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Sample records for micro rna cluster

  1. MicroRNA Clusters in the Adult Mouse Heart: Age-Associated Changes

    PubMed Central

    Zhang, Xiaomin; Azhar, Gohar; Williams, Emmanuel D.; Rogers, Steven C.; Wei, Jeanne Y.

    2015-01-01

    The microRNAs and microRNA clusters have been implicated in normal cardiac development and also disease, including cardiac hypertrophy, cardiomyopathy, heart failure, and arrhythmias. Since a microRNA cluster has from two to dozens of microRNAs, the expression of a microRNA cluster could have a substantial impact on its target genes. In the present study, the configuration and distribution of microRNA clusters in the mouse genome were examined at various inter-microRNA distances. Three important microRNA clusters that are significantly impacted during adult cardiac aging, the miR-17-92, miR-106a-363, and miR-106b-25, were also examined in terms of their genomic location, RNA transcript character, sequence homology, and their relationship with the corresponding microRNA families. Multiple microRNAs derived from the three clusters potentially target various protein components of the cdc42-SRF signaling pathway, which regulates cytoskeleton dynamics associated with cardiac structure and function. The data indicate that aging impacted the expression of both guide and passenger strands of the microRNA clusters; nutrient stress also affected the expression of the three microRNA clusters. The miR-17-92, miR-106a-363, and miR-106b-25 clusters are likely to impact the Cdc42-SRF signaling pathway and thereby affect cardiac morphology and function during pathological conditions and the aging process. PMID:26221604

  2. Helicobacter pylori interferes with an embryonic stem cell micro RNA cluster to block cell cycle progression

    PubMed Central

    2011-01-01

    Background MicroRNAs, post-transcriptional regulators of eukaryotic gene expression, are implicated in host defense against pathogens. Viruses and bacteria have evolved strategies that suppress microRNA functions, resulting in a sustainable infection. In this work we report that Helicobacter pylori, a human stomach-colonizing bacterium responsible for severe gastric inflammatory diseases and gastric cancers, downregulates an embryonic stem cell microRNA cluster in proliferating gastric epithelial cells to achieve cell cycle arrest. Results Using a deep sequencing approach in the AGS cell line, a widely used cell culture model to recapitulate early events of H. pylori infection of gastric mucosa, we reveal that hsa-miR-372 is the most abundant microRNA expressed in this cell line, where, together with hsa-miR-373, it promotes cell proliferation by silencing large tumor suppressor homolog 2 (LATS2) gene expression. Shortly after H. pylori infection, miR-372 and miR-373 synthesis is highly inhibited, leading to the post-transcriptional release of LATS2 expression and thus, to a cell cycle arrest at the G1/S transition. This downregulation of a specific cell-cycle-regulating microRNA is dependent on the translocation of the bacterial effector CagA into the host cells, a mechanism highly associated with the development of severe atrophic gastritis and intestinal-type gastric carcinoma. Conclusions These data constitute a novel example of host-pathogen interplay involving microRNAs, and unveil the couple LATS2/miR-372 and miR-373 as an unexpected mechanism in infection-induced cell cycle arrest in proliferating gastric cells, which may be relevant in inhibition of gastric epithelium renewal, a major host defense mechanism against bacterial infections. PMID:22027184

  3. Altered Spinal MicroRNA-146a and the MicroRNA-183 Cluster Contribute to Osteoarthritic Pain in Knee Joints

    PubMed Central

    Li, Xin; Kroin, Jeffrey S; Kc, Ranjan; Gibson, Gary; Chen, Di; Corbett, Grant T; Pahan, Kalipada; Fayyaz, Sana; Kim, Jae-Sung; van Wijnen, Andre J.; Suh, Joon; Kim, Su-Gwan; Im, Hee-Jeong

