A Genome-Wide Map of Conserved MicroRNA Targets in C. elegans

Center for Comparative Functional Genomics, Department of Biology, New York University, New York, New York 10003, USA.
Current Biology (Impact Factor: 9.57). 04/2006; 16(5):460-71. DOI: 10.1016/j.cub.2006.01.050
Source: PubMed


Metazoan miRNAs regulate protein-coding genes by binding the 3' UTR of cognate mRNAs. Identifying targets for the 115 known C. elegans miRNAs is essential for understanding their function.
By using a new version of PicTar and sequence alignments of three nematodes, we predict that miRNAs regulate at least 10% of C. elegans genes through conserved interactions. We have developed a new experimental pipeline to assay 3' UTR-mediated posttranscriptional gene regulation via an endogenous reporter expression system amenable to high-throughput cloning, demonstrating the utility of this system using one of the most intensely studied miRNAs, let-7. Our expression analyses uncover several new potential let-7 targets and suggest a new let-7 activity in head muscle and neurons. To explore genome-wide trends in miRNA function, we analyzed functional categories of predicted target genes, finding that one-third of C. elegans miRNAs target gene sets are enriched for specific functional annotations. We have also integrated miRNA target predictions with other functional genomic data from C. elegans.
At least 10% of C. elegans genes are predicted miRNA targets, and a number of nematode miRNAs seem to regulate biological processes by targeting functionally related genes. We have also developed and successfully utilized an in vivo system for testing miRNA target predictions in likely endogenous expression domains. The thousands of genome-wide miRNA target predictions for nematodes, humans, and flies are available from the PicTar website and are linked to an accessible graphical network-browsing tool allowing exploration of miRNA target predictions in the context of various functional genomic data resources.

