Divergent target recognition by coexpressed 5′-isomiRs of miR-142-3p and selective viral mimicry

Sequence heterogeneity at the ends of mature microRNAs (miRNAs) is well documented, but its effects on miRNA function are largely unexplored. Here we studied the impact of miRNA 5′-heterogeneity, which affects the seed region critical for target recognition. Using the example of miR-142-3p, an emerging regulator of the hematopoietic lineage in vertebrates, we show that naturally coexpressed 5′-variants (5′-isomiRs) can recognize largely distinct sets of binding sites. Despite this, both miR-142-3p isomiRs regulate exclusive and shared targets involved in actin dynamics. Thus, 5′-heterogeneity can substantially broaden and enhance regulation of one pathway. Other 5′-isomiRs, in contrast, recognize largely overlapping sets of binding sites. This is exemplified by two herpesviral 5′-isomiRs that selectively mimic one of the miR-142-3p 5′-isomiRs. We hypothesize that other cellular and viral 5′-isomiRs can similarly be grouped into those with divergent or convergent target repertoires, based on 5′-sequence features. Taken together, our results provide a detailed characterization of target recognition by miR-142-3p and its 5′-isomiR-specific viral mimic. We furthermore demonstrate that miRNA 5′-end variation leads to differential targeting and can thus broaden the target range of miRNAs.     

The dynamic recruitment of TRBP to neuronal membranes mediates dendritogenesis during development

MicroRNAs are important regulators of local protein synthesis during neuronal development. We investigated the dynamic regulation of microRNA production and found that the majority of the microRNA‐generating complex, consisting of Dicer, TRBP, and PACT, specifically associates with intracellular membranes in developing neurons. Stimulation with brain‐derived neurotrophic factor (BDNF), which promotes dendritogenesis, caused the redistribution of TRBP from the endoplasmic reticulum into the cytoplasm, and its dissociation from Dicer, in a Ca²⁺‐dependent manner. As a result, the processing of a subset of neuronal precursor microRNAs, among them the dendritically localized pre‐miR16, was impaired. Decreased production of miR‐16‐5p, which targeted the BDNF mRNA itself, was rescued by expression of a membrane‐targeted TRBP. Moreover, miR‐16‐5p or membrane‐targeted TRBP expression blocked BDNF‐induced dendritogenesis, demonstrating the importance of neuronal TRBP dynamics for activity‐dependent neuronal development. We propose that neurons employ specialized mechanisms to modulate local gene expression in dendrites, via the dynamic regulation of microRNA biogenesis factors at intracellular membranes of the endoplasmic reticulum, which in turn is crucial for neuronal dendrite complexity and therefore neuronal circuit formation and function.     

microRNA序列和表达模式的多样性

microRNA(miRNA)在后生动植物细胞分化、生长发育、正常生理功能的维持、疾病的发生发展和转归中发挥重要功能.通常认为,已知miRNA对应的序列就是存储于miRBase数据库中相应的成熟miRNA序列.然而,人们忽视了miRNA序列长度和碱基的可变性,低估了来源于同一miRNA基因、非主要表达的miRNA的多样性及其功能的重要性.由于测序技术的快速发展和广泛使用,人们利用二代测序技术数据发现了miRNA序列和miRNA表达水平的多样性.基于miRNA序列和表达的多样性,本文介绍了miRNA序列5′端、3′端和内部的变异体、miRNA变异体的时空差异性表达和miRNA优势表达臂转换等方面近年来的研究进展.     

Identification of novel microRNA-like-coding sites on the long-stem microRNA precursors in Arabidopsis

Plant microRNA (miRNA) is a crucial regulator of gene expression. It has been reported that more than one miRNA/miRNA* duplex could be produced from a microRNA precursor (pre-miRNA). In this study, we performed a comprehensive search for the novel miRNA candidates on the pre-miRNAs of Arabidopsis. AGO1 enrichment, co-existence of the miRNA*-like coordinates, and unique genome-wide match sites were taken into consideration for candidate screening. As a result, 43 miRNA-like candidates derived from 25 pre-miRNAs were identified. Among these candidates, 31 strong candidates from 22 pre-miRNAs passed all the filtering steps. Interestingly, some of these miRNA-like candidates showed organ-specific expression patterns. After target prediction and degradome sequencing data-based validation, five miRNA candidate–target pairs (ath-miR863-5p.2–AT1G76550.1, ath-miR822.2–AT5G03552.1, ath-miR822.3–AT5G02350.1, sRNA4–AT1G66290.1 and sRNA6–AT1G66310.1) were identified, providing a basis for in-depth functional analysis of these miRNA candidates. These results could update the current understanding of the biogenesis and the action of the plant miRNAs.     