    2015-01-01

    Objective Examine whether altered expression of microRNAs in central nervous system components is pathologically linked to chronic knee joint pain in osteoarthritis. Methods A surgical animal model for knee joint OA was generated by medial meniscus transection in rats followed by behavioral pain tests. Relationships between pathological changes in knee joint and development of chronic joint pain were examined by histology and imaging analyses. Alterations in microRNAs associated with OA-evoked pain sensation were determined in bilateral lumbar dorsal root ganglia (DRG) and the spinal dorsal horn by microRNA array followed by individual microRNA analyses. Gain- and loss-of-function studies of selected microRNAs (miR-146a and miR-183 cluster) were conducted to identify target pain mediators regulated by these selective microRNAs in glial cells. Results The ipsilateral hind leg displayed significantly increased hyperalgesia after 4 weeks of surgery and sensitivity was sustained for the remainder of the 8 week experimental period (F=341, P<0.001). The development of OA-induced chronic pain was correlated with pathological changes in the knee joints as assessed by histological and imaging analyses. MicroRNA analyses showed that miR-146a and the miR-183 cluster were markedly reduced in the sensory neurons in DRG (L4/L5) and spinal cord from animals experiencing knee joint OA pain. The downregulation of miR-146a and/or the miR-183 cluster in the central compartments (DRG and spinal cord) are closely associated with the upregulation of inflammatory pain mediators. The corroboration between decreases in these signature microRNAs and their specific target pain mediators were further confirmed by gain- and loss-of-function analyses in glia, the major cellular component of the central nervous system (CNS). Conclusion MicroRNA therapy using miR-146a and the miR-183 cluster could be powerful therapeutic intervention for OA in alleviating joint pain and concomitantly regenerating

  4. The microRNA-183 cluster: the family that plays together stays together

    PubMed Central

    Dambal, Shweta; Shah, Mit; Mihelich, Brittany; Nonn, Larisa

    2015-01-01

    The microRNA (miR)183 cluster, which is comprised of miRs-183, -96 and -182, is also a miR family with sequence homology. Despite the strong similarity in the sequences of these miRs, minute differences in their seed sequences result in both overlapping and distinct messenger RNA targets, which are often within the same pathway. These miRs have tightly synchronized expression during development and are required for maturation of sensory organs. In comparison to their defined role in normal development, the miR-183 family is frequently highly expressed in a variety of non-sensory diseases, including cancer, neurological and auto-immune disorders. Here, we discuss the conservation of the miR-183 cluster and the functional role of this miR family in normal development and diseases. We also describe the regulation of vital cellular pathways by coordinated expression of these miR siblings. This comprehensive review sheds light on the likely reasons why the genomic organization and seeming redundancy of the miR-183 family cluster was conserved through 600 million years of evolution. PMID:26170234

  5. The microRNA-212/132 cluster regulates B cell development by targeting Sox4

    PubMed Central

    Mehta, Arnav; Mann, Mati; Zhao, Jimmy L.; Marinov, Georgi K.; Majumdar, Devdoot; Garcia-Flores, Yvette; Du, Xiaomi; Erikci, Erdem; Chowdhury, Kamal

    2015-01-01

    MicroRNAs have emerged as key regulators of B cell fate decisions and immune function. Deregulation of several microRNAs in B cells leads to the development of autoimmune disease and cancer in mice. We demonstrate that the microRNA-212/132 cluster (miR-212/132) is induced in B cells in response to B cell receptor signaling. Enforced expression of miR-132 results in a block in early B cell development at the prepro–B cell to pro–B cell transition and induces apoptosis in primary bone marrow B cells. Importantly, loss of miR-212/132 results in accelerated B cell recovery after antibody-mediated B cell depletion. We find that Sox4 is a target of miR-132 in B cells. Co-expression of SOX4 with miR-132 rescues the defect in B cell development from overexpression of miR-132 alone, thus suggesting that miR-132 may regulate B lymphopoiesis through Sox4. In addition, we show that the expression of miR-132 can inhibit cancer development in cells that are prone to B cell cancers, such as B cells expressing the c-Myc oncogene. We have thus uncovered miR-132 as a novel contributor to B cell development. PMID:26371188

  6. MicroRNA-Target Network Inference and Local Network Enrichment Analysis Identify Two microRNA Clusters with Distinct Functions in Head and Neck Squamous Cell Carcinoma

    PubMed Central

    Sass, Steffen; Pitea, Adriana; Unger, Kristian; Hess, Julia; Mueller, Nikola S.; Theis, Fabian J.