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    • "While there is a growing number of online databases available for TG prediction (TargetScan (Lewis et al. 2003), miRanda (John et al. 2004), miRBase (Griffiths-Jones et al. 2008), Pic- Tar (Lall et al. 2006), PITA (Kertesz et al. 2007), DIANA-microT (Miranda et al. 2006), GeneMir (Huang et al. 2007), miRDB (Wang et al. 2008), mirDIP (Shirdel et al. 2011), only those utilized in this study) there are few validated miRNA TG databases (miRecords (Xiao et al. 2009), mirWalk (Dweep et al. 2011), miRTarBase (Vergoulis et al. 2012)). Many of these online databases store miRNA predicted TG's from multiple different prediction algorithms, and provide thousands of potential targets. "
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    ABSTRACT: Disruption in homeostatic levels of gene expression can lead directly to disease phenotype. miRNAs have key regulatory roles in modulating gene expression and have been shown to act as oncogenes, with their altered expression disrupting homeostatic biological mechanisms and adding to a disease phenotype. Using the Illumina MiSeq and HiSeq RNA sequencing platform data from the TCGA online resource, the objectives of the current research were to 1 Assess and quantify the existing online resource for miRNA target gene (TG) association, and 2 Define TG lists that can be used for genome-wide miRNA-mRNA-disease association analyses. Using the integration of miRNA lists from the Illumina platform and validated TG online databases, the researchers identified 307 miRNAs mapping to 3,358 validated TG’s, with 9,858 miRNA-TG connections. From eight online predicted TG databases, they find 547 miRNAs that map to 18,271 unique TG’s, with nearly three and a half million connections. Using the genomic location of miRNA and mRNAs assessed on the Illumina platforms, they identified 434 genes where miRNAs are co-located, and suggest that hypo/hyper methylation of these sites may play a key role in aberrant miRNA expression. In conclusion, using the Illumina miRNA and mRNA sequencing platforms, the researchers have created informative databases for the analyses of the complex interactions between miRNA and their target genes. The researchers’ approaches can be applied to similar data sets for any other disease.
    International Journal of Human Genetics 04/2014; 14(1):17-22. · 0.37 Impact Factor
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    • "Differentially expressed miRNAs (p<0.05) were selected for further study after bioinformatic analysis of target prediction data bases and thorough analysis of bibliography, to select potential candidates that could be involved in MRCC-related pathways and response to sunitinib. The following databases were used for target prediction: TargetScan release 5·1 [12], PicTac [13], PITA release 6 [14], miranda release sept2008 [15], and microcosm released 5 [16]. "
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    ABSTRACT: To identify tissue microRNAs predictive of sunitinib activity in patients with metastatic renal-cell-carcinoma (MRCC) and to evaluate in vitro their mechanism of action in sunitinib resistance. We screened 673 microRNAs using TaqMan Low-density-Arrays (TLDAs) in tumors from MRCC patients with extreme phenotypes of marked efficacy and resistance to sunitinib, selected from an identification cohort (n = 41). The most relevant differentially expressed microRNAs were selected using bioinformatics-based target prediction analysis and quantified by qRT-PCR in tumors from patients presenting similar phenotypes selected from an independent cohort (n = 101). In vitro experiments were conducted to study the role of miR-942 in sunitinib resistance. TLDAs identified 64 microRNAs differentially expressed in the identification cohort. Seven candidates were quantified by qRT-PCR in the independent series. MiR-942 was the most accurate predictor of sunitinib efficacy (p = 0.0074). High expression of miR-942, miR-628-5p, miR-133a, and miR-484 was significantly associated with decreased time to progression and overall survival. These microRNAs were also overexpressed in the sunitinib resistant cell line Caki-2 in comparison with the sensitive cell line. MiR-942 overexpression in Caki-2 up-regulates MMP-9 and VEGF secretion which, in turn, promote HBMEC endothelial migration and sunitinib resistance. We identified differentially expressed microRNAs in MRCC patients presenting marked sensitivity or resistance to sunitinib. MiR-942 was the best predictor of efficacy. We describe a novel paracrine mechanism through which high miR-942 levels in MRCC cells up-regulates MMP-9 and VEGF secretion to enhance endothelial migration and sunitinib resistance. Our results support further validation of these miRNA in clinical confirmatory studies.
    PLoS ONE 01/2014; 9(1):e86263. DOI:10.1371/journal.pone.0086263 · 3.23 Impact Factor
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    • "In order to better understand the potential downstream effects of the activity-dependent enhanced and depleted Ago2-associated miRNAs we integrated predictions from four of the most commonly used miRNA target prediction resources; DIANA (Maragkakis et al., 2011), miRanda (Griffiths-Jones et al., 2008), TargetScan (Friedman et al., 2009), and PicTar (Lall et al., 2006) using the RP method (Breitling et al., 2004). This allowed us to mitigate for the poor agreement normally found between "
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    ABSTRACT: microRNAs (miRNAs) are major regulators of protein synthesis in the brain. A major goal is to identify changes in miRNA expression underlying protein synthesis-dependent forms of synaptic plasticity such as long-term potentiation (LTP). Previous analyses focused on changes in miRNA levels in total lysate samples. Here, we asked whether changes in total miRNA accurately reflect changes in the amount of miRNA bound to Argonaute protein within the miRNA-induced silencing complex (miRISC). Ago2 immunoprecipitation was used to isolate RISC-associated miRNAs following high-frequency stimulation (HFS)-induced LTP in the dentate gyrus of anesthetized rats. Using locked-nucleic acid-based PCR cards for high-throughput screening and independent validation by quantitative TaqMan RT-PCR, we identified differential regulation of Ago2-associated and total miRNA expression. The ratio of Ago2/total miRNA expression was regulated bidirectionally in a miRNA-specific manner and was largely dependent on N-methyl-D-aspartate receptor (NMDA) activation during LTP induction. The present results identify miRNA association with Ago2 as a potential control point in activity-dependent synaptic plasticity in the adult brain. Finally, novel computational analysis for targets of the Ago2-associated miRNAs identifies 21 pathways that are enriched and differentially targeted by the miRNAs including axon guidance, mTOR, MAPK, Ras, and LTP.
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