primers to quantify pre-miRNAs by qPCR specicially, and thus this qPCR can distinguish pre-miRNA from pri-miRNAs. In addition, the iLIME also allows us to sequence pre-miRNAs using next-generation sequencing. The iLIME method offers a simple and effective way to quantify and sequence pre-miRNAs. This will be useful in investigating pre-miRNAs for addressing research questions for medical applications. The iLIME can be potentially extended to other hairpin-containing RNAs, such as tRNAs and snRNAs.
2021-10-11 | GSE185524 | GEO
Project description:Sus scrofa (pig, or swine) is one of the most important economic animals and a close biological model for complex human diseases such as obesity and diabetes. It is therefore utterly important to decode the porcine microRNAome (miRNAome) as in the literature only a small portion of it is known. In this work, a comprehensive search for porcine microRNAs (miRNAs) by Illumina sequencing was performed in ten small RNA libraries prepared from mixtures of assorted tissues, which included those collected from fetuses to adult pigs. The millions of the sequencing reads were analyzed with reference to 77 known porcine miRNA precursors (pre-miRNAs) and 3,443 distinct pre-miRNAs of other mammals listed in miRBase 13.0, and the most updated porcine genome (Sscrofa9, April 2009) and available EST sequences. Additionally, miRNA candidates specific to pig are predicated by genome & EST match and hairpin folding. Our search found 72 out of 78 (~92%) known porcine miRNAs and miRNA*s, and 36 previously unannotated miRNA*s are also indentified. Furthermore, we discovered 397 novel miRNAs by mapping to the sequencing transcripts to other mammalian pre-miRNAs and 493 candidate miRNAs which do not map to other mammalian miRNAomes and could be pig-specific. We constructed sequence- and genome-position clusters for the total of 998 miRNA candidates originating from 862 pre-miRNAs, which represent 777 unique miRNA sequences. Together with the six known porcine miRNAs that not been observed in our study, we report herein the sequence families of 783 unique miRNAs and genomic distribution patterns of 622 pre-miRNAs. We preformed q-PCR experiments for selected 30 miRNAs in 47 tissue-specific samples and found agreement between the sequencing data and the q-PCR data. We envision that our report will serve as a valuable resource for future studies aimed at understanding miRNAome of pig
2010-02-27 | GSE17885 | GEO
Project description:miRNAs are short regulatory single stranded RNA sequences that upon complementary binding to mRNAs lead to the inhibition or degradation of their targets. This regulatory mechanisms has been shown to play crucial roles throughout the whole life cycle of animals and plants as well as in disease. While a plethora of methods exist to predict targets of miRNA, which suggest that up to 80% of the genome is miRNA regulated, it has recently been reported that many of these predictions are false positives, cell type specific or represent non-functional binding. In order to identify the subset of real functional miRNAs and their targets, we established miRNA pathway mutants in mouse embryonic stem cells (mESC), allowing the dissection of canonical and non-canonical functions of pathway members. Additional data integration of downstream regulatory layers (CLIP-seq, ribosome profiling and MS) enabled us to follow and track down real functional miRNA-gene interactions, which reduced the miRNA genome regulation to approximately 1%.
2022-08-31 | PXD014484 | Pride
Project description:In the current study, we investigated the modulatory effect of epigallocatechin gallate (EGCG, an active derivative of green tea) on cellular miRNA expression and cell cycle proliferation in NSCLCs (A549). As the IC50 value of EGCG is approximately 304 μM, our studies support the resistance of A549 lung cancer cell by EGCG. In addition, it was interesting to observe its G0/G1 cell cycle arrest activity at 40μM EGCG treatment. So, in the present study, next-generation sequencing (NGS) was used to study the miRNA expression between the cancer control and EGCG-treated samples. A549 cells were treated with 40 μM and 100 μM EGCG. NGS was employed to study the differential expression of novel and known miRNAs. The data generated was analysed with computational approach to study and correlate the expression profiles of both known and novel miRNAs. Differential expression of known miRNAs was observed between control and EGCG-treated samples. Some putative novel miRNA sequences were also observed in the current study. The differential expression levels of putative novel miRNA sequences were estimated by counting the number of reads in the given dataset. Together the analysis of these miRNA profiles revealed log2-fold modulation in the expression levels. We detected 344, 337, and 346 number of known mature miRNAs in control, 40 μM EGCG treated, and 100 μM EGCG treated samples, respectively. We also reported 119, 126, and 125 number of putative novel mature miRNA sequences in control, 40 μM EGCG treated, and 100 μM EGCG treated samples, respectively. In conclusion, the analysis demonstrates that EGCG is actively involved in miRNA modulation in NSCLCs, which further attests to its chemoprotective role and could play a major role in therapeutic applications.