    2015-01-01

    MicroRNAs represent ~22 nt long endogenous small RNA molecules that have been experimentally shown to regulate gene expression post-transcriptionally. One main interest in miRNA research is the investigation of their functional roles, which can typically be accomplished by identification of mi-/mRNA interactions and functional annotation of target gene sets. We here present a novel method “miRlastic”, which infers miRNA-target interactions using transcriptomic data as well as prior knowledge and performs functional annotation of target genes by exploiting the local structure of the inferred network. For the network inference, we applied linear regression modeling with elastic net regularization on matched microRNA and messenger RNA expression profiling data to perform feature selection on prior knowledge from sequence-based target prediction resources. The novelty of miRlastic inference originates in predicting data-driven intra-transcriptome regulatory relationships through feature selection. With synthetic data, we showed that miRlastic outperformed commonly used methods and was suitable even for low sample sizes. To gain insight into the functional role of miRNAs and to determine joint functional properties of miRNA clusters, we introduced a local enrichment analysis procedure. The principle of this procedure lies in identifying regions of high functional similarity by evaluating the shortest paths between genes in the network. We can finally assign functional roles to the miRNAs by taking their regulatory relationships into account. We thoroughly evaluated miRlastic on a cohort of head and neck cancer (HNSCC) patients provided by The Cancer Genome Atlas. We inferred an mi-/mRNA regulatory network for human papilloma virus (HPV)-associated miRNAs in HNSCC. The resulting network best enriched for experimentally validated miRNA-target interaction, when compared to common methods. Finally, the local enrichment step identified two functional clusters of mi

  7. Micro RNA-17-92 cluster mediates interleukin-4-suppressed IL-10 expression in B cells

    PubMed Central

    Liu, Zhi-Qiang; Yang, Gui; Geng, Xiao-Rui; Liu, Jiang-Qi; Mo, Li-Hua; Liu, Zhi-Gang; Yang, Ping-Chang

    2016-01-01

    The pathogenesis of allergen-related inflammation in the intestine is to be further understood. Micro RNA (miR) can regulate immune responses. This study aims to investigate the role of miR-17-92 cluster in the induction of food allergen-related inflammation in the intestine. In this study, a mouse model of food allergen-related intestinal inflammation was developed. Expression of miR-17-92 cluster in B cells of the intestinal mucosa was analyzed by real time quantitative RT-PCR. The results showed that the levels of miR-19a, one of the members of the miR-17-92 cluster, were detected in the B cells of the intestine of mice sensitized to ovalbumin, which was significantly higher than that in naïve control mice. The expression of IL-10 by B cells was significantly lower in the sensitized mice as compared with naive control mice. Exposure to IL-4 in the culture increased the expression of miR-19a as well as suppression the expression of IL-10 in B cells via remolding DNA structure at the IL-10 promoter locus. We conclude that B cells from sensitized mice show higher levels of miR-19a, which plays an important role in the suppression of IL-10 in the B cells. PMID:27347339

  8. Evolution of an X-linked primate-specific micro RNA cluster.

    PubMed

    Li, Jingjing; Liu, Yu; Dong, Dong; Zhang, Zhaolei

    2010-03-01

    Micro RNAs (miRNAs) are a class of small regulatory RNAs, which posttranscriptionally repress protein production of the targeted messenger RNAs (mRNAs). Accumulating evidence has suggested lineage-specific miRNAs have contributed to lineage-specific characteristics. However, the birth and death of these miRNAs, particularly in primates, largely remain unexplored. We herein characterized the evolutionary history of a newly discovered miRNA cluster on primate X-chromosome, spanning a approximately 33-kb region in human Xq27.3. The cluster consists of six distinct miRNAs, four of which are compactly organized in a 3-kb region belonging to a phylogenetic group distinct from the other two miRNAs. By comparing the genomic structure of this cluster in human with four other primates (chimpanzee, orangutan, rhesus macaque, and marmoset), we identified several previously uncovered miRNAs in these primates that share orthology with the human miRNAs. We found the entire miRNA cluster was well conserved among primate species but unidentifiable in other mammalian species (including mouse, rat, cat, dog, horse, cow, opossum, and platypus), suggesting that the formation of this cluster was after the primate-rodent split but before the emergence of New-World Monkey (represented by marmoset). Our analysis further revealed complex evolutionary dynamics on this locus, characterized by extensive duplication events. Phylogenetic analysis revealed birth and death of the miRNAs within this region, accompanied by rapid evolution, which highlighted their functional importance. These miRNAs are primarily expressed in primate epididymis, part of the male reproductive system. Our analysis showed that their predicted target mRNAs are significantly enriched for several functional classes relevant to epididymal physiology, such as morphogenesis of epithelium and tube development. Furthermore, several genes controlling sperm maturation and male fertility are confidently predicted to be their

  9. The microRNA-132 and microRNA-212 cluster regulates hematopoietic stem cell maintenance and survival with age by buffering FOXO3 expression

