Cloning and characterization of miRNAs from maize seedling roots under low phosphorus stress

MicroRNAs (miRNAs) are a class of small, non-coding regulatory RNAs that regulate gene expression by guiding target mRNA cleavage or translational inhibition in plants and animals. In this study, a small RNA library was constructed to identify conserved miRNAs as well as novel miRNAs in maize seedling roots under low level phosphorus stress. Twelve miRNAs were identified by high throughput sequencing of the library and subsequent analysis, two belong to conserved miRNA families (miRNA399b and miRNA156), and the remaining ten are novel and one of latter is conserved in gramineous species. Based on sequence homology, we predicted 125 potential target genes of these miRNAs and then expression patterns of 7 miRNAs were validated by semi-RT-PCR analysis. MiRNA399b, Zma-miR3, and their target genes (Zmpt1 and Zmpt2) were analyzed by real-time PCR. It is shown that both miRNA399b and Zma-miR3 are induced by low phosphorus stress and regulated by their target genes (Zmpt1 and Zmpt2). Moreover, Zma-miR3, regulated by two maize inorganic phosphate transporters as a newly identified miRNAs, would likely be directly involved in phosphate homeostasis, so was miRNA399b in Arabidopsis and rice. These results indicate that both conserved and maize-specific miRNAs play important roles in stress responses and other physiological processes correlated with phosphate starvation, regulated by their target genes. Identification of these differentially expressed miRNAs will facilitate us to uncover the molecular mechanisms underlying the progression of maize seedling roots development under low level phosphorus stress.     

miSSING LINKS: miRNAs and plant development

The discovery of hundreds of plant micro RNAs (miRNAs) has triggered much speculation about their potential roles in plant development. The search for plant genes involved in miRNA processing has revealed common factors such as DICER, and new molecules, including HEN1. Progress is also being made toward identifying miRNA target genes and understanding the mechanisms of miRNA-mediated gene regulation in plants. This work has lead to a reexamination of many previously characterized mutations that are now known to affect components or targets of miRNA-mediated pathways.     

Identification of conserved microRNAs and their target genes in tomato (Lycopersicon esculentum)

MicroRNAs (miRNAs) are a class of non-coding RNAs that have important gene regulation roles in various organisms. To date, a total of 1279 plant miRNAs have been deposited in the miRNA miRBase database (Release 10.1). Many of them are conserved during the evolution of land plants suggesting that the well-conserved miRNAs may also retain homologous target interactions. Recently, little is known about the experimental or computational identification of conserved miRNAs and their target genes in tomato. Here, using a computational homology search approach, 21 conserved miRNAs were detected in the Expressed Sequence Tags (EST) and Genomic Survey Sequence (GSS) databases. Following this, 57 potential target genes were predicted by searching the mRNA database. Most of the target mRNAs appeared to be involved in plant growth and development. Our findings verified that the well-conserved tomato miRNAs have retained homologous target interactions amongst divergent plant species. Some miRNAs express diverse combinations in different cell types and have been shown to regulate cell-specific target genes coordinately. We believe that the targeting propensity for genes in different biological processes can be explained largely by their protein connectivity.     

植物miRNA参与调控作物农艺性状的研究进展

植物microRNA (miRNA)是一类长度约为20～24 nt的内源非编码小RNA,它们通过在转录后水平调控靶基因的表达,在植物的生长发育、逆境响应和环境适应等过程中起着关键作用. miRNA对水稻、玉米、大豆等重要经济作物的农艺性状也起着重要的调控作用,在改良农作物性状上具有重大的应用潜能.本文重点介绍了参与作物农艺性状(如株型、花期、种子发育及抗逆等)调控的关键miRNA及其调控途径,同时总结了miRNA参与作物性状改良的主要研究方法和手段,并讨论了miRNA在作物性状改良应用中的前景.     

MicroRNAs target gene and signaling pathway by bioinformatics analysis in the cardiac hypertrophy

Cardiac hypertrophy is a physiological adaptive response of the heart to diverse pathophysiological stimuli. Initially, it may be adaptive to normalize wall stress and to preserve contractile performance. This adaptive process may gradually progress to dilated cardiomyopathy, fibrotic diseases, arrhythmia, heart failure and even sudden death. Although various molecular pathways responsible for the coordinated control of the hypertrophic program, little is known about their underlying molecular mechanisms. Very recently, increasing evidence showed that miRNAs are key modulators of both cardiovascular development and function, which govern the process of cardiac hypertrophy and heart failure. MicroRNAs (miRNAs) act in a complex functional network in which each single miRNAs might control thousands of distinct target genes, and each single protein-coding gene can be regulated by many different miRNAs. Identifying the roles of miRNAs, their target genes and signaling pathways in cardiac hypertrophy by bioinformatic analysis will provide more insight into the molecular mechanisms underlying this disease process. Currently, bioinformatics resource such as GO and KEGG was applied to describe the miRNAs target genes function and identify the mRNA interaction networks that are responsible for various cellular processes. It provides a useful approach to observe the function of microRNA in physiological and pathological conditions. In this review, we will give a discussion on the dysregulation of specific miRNAs in cardiac hypertrophy and signaling pathways linking the hypertrophy-regulating miRNAs to the pathological process of cardiac hypertrophy. Finally, we place special emphasis on the essential role of bioinformatics analysis to predict the target genes and miRNAs gene networks.     

