Regulation by let-7 and lin-4 miRNAs results in target mRNA degradation

MicroRNAs (miRNAs) are approximately 22 nucleotide RNAs that negatively regulate the expression of protein-coding genes. In a present model of miRNA function in animals, miRNAs that form imperfect duplexes with their targets inhibit protein expression without affecting mRNA levels. Here, we report that in C. elegans, regulation by the let-7 miRNA results in degradation of its lin-41 target mRNA, despite the fact that its 3'UTR regulatory sequences can only partially base-pair with the miRNA. Furthermore, lin-14 and lin-28 are targets of the lin-4 miRNA, and we show that the mRNA levels for these protein-coding genes significantly decrease in response to lin-4 expression. This study reveals that mRNAs containing partial miRNA complementary sites can be targeted for degradation in vivo, raising the possibility that regulation at the level of mRNA stability may be more common than previously appreciated for the miRNA pathway.     

A whole-genome RNAi Screen for C. elegans miRNA pathway genes

BACKGROUND: miRNAs are an abundant class of small, endogenous regulatory RNAs. Although it is now appreciated that miRNAs are involved in a broad range of biological processes, relatively little is known about the actual mechanism by which miRNAs downregulate target gene expression. An exploration of which protein cofactors are necessary for a miRNA to downregulate a target gene should reveal more fully the molecular mechanisms by which miRNAs are processed, trafficked, and regulate their target genes. RESULTS: A weak allele of the C. elegans miRNA gene let-7 was used as a sensitized genetic background for a whole-genome RNAi screen to detect miRNA pathway genes, and 213 candidate miRNA pathway genes were identified. About 2/3 of the 61 candidates with the strongest phenotype were validated through genetic tests examining the dependence of the let-7 phenotype on target genes known to function in the let-7 pathway. Biochemical tests for let-7 miRNA production place the function of nearly all of these new miRNA pathway genes downstream of let-7 expression and processing. By monitoring the downregulation of the protein product of the lin-14 mRNA, which is the target of the lin-4 miRNA, we have identified 19 general miRNA pathway genes. CONCLUSIONS: The 213 candidate miRNA pathway genes identified could act at steps that produce and traffic miRNAs or in downstream steps that detect miRNA::mRNA duplexes to regulate mRNA translation. The 19 validated general miRNA pathway genes are good candidates for genes that may define protein cofactors for sorting or targeting miRNA::mRNA duplexes, or for recognizing the miRNA base-paired to the target mRNA to downregulate translation.     

CID-miRNA: a web server for prediction of novel miRNA precursors in human genome

microRNAs (miRNA) are a class of non-protein coding functional RNAs that are thought to regulate expression of target genes by direct interaction with mRNAs. miRNAs have been identified through both experimental and computational methods in a variety of eukaryotic organisms. Though these approaches have been partially successful, there is a need to develop more tools for detection of these RNAs as they are also thought to be present in abundance in many genomes. In this report we describe a tool and a web server, named CID-miRNA, for identification of miRNA precursors in a given DNA sequence, utilising secondary structure-based filtering systems and an algorithm based on stochastic context free grammar trained on human miRNAs. CID-miRNA analyses a given sequence using a web interface, for presence of putative miRNA precursors and the generated output lists all the potential regions that can form miRNA-like structures. It can also scan large genomic sequences for the presence of potential miRNA precursors in its stand-alone form. The web server can be accessed at http://mirna.jnu.ac.in/cidmirna/.     