    PubMed Central

    Mehta, Arnav; Zhao, Jimmy L.; Sinha, Nikita; Marinov, Georgi K.; Mann, Mati; Kowalczyk, Monika S.; Galimidi, Rachel P.; Du, Xiaomi; Erikci, Erdem; Regev, Aviv; Chowdhury, Kamal; Baltimore, David

    2015-01-01

    Summary MicroRNAs are critical post-transcriptional regulators of hematopoietic cell-fate decisions, though little remains known about their role in aging hematopoietic stem cells (HSCs). We found that the microRNA-212/132 cluster (Mirc19) is enriched in HSCs and is up-regulated during aging. Both over-expression and deletion of microRNAs in this cluster leads to inappropriate hematopoiesis with age. Enforced expression of miR-132 in the bone marrow of mice led to rapid HSC cycling and depletion. A genetic deletion of Mirc19 in mice resulted in HSCs that had altered cycling, function, and survival in response to growth factor starvation. We found that miR-132 exerted its effect on aging HSCs by targeting the transcription factor FOXO3, a known aging associated gene. Our data demonstrates that Mirc19 plays a role in maintaining balanced hematopoietic output by buffering FOXO3 expression. We have thus identified it as a potential target that may play a role in age-related hematopoietic defects. PMID:26084022

  10. MicroRNA 17-92 cluster mediates ETS1 and ETS2-dependent RAS-oncogenic transformation.

    PubMed

    Kabbout, Mohamed; Dakhlallah, Duaa; Sharma, Sudarshana; Bronisz, Agnieszka; Srinivasan, Ruchika; Piper, Melissa; Marsh, Clay B; Ostrowski, Michael C

    2014-01-01

    The ETS-family transcription factors Ets1 and Ets2 are evolutionarily conserved effectors of the RAS/ERK signaling pathway, but their function in Ras cellular transformation and biology remains unclear. Taking advantage of Ets1 and Ets2 mouse models to generate Ets1/Ets2 double knockout mouse embryonic fibroblasts, we demonstrate that deletion of both Ets1 and Ets2 was necessary to inhibit HrasG12V induced transformation both in vitro and in vivo. HrasG12V expression in mouse embryonic fibroblasts increased ETS1 and ETS2 expression and binding to cis-regulatory elements on the c-Myc proximal promoter, and consequently induced a robust increase in MYC expression. The expression of the oncogenic microRNA 17-92 cluster was increased in HrasG12V transformed cells, but was significantly reduced when ETS1 and ETS2 were absent. MYC and ETS1 or ETS2 collaborated to increase expression of the oncogenic microRNA 17-92 cluster in HrasG12V transformed cells. Enforced expression of exogenous MYC or microRNA 17-92 rescued HrasG12V transformation in Ets1/Ets2-null cells, revealing a direct function for MYC and microRNA 17-92 in ETS1/ETS2-dependent HrasG12V transformation. PMID:24968297

  11. A tripartite clustering analysis on microRNA, gene and disease model.

    PubMed

    Shen, Chengcheng; Liu, Ying

    2012-02-01

    Alteration of gene expression in response to regulatory molecules or mutations could lead to different diseases. MicroRNAs (miRNAs) have been discovered to be involved in regulation of gene expression and a wide variety of diseases. In a tripartite biological network of human miRNAs, their predicted target genes and the diseases caused by altered expressions of these genes, valuable knowledge about the pathogenicity of miRNAs, involved genes and related disease classes can be revealed by co-clustering miRNAs, target genes and diseases simultaneously. Tripartite co-clustering can lead to more informative results than traditional co-clustering with only two kinds of members and pass the hidden relational information along the relation chain by considering multi-type members. Here we report a spectral co-clustering algorithm for k-partite graph to find clusters with heterogeneous members. We use the method to explore the potential relationships among miRNAs, genes and diseases. The clusters obtained from the algorithm have significantly higher density than randomly selected clusters, which means members in the same cluster are more likely to have common connections. Results also show that miRNAs in the same family based on the hairpin sequences tend to belong to the same cluster. We also validate the clustering results by checking the correlation of enriched gene functions and disease classes in the same cluster. Finally, widely studied miR-17-92 and its paralogs are analyzed as a case study to reveal that genes and diseases co-clustered with the miRNAs are in accordance with current research findings. PMID:22809308

  12. The expression of a viral microRNA is regulated by clustering to allow optimal B cell transformation.

    PubMed

    Haar, Janina; Contrant, Maud; Bernhardt, Katharina; Feederle, Regina; Diederichs, Sven; Pfeffer, Sébastien; Delecluse, Henri-Jacques