2019-02-07 | GSE110514 | GEO
Project description:Sus scrofa (pig, or swine) is one of the most important economic animals and a close biological model for complex human diseases such as obesity and diabetes. It is therefore utterly important to decode the porcine microRNAome (miRNAome) as in the literature only a small portion of it is known. In this work, a comprehensive search for porcine microRNAs (miRNAs) by Illumina sequencing was performed in ten small RNA libraries prepared from mixtures of assorted tissues, which included those collected from fetuses to adult pigs. The millions of the sequencing reads were analyzed with reference to 77 known porcine miRNA precursors (pre-miRNAs) and 3,443 distinct pre-miRNAs of other mammals listed in miRBase 13.0, and the most updated porcine genome (Sscrofa9, April 2009) and available EST sequences. Additionally, miRNA candidates specific to pig are predicated by genome & EST match and hairpin folding. Our search found 72 out of 78 (~92%) known porcine miRNAs and miRNA*s, and 36 previously unannotated miRNA*s are also indentified. Furthermore, we discovered 397 novel miRNAs by mapping to the sequencing transcripts to other mammalian pre-miRNAs and 493 candidate miRNAs which do not map to other mammalian miRNAomes and could be pig-specific. We constructed sequence- and genome-position clusters for the total of 998 miRNA candidates originating from 862 pre-miRNAs, which represent 777 unique miRNA sequences. Together with the six known porcine miRNAs that not been observed in our study, we report herein the sequence families of 783 unique miRNAs and genomic distribution patterns of 622 pre-miRNAs. We preformed q-PCR experiments for selected 30 miRNAs in 47 tissue-specific samples and found agreement between the sequencing data and the q-PCR data. We envision that our report will serve as a valuable resource for future studies aimed at understanding miRNAome of pig Ten small RNA libraries from samples of porcine fetuses (30 days after insemination) to mixed tissues (180 days after birth) at ten developmental stages were sequenced.
2010-02-27 | E-GEOD-17885 | biostudies-arrayexpress
Project description:MicroRNAs or miRNAs are endogenously encoded small RNAs that play a key role in diverse plant biological processes. Jatropha curcas L. has received much attention as a potential oilseed crop for the production of renewable oil. Here, a sRNA library from mature seeds and three mRNA libraries, from three different stages of seed development, were generated by deep sequencing in order to identify and characterize miRNAs and pre-miRNAs of J. curcas. Computation analysis allowed the identification of 180 conserved miRNAs and 41 precursors (pre-miRNAs), as well as 16 novel pre-miRNAs. The predicted miRNA target genes are involved in a broad range of physiological functions, including cellular structure, nuclear function, translation, transport, hormone synthesis, defense and also lipid metabolism. Some of pre-miRNA and miRNA targets have different abundance among the three stages of seed development. A search for sequences that produce siRNA was performed indicating that J. curcas siRNAs play a role in nuclear functions, transport, catalytic process and diseases resistance. This study presents the first large scale identification of J. curcas miRNAs and their targets in mature seeds based on deep sequencing and contributes to understand the function of these miRNAs.
2016-09-06 | GSE50669 | GEO
Project description:A key step in microRNA (miRNA) biogenesis is the processing of a primary precursor RNA by the microprocessor into a precursor miRNA (pre-miRNA) intermediate. In plants, little is known about the processes that act on pre-miRNAs to influence miRNA biogenesis. Here, we performed 3’ RACE-seq to profile pre-miRNA 3’ ends in Arabidopsis. 3’ end heterogeneity was prevalent and the three microprocessor components promoted 3’ end precision. Extensive cytidylation and uridylation of precise and imprecise pre-miRNA 3’ ends were uncovered. The nucleotidyl transferase HESO1 uridylated pre-miRNAs in vitro and was responsible for most pre-miRNA uridylation in vivo. HESO1, NTP6 and NTP7 contribute to pre-miRNA cytidylation. Tailing of pre-miRNAs tended to restore trimmed pre-miRNAs to intact lengths to promote further processing. In addition, HESO1-mediated uridylation led to the degradation of some imprecisely processed pre-miRNAs. Thus, we uncovered widespread cytidylation and uridylation of pre-miRNAs and demonstrated diverse functions of pre-miRNA tailing in plants.