Characteristics of the competition among RNAs for the binding of shared miRNAs

Competing endogenous RNAs (ceRNAs) are RNAs that share common miRNA binding sites and compete with each other for the miRNA association at these sites. The observation of this phenomenon in the cells altered the view of the miRNA target RNAs from molecules that are passively controlled by miRNAs to molecules that also modulate the miRNAs activity. In this review, we build a general profile of ceRNAS characteristics in order to facilitate ceRNAs identification by researchers. The information summarized here contains an actualized list of previously reported ceRNAs and classes of RNAs that can participate in this type of interaction, the expression behavior and characteristics of ceRNAs and miRNAs in the context of competition, the influence of the shared MREs/miRNAs numbers and the miRNA binding strength on the competition, reports on competition between RNAs in different subcellular localizations and the concept that ceRNAs may form a huge regulatory network in the cell.     

miRNAs的表达调控机制

microRNAs(miRNAs)是一类在转录后基因调控中发挥功能的非编码小RNAs,在发育、生长和分化等过程中发挥重要作用.至今已经在动物、植物和微生物等不同生物体中鉴定出来数千种miRNAs. miRNAs可以通过降解mRNA或抑制蛋白翻译的方式调节特异基因表达.生物体内约30%的基因都受miRNAs的调节.miRNAs的表达与功能受到转录因子、表观遗传学、多核苷酸多态性及其RNA编辑等多种因素的调节.此外,特异miRNA基因敲除的成功为研究miRNAs功能提供了有力的实验模型.     

Anti-Argonaute RIP-Chip shows that miRNA transfections alter global patterns of mRNA recruitment to microribonucleoprotein complexes

MicroRNAs (miRNAs) play key roles in gene expression regulation by guiding Argonaute (AGO)-containing microribonucleoprotein (miRNP) effector complexes to target polynucleotides. There are still uncertainties about how miRNAs interact with mRNAs. Here we employed a biochemical approach to isolate AGO-containing miRNPs from human H4 tumor cells by co-immunoprecipitation (co-IP) with a previously described anti-AGO antibody. Co-immunoprecipitated (co-IPed) RNAs were subjected to downstream Affymetrix Human Gene 1.0 ST microarray analysis. During rigorous validation, the “RIP-Chip” assay identified target mRNAs specifically associated with AGO complexes. RIP-Chip was performed after transfecting brain-enriched miRNAs (miR-107, miR-124, miR-128, and miR-320) and nonphysiologic control miRNA to identify miRNA targets. As expected, the miRNA transfections altered the mRNA content of the miRNPs. Specific mRNA species recruited to miRNPs after miRNA transfections were moderately in agreement with computational target predictions. In addition to recruiting mRNA targets into miRNPs, miR-107 and to a lesser extent miR-128, but not miR-124 or miR-320, caused apparent exclusion of some mRNAs that are normally associated with miRNPs. MiR-107 and miR-128 transfections also result in decreased AGO mRNA and protein levels. However, AGO mRNAs were not recruited to miRNPs after either miR-107 or miR-128 transfection, confirming that miRNAs may alter gene expression without stable association between particular mRNAs and miRNPs. In summary, RIP-Chip assays constitute an optimized, validated, direct, and high-throughput biochemical assay that provides data about specific miRNA:mRNA interactions, as well as global patterns of regulation by miRNAs.     

Individual mRNA expression profiles reveal the effects of specific microRNAs

Background  

MicroRNAs (miRNAs) are oligoribonucleotides with an important role in regulation of gene expression at the level of translation. Despite imperfect target complementarity, they can also significantly reduce mRNA levels. The validity of miRNA target gene predictions is difficult to assess at the protein level. We sought, therefore, to determine whether a general lowering of predicted target gene mRNA expression by endogenous miRNAs was detectable within microarray gene expression profiles.     

miRNA对水稻生长发育的调控

MicroRNA(miRNA)是一类小的非编码RNA,它们主要在转录后水平对靶mRNA进行切割或抑制mRNA的翻译来调控基因的表达. miRNA通过对靶基因的调控参与植物的生长发育、胁迫应答和代谢过程.在水稻中,已经发现并初步验证了许多与生长发育相关的miRNA,它们对水稻不同器官和形态发育发挥着重要作用.本文综述了水稻miRNA的发生和调控机制、靶基因的预测,重点介绍了miRNA对水稻生长发育和形态建成的研究进展,并对研究过程中存在的问题进行了讨论.为更好地了解miRNA及其靶基因在提高水稻产量和品质方面的作用,进一步解析miRNA在水稻中的调控机制提供参考.