    2016-02-18

    The Epstein-Barr virus (EBV) transforms B cells by expressing latent proteins and the BHRF1 microRNA cluster. MiR-BHRF1-3, its most transforming member, belongs to the recently identified group of weakly expressed microRNAs. We show here that miR-BHRF1-3 displays an unusually low propensity to form a stem-loop structure, an effect potentiated by miR-BHRF1-3's proximity to the BHRF1 polyA site. Cloning miR-BHRF1-2 or a cellular microRNA, but not a ribozyme, 5' of miR-BHRF1-3 markedly enhanced its expression. However, a virus carrying mutated miR-BHRF1-2 seed regions expressed miR-BHRF1-3 at normal levels and was fully transforming. Therefore, miR-BHRF1-2's role during transformation is independent of its seed regions, revealing a new microRNA function. Increasing the distance between miR-BHRF1-2 and miR-BHRF1-3 in EBV enhanced miR-BHRF1-3's expression but decreased its transforming potential. Thus, the expression of some microRNAs must be restricted to a narrow range, as achieved by placing miR-BHRF1-3 under the control of miR-BHRF1-2. PMID:26635399

  13. The role, mechanism and potentially novel biomarker of microRNA-17-92 cluster in macrosomia

    PubMed Central

    Li, Jing; Chen, Liping; Qiuqin Tang; Wu, Wei; Hao Gu; Lou Liu; Jie Wu; Hua Jiang; Hongjuan Ding; Xia, Yankai; Chen, Daozhen; Hu, Yali; Wang, Xinru

    2015-01-01

    Macrosomia is one of the most common perinatal complications of pregnancy and has life-long health implications for the infant. microRNAs (miRNAs) have been identified to regulate placental development, yet the role of miRNAs in macrosomia remains poorly understood. Here we investigated the role of miR-17-92 cluster in macrosomia. The expression levels of five miRNAs in miR-17-92 cluster were significantly elevated in placentas of macrosomia, which may due to the up-regulation of miRNA-processing enzyme Drosha and Dicer. Cell cycle pathway was identified to be the most relevant pathways regulated by miR-17-92 cluster miRNAs. Importantly, miR-17-92 cluster increased proliferation, attenuated cell apoptosis and accelerated cells entering S phase by targeting SMAD4 and RB1 in HTR8/SVneo cells. Furthermore, we found that expression of miR-17-92 cluster in serum had a high diagnostic sensitivity and specificity for macrosomia (AUC: 80.53%; sensitivity: 82.61%; specificity: 69.57%). Our results suggested that miR-17-92 cluster contribute to macrosomia development by targeting regulators of cell cycle pathway. Our findings not only provide a novel insight into the molecular mechanisms of macrosomia, but also the clinical value of miR-17-92 cluster as a predictive biomarker for macrosomia. PMID:26598317

  14. High expression of microRNA-183/182/96 cluster as a prognostic biomarker for breast cancer

    PubMed Central

    Song, Cailu; Zhang, Lijuan; Wang, Jin; Huang, Zhongying; Li, Xing; Wu, Mingqing; Li, Shuaijie; Tang, Hailin; Xie, Xiaoming

    2016-01-01

    More sensitive and effective diagnostic markers for the detection of breast cancer are urgently needed. The microRNA-183/182/96 cluster has been reported to be involved in tumorigenesis and progression in a variety of cancers, and it is a promising cancer prognostic biomarker. The goal of this study was to determine the expression levels of the miR-183/182/96 cluster in breast cancer tissues and evaluate its prognostic role in breast cancer. Real-time quantitative polymerase chain reaction analysis (qRT-PCR) was used to detect the expression levels of the miR-183/182/96 cluster in 41 breast cancer tissues and adjacent normal tissues (control tissues) and also in different mammary cell lines. In situ hybridization (ISH) of the miR-183/182/96 cluster on 131 tissue microarrays (TMAs) was used to statistically analyze its prognostic role. The miR-183/182/96 cluster levels were significantly higher in breast cancer tissues than in control tissues. The miR-183/182/96 cluster was also upregulated in human breast cancer cell lines. An increased miR-183/182/96 cluster level was correlated with local relapse, distant metastasis and poor clinical outcomes. Our findings improve our understanding of the expression level of the miR-183/182/96 cluster in breast cancer and clarify the role of the miR-183/182/96 cluster as a novel prognostic biomarker for breast cancer. PMID:27071841

