Conserved seed pairing, often flanked by adenosines, indicates that thousands of human genes are microRNA targets

We predict regulatory targets of vertebrate microRNAs (miRNAs) by identifying mRNAs with conserved complementarity to the seed (nucleotides 2-7) of the miRNA. An overrepresentation of conserved adenosines flanking the seed complementary sites in mRNAs indicates that primary sequence determinants can supplement base pairing to specify miRNA target recognition. In a four-genome analysis of 3' UTRs, approximately 13,000 regulatory relationships were detected above the estimate of false-positive predictions, thereby implicating as miRNA targets more than 5300 human genes, which represented 30% of our gene set. Targeting was also detected in open reading frames. In sum, well over one third of human genes appear to be conserved miRNA targets.     

Isolation and identification of miRNAs in Jatropha curcas

MicroRNAs (miRNAs) are small noncoding RNAs that play crucial regulatory roles by targeting mRNAs for silencing. To identify miRNAs in Jatropha curcas L, a bioenergy crop, cDNA clones from two small RNA libraries of leaves and seeds were sequenced and analyzed using bioinformatic tools. Fifty-two putative miRNAs were found from the two libraries, among them six were identical to known miRNAs and 46 were novel. Differential expression patterns of 15 miRNAs in root, stem, leave, fruit and seed were detected using quantitative real-time PCR. Ten miRNAs were highly expressed in fruit or seed, implying that they may be involved in seed development or fatty acids synthesis in seed. Moreover, 28 targets of the isolated miRNAs were predicted from a jatropha cDNA library database. The miRNA target genes were predicted to encode a broad range of proteins. Sixteen targets had clear BLASTX hits to the Uniprot database and were associated with genes belonging to the three major gene ontology categories of biological process, cellular component, and molecular function. Four targets were identified for JcumiR004. By silencing JcumiR004 primary miRNA, expressions of the four target genes were up-regulated and oil composition were modulated significantly, indicating diverse functions of JcumiR004.     

Experimental identification of microRNA-140 targets by silencing and overexpressing miR-140

MicroRNAs (miRNAs) are short noncoding RNA molecules regulating the expression of mRNAs. Target identification of miRNAs is computationally difficult due to the relatively low homology between miRNAs and their targets. We present here an experimental approach to target identification where the cartilage-specific miR-140 was overexpressed and silenced in cells it is normally expressed in separate experiments. Expression of mRNAs was profiled in both experiments and the intersection of mRNAs repressed by miR-140 overexpression and derepressed by silencing of miR-140 was identified. The intersection contained only 49 genes, although both treatments affected the accumulation of hundreds of mRNAs. These 49 genes showed a very strong enrichment for the miR-140 seed sequence implying that the approach is efficient and specific. Twenty-one of these 49 genes were predicted to be direct targets based on the presence of the seed sequence. Interestingly, none of these were predicted by the published target prediction methods we used. One of the potential target mRNAs, Cxcl12, was experimentally validated by Northern blot analysis and a luciferase reporter assay.     

A transcriptome‐wide study on the microRNA‐ and the Argonaute 1‐enriched small RNA‐mediated regulatory networks involved in plant leaf senescence

Leaf senescence is an important physiological process during the plant life cycle. However, systemic studies on the impact of microRNAs (miRNAs) on the expression of senescence‐associated genes (SAGs) are lacking. Besides, whether other Argonaute 1 (AGO1)‐enriched small RNAs (sRNAs) play regulatory roles in leaf senescence remains unclear. In this study, a total of 5,123 and 1,399 AGO1‐enriched sRNAs, excluding miRNAs, were identified in Arabidopsis thaliana and rice (Oryza sativa), respectively. After retrieving SAGs from the Leaf Senescence Database, all of the AGO1‐enriched sRNAs and the miRBase‐registered miRNAs of these two plants were included for target identification. Supported by degradome signatures, 200 regulatory pairs involving 120 AGO1‐enriched sRNAs and 40 SAGs, and 266 regulatory pairs involving 64 miRNAs and 42 SAGs were discovered in Arabidopsis. Moreover, 13 genes predicted to interact with some of the above‐identified target genes at protein level were validated as regulated by 17 AGO1‐enriched sRNAs and ten miRNAs in Arabidopsis. In rice, only one SAG was targeted by three AGO1‐enriched sRNAs, and one SAG was targeted by miR395. However, five AGO1‐enriched sRNAs were conserved between Arabidopsis and rice. Target genes conserved between the two plants were identified for three of the above five sRNAs, pointing to the conserved roles of these regulatory pairs in leaf senescence or other developmental procedures. Novel targets were discovered for three of the five AGO1‐enriched sRNAs in rice, indicating species‐specific functions of these sRNA–target pairs. These results could advance our understanding of the sRNA‐involved molecular processes modulating leaf senescence.     

Macro effects of microRNAs in plants

MicroRNAs (miRNAs) are 20- to 22-nucleotide fragments that regulate expression of mRNAs that have complementary sequences. They are numerous and widespread among eukaryotes, being conserved throughout evolution. The few miRNAs that have been fully characterized were found in Caenorhabditis elegans and are required for development. Recently, a study of miRNAs isolated from Arabidopsis showed that here also developmental genes are putative regulatory targets. A role for miRNAs have in plant development is supported by the developmental phenotypes of mutations in the genes required for miRNA processing.     

急性髓系白血病mRNA与非编码RNA差异表达谱及CeRNA调控网络分析

利用GEO数据库(gene expression omnibus database)通过生物信息学分析方法探讨急性髓系白血病(acute myelogenous leukemia,AML)的发病机制。检索GEO数据库中AML相关芯片数据集GSE142698、GSE142699和GSE96535。利用GEO2R分析得到差异mRNAs、miRNAs以及差异lncRNAs。利用在线生物信息学分析工具DAVID对差异mRNAs进行GO富集分析和KEGG通路分析。利用miRWalk数据库预测AML相关miRNAs的靶向mRNAs,利用Spongescan数据库预测AML相关miRNAs的靶向lncRNAs,构建lncRNA-miRNA-mRNA竞争性内源RNA （competing endogenous RNA,ceRNA）调控网络。共筛选出29个显著差异mRNAs、70个显著差异miRNAs和20 005个显著差异lncRNAs。GO富集分析和KEGG通路分析显示,差异表达基因主要涉及蛋白磷酸化、细胞分裂、细胞增殖的负调控、基因表达的正向调节、周期蛋白依赖的丝氨酸/苏氨酸激酶活性的调节等生物过程以及细胞周期、细胞衰老、癌症通路、PI3K-Akt通路等信号通路。将miRWalk数据库预测的靶向mRNAs与差异mRNAs取交集,Spongescan数据库预测的靶向lncRNAs与差异lncRNAs取交集,分别确定了25个mRNAs、6个lncRNAs参与AML相关ceRNA调控网络的构建。结果表明,lncRNAs可能作为关键的ceRNA,通过调控miRNA和相关靶基因参与AML的发生与发展,研究结果为AML诊断和治疗的分子生物学研究提供了新的依据。