Identification of novel microRNAs in rice (Oryza sativa) based on the cleavage signals in precursors

MicroRNAs (miRNAs) are crucial regulators of gene expression in plants and a growing number of novel miRNA genes have been cloned in rice in recent years. However, there is no evidence that all miRNAs have been discovered, especially for those low expression ones which are difficult to be found by conventional methods. By taking advantage of the finding that DCL1-mediated cleavage signals for the processing of the miRNA precursors could be used as the clues for novel miRNAs’ discovery, a genome-wide search for rice miRNA candidates was carried out. As a result, 51 previously validated miRNAs and 24 novel miRNA candidates were discovered. After target prediction and degradome sequencing data-based validation, coupled with reverse approach retest, 10 miRNA candidate–mRNA target pairs were further identified, providing a basis for in-depth functional analysis of these miRNA candidates. Besides, some isomiRs found in this study showed more likely to be the real miRNAs. We also found an exceptional example which did not obey the rule that 22-nt miRNAs have the ability to trigger the phased siRNAs production from the cleaved targets.     

Deep-sequencing of endothelial cells exposed to hypoxia reveals the complexity of known and novel microRNAs

In order to understand the role of microRNAs (miRNAs) in vascular physiopathology, we took advantage of deep-sequencing techniques to accurately and comprehensively profile the entire miRNA population expressed by endothelial cells exposed to hypoxia. SOLiD sequencing of small RNAs derived from human umbilical vein endothelial cells (HUVECs) exposed to 1% O₂ or normoxia for 24 h yielded more than 22 million reads per library. A customized bioinformatic pipeline identified more than 400 annotated microRNA/microRNA* species with a broad abundance range: miR-21 and miR-126 totaled almost 40% of all miRNAs. A complex repertoire of isomiRs was found, displaying also 5′ variations, potentially affecting target recognition. High-stringency bioinformatic analysis identified microRNA candidates, whose predicted pre-miRNAs folded into a stable hairpin. Validation of a subset by qPCR identified 18 high-confidence novel miRNAs as detectable in independent HUVEC cultures and associated to the RISC complex. The expression of two novel miRNAs was significantly down-modulated by hypoxia, while miR-210 was significantly induced. Gene ontology analysis of their predicted targets revealed a significant association to hypoxia-inducible factor signaling, cardiovascular diseases, and cancer. Overexpression of the novel miRNAs in hypoxic endothelial cells affected cell growth and confirmed the biological relevance of their down-modulation. In conclusion, deep-sequencing accurately profiled known, variant, and novel microRNAs expressed by endothelial cells in normoxia and hypoxia.     

miRNA的异构体——isomiR

最近,深度测序技术揭示:同一个miRNA前体可能由于Drosha或Dicer的剪切位点改变,外切核酸酶介导的miRNA末端缩短,miRNA编辑或miRNA 3'末端无需模板的核苷酸添加等4种原因,而形成多种长度或序列不同的miRNAs异构体—isomiR.因为这些isomiR与已注解的miRNA可以调节同一个靶标,也可以靶向不同的靶标,所以它们不仅扩大了miRNA调节的范围,而且还有可能代表了每种miRNA基于isomiR的一种微型调节网络.研究发现,isomiR的表达具有细胞、组织、发育和疾病状况等特异性,并且很多人类疾病的致病机制也与它们有关,推测isomiR将来不仅有可能成为疾病诊断或治疗的生物学标记或靶标,而且相关的研究还对于RNA干扰技术也具有重要的指导意义.本文主要综述了isomiR的研究进展,并对isomiR应用前景做了展望.     

Structural basis of microRNA biogenesis by Dicer-1 and its partner protein Loqs-PB

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