MicroRNA Profiling in Human Colon Cancer Cells during 5-Fluorouracil-Induced Autophagy

Autophagy modulation is now recognized as a potential therapeutic approach for cancer (including colorectal cancer), yet the molecular mechanisms regulating autophagy in response to cellular stress are still not well understood. MicroRNAs (miRNAs) have been found to play important roles in controlling many cellular functions, including growth, metabolism and stress response. The physiological importance of the miRNA-autophagy interconnection is only beginning to be elucidated. MiRNA microarray technology facilitates analysis of global miRNA expression in certain situations. In this study, we explored the expression profile of miRNAs during the response of human colon cancer cells (HT29s) to 5-FU treatment and nutrient starvation using miRNA microarray analysis. The alteration of miRNA expression showed the same pattern under both conditions was further testified by qRT-PCR in three human colon cancer cell lines. In addition, bioinformatic prediction of target genes, pathway analysis and gene network analysis were performed to better understand the roles of these miRNAs in the regulation of autophagy. We identified and selected four downregulated miRNAs including hsa-miR-302a-3p and 27 upregulated miRNAs under these two conditions as having the potential to target genes involved in the regulation of autophagy in human colon cancer cells. They have the potential to modulate autophagy in 5-FU-based chemotherapy in colorectal cancer.     

Identification of the Conserved and Novel miRNAs in Mulberry by High-Throughput Sequencing

miRNAs are a class of non-coding endogenous small RNAs. They play vital roles in plant growth, development, and response to biotic and abiotic stress by negatively regulating genes. Mulberry trees are economically important species with multiple uses. However, to date, little is known about mulberry miRNAs and their target genes. In the present study, three small mulberry RNA libraries were constructed and sequenced using high-throughput sequencing technology. Results showed 85 conserved miRNAs belonging to 31 miRNA families and 262 novel miRNAs at 371 loci. Quantitative real-time PCR (qRT-PCR) analysis confirmed the expression pattern of 9 conserved and 5 novel miRNAs in leaves, bark, and male flowers. A total of 332 potential target genes were predicted to be associated with these 113 novel miRNAs. These results provide a basis for further understanding of mulberry miRNAs and the biological processes in which they are involved.     

植物体内调控miRNA合成与功能的机制研究进展

microRNA(miRNA)能够在转录后水平通过剪切或抑制翻译对靶mRNA进行调节,并且广泛参与植物的生长发育、生物胁迫和非生物胁迫等过程,因此引起了众多研究者的关注。随着对miRNA研究的深入,人们发现miRNA从转录到转录后成熟再到行使功能的过程中,会受到很多因子的调控。这些因子可以是蛋白质、核酸序列、基因、甚至miRNA本身,由于这些因子的参与,使得miRNA调节的生物学过程更具复杂性和灵活性。本文从转录、加工、活性调节和反馈调控等层次综述了近年来调控植物miRNA合成与功能方面取得的进展,以期为miRNA调控机制的研究提供理论基础和新思路。     

植物激素相关microRNA研究进展

microRNA（miRNA）是22nt左右的非编码RNA,主要在转录后水平调节基因的活性。miRNA通过与靶基因的互补位点结合从而降解靶基因mRNA或抑制其翻译。近年的研究发现,miRNA在植物生长发育中发挥着重要的调控作用。目前已知一些miRNA参与植物激素信号途径的切入点,这为我们了解miRNA和植物激素在植物发育中的作用提供了新思路。本文综述了miRNA参与植物激素信号应答及生物合成的研究进展,并对一些miINA在植物激素信号应答中可能的作用进行了讨论。     

Computational identification of citrus microRNAs and target analysis in citrus expressed sequence tags

MicroRNAs (miRNAs) are a new family of small RNA molecules found in plants and animals. We developed a comprehensive strategy for identifying new miRNA homologues by mining the repository of available citrus expressed sequence tags (ESTs). By adopting a range of filtering criteria, we identified a total of 38 potential miRNAs--nine, five, nine and 15 miRNAs in Citrus trifoliata (ctr-miRNAs), C. clementina (ccl-miRNAs), C. reticulata (crt-miRNAs) and C. sinensis (csi-miRNAs), respectively--from more than 430,000 EST sequences in citrus. Using the potential miRNA sequences, we conducted a further BLAST search of the mRNA database and found six potential target genes in these citrus species. Eight miRNAs were selected in order to verify their existence in citrus using Northern blotting, and the functions of several miRNAs in miRNA-mediated gene regulation are experimentally suggested. It appears that all these miRNAs regulate expression of their target genes by cleavage, which is the most common situation in gene regulation mediated by plant miRNAs. Our achievement in identifying new miRNAs in citrus provides a powerful incentive for further studies on the important roles of these miRNAs.     

MicroRNA Expression Patterns of CD8+ T Cells in Acute and Chronic Brucellosis

Although our knowledge about Brucella virulence factors and the host response increase rapidly, the mechanisms of immune evasion by the pathogen and causes of chronic disease are still unknown. Here, we aimed to investigate the immunological factors which belong to CD8+ T cells and their roles in the transition of brucellosis from acute to chronic infection. Using miRNA microarray, more than 2000 miRNAs were screened in CD8+ T cells of patients with acute or chronic brucellosis and healthy controls that were sorted from peripheral blood with flow cytometry and validated through qRT-PCR. Findings were evaluated using GeneSpring GX (Agilent) 13.0 software and KEGG pathway analysis. Expression of two miRNAs were determined to display a significant fold change in chronic group when compared with acute or control groups. Both miRNAs (miR-126-5p and miR-4753-3p) were decreased (p <0.05 or fold change > 2). These miRNAs have the potential to be the regulators of CD8+ T cell-related marker genes for chronic brucellosis infections. The differentially expressed miRNAs and their predicted target genes are involved in MAPK signaling pathway, cytokine-cytokine receptor interactions, endocytosis, regulation of actin cytoskeleton, and focal adhesion indicating their potential roles in chronic brucellosis and its progression. It is the first study of miRNA expression analysis of human CD8+ T cells to clarify the mechanism of inveteracy in brucellosis.