2019-10-22 | GSE134124 | GEO
Project description:MicroRNA precursors (pre-miRNAs) are short hairpin RNAs that are rapidly processed into mature microRNAs (miRNAs) in the cytoplasm. Due to their low abundance in cells, sequencing-based studies of pre-miRNAs have been limited. We successfully enriched for and deep sequenced pre-miRNAs in human cells by capturing these RNAs during their interaction with Argonaute (AGO) proteins. Using this approach, we detected > 350 pre-miRNAs in human cells and > 250 pre-miRNAs in a reanalysis of a similar study in mouse cells. We uncovered widespread trimming and non-templated additions to 3â?? ends of pre-miRNAs and mature miRNAs. Additionally, we identified novel AGO2-cleaved pre-miRNAs and created an index for microRNA precursor processing efficiency. This analysis revealed a subset of pre-miRNAs that produce low levels of mature miRNAs despite abundant precursors, including an annotated miRNA in the 5â?? UTR of the DiGeorge syndrome critical region 8 (Dgcr8) mRNA transcript. This led us to search for other AGO-associated stem-loops originating from other mRNA species, which identified hundreds of putative pre-miRNAs derived from mRNA sequences in both the mouse and human transcriptomes. Intriguingly, we found that iron responsive elements in ferritin heavy and light chain mRNAs are processed into AGO-associated stem-loops in both mouse and humans but do not produce functional small RNAs. In summary, we provide a wealth of information on mammalian pre-miRNAs, and identify novel microRNA and microRNA-like elements localized in mRNAs. pre-miRNA-seq and miRNA-seq from HEK293T cells. Reanalysis of HEK293T smRNA-seq, MEF pre-miRNA and MEF smRNA-seq (processed data for these Samples are linked below). The reanalyzed HEK293T Samples are from Series GSE66224 and consists of Samples GSM1617437 and GSM1617438. The reanalyzed MEF pre-miRNA and MEF smRNA-seq Samples were downloaded from European Nucleotide Archive (http://www.ebi.ac.uk/ena/) under the accession number PRJEB6756 (ERX525474 and ERX525475). RNA-seq analysis of HEK293T cells following knockdown of DGCR8, DROSHA or Luciferase.
2016-07-30 | E-GEOD-71710 | biostudies-arrayexpress
Project description:DEAD-box RNA-binding proteins (RBPs) play a significant role in RNA metabolism to achieve cellular homeostasis, including miRNA biogenesis and transcription. Hypoxia induces stemness cell-like characteristics in cancer cells and promotes malignant progression. Despite the fact that hypoxia can induce the changes in protein and RNA modification, thereby regulating downstream gene expressions, how modifications at different molecular layers interplay with each other are poorly understood. Here we show that hypoxia induces HectH9-mediated K63-linked polyubiquitination of the DEAD-box protein DDX17 as well as reduces N6-methyladenosine (m6A) marks in pri-miRNAs. While m6A potentiates DDX17 binding to pri-miRNAs, decreased m6A modifications of pri-miRNAs and increased polyubiquitination of DDX17 under hypoxia lead to decreased DDX17 binding to pri-miRNAs binding. These events enhance the association of DDX17 with the ubiquitin receptor p300 and lead to a decrease in miRNA biogenesis, especially for miRNAs regulating stemness and stemness-related genes. In addition, polyubiquitinated DDX17 together with p300 upregulates H3K56Ac levels on the stemness and stemness-related genes, resulting in enhancement of tumor initiating ability. Post-transcriptionally, decreased miRNA production, including those targeting stemness genes or stemness-related genes, also facilitates tumor initiation. Together, hypoxia triggers DDX17 poly-ubiquitination, which orchestrates dual mechanisms to increase tumor initiating ability and promote tumor progression.
2018-03-09 | PXD006059 | JPOST Repository