  15. High expression of microRNA-183/182/96 cluster as a prognostic biomarker for breast cancer.

    PubMed

    Song, Cailu; Zhang, Lijuan; Wang, Jin; Huang, Zhongying; Li, Xing; Wu, Mingqing; Li, Shuaijie; Tang, Hailin; Xie, Xiaoming

    2016-01-01

    More sensitive and effective diagnostic markers for the detection of breast cancer are urgently needed. The microRNA-183/182/96 cluster has been reported to be involved in tumorigenesis and progression in a variety of cancers, and it is a promising cancer prognostic biomarker. The goal of this study was to determine the expression levels of the miR-183/182/96 cluster in breast cancer tissues and evaluate its prognostic role in breast cancer. Real-time quantitative polymerase chain reaction analysis (qRT-PCR) was used to detect the expression levels of the miR-183/182/96 cluster in 41 breast cancer tissues and adjacent normal tissues (control tissues) and also in different mammary cell lines. In situ hybridization (ISH) of the miR-183/182/96 cluster on 131 tissue microarrays (TMAs) was used to statistically analyze its prognostic role. The miR-183/182/96 cluster levels were significantly higher in breast cancer tissues than in control tissues. The miR-183/182/96 cluster was also upregulated in human breast cancer cell lines. An increased miR-183/182/96 cluster level was correlated with local relapse, distant metastasis and poor clinical outcomes. Our findings improve our understanding of the expression level of the miR-183/182/96 cluster in breast cancer and clarify the role of the miR-183/182/96 cluster as a novel prognostic biomarker for breast cancer. PMID:27071841

  16. MicroRNA-183-96-182 Cluster Regulates Bovine Granulosa Cell Proliferation and Cell Cycle Transition by Coordinately Targeting FOXO1.

    PubMed

    Gebremedhn, Samuel; Salilew-Wondim, Dessie; Hoelker, Michael; Rings, Franca; Neuhoff, Christiane; Tholen, Ernst; Schellander, Karl; Tesfaye, Dawit

    2016-06-01

    Large-scale expression profiling of micro-RNAs (miRNAs) in bovine granulosa cells from dominant and subordinate follicles on Day 19 of the estrous cycle revealed enriched micro-RNA-183-96-182 cluster miRNAs in preovulatory dominant follicles that coordinately regulate the forkhead box protein O1 (FOXO1) gene. However, little is known about the role of this cluster in bovine granulosa cell function. We used an in vitro granulosa cell culture model to investigate this role. Granulosa cells aspirated from small growing follicles (3-5 mm in diameter) were cultured in Dulbecco modified Eagle medium/F-12 medium supplemented with fetal bovine serum and transfected with locked nucleic acid-based miRNA mimics, inhibitors, and corresponding negative controls. Overexpression of the miRNA cluster resulted in suppression of FOXO1 mRNA and protein, whereas inhibition of the cluster increased expression of FOXO1 mRNA. Overexpression also increased the relative rate of cell proliferation, whereas inhibition slowed it down. Similarly, the proportion of cells under G0/G1 arrest declined, whereas the ratio of cells in S phase increased in response to miR-183-96-182 overexpression. Selective knockdown of FOXO1 mRNA using anti-FOXO1 small interfering RNA increased the rate of granulosa cell proliferation, decreased the proportion of cells under G0/G1 arrest, and increased the proportion of cells in the S phase of cell cycle. Our data suggest that miR-183-96-182 cluster miRNAs promote proliferation and G1/S transition of bovine granulosa cells by coordinately targeting FOXO1, suggesting a critical role in granulosa cell function. MicroRNA-183-96-182 cluster regulates bovine granulosa cell function by targeting FOXO1 gene. PMID:27122636

  17. Sponge Transgenic Mouse Model Reveals Important Roles for the MicroRNA-183 (miR-183)/96/182 Cluster in Postmitotic Photoreceptors of the Retina*

    PubMed Central

    Zhu, Qubo; Sun, Wenyu; Okano, Kiichiro; Chen, Yu; Zhang, Ning; Maeda, Tadao; Palczewski, Krzysztof

    2011-01-01

    MicroRNA-183 (miR-183), miR-96, and miR-182 comprising the miR-183/96/182 cluster are highly expressed in photoreceptor cells. Although in vitro data have indicated an important role for this cluster in the retina, details of its in vivo biological activity are still unknown. To observe the impact of the miR-183/96/182 cluster on retinal maintenance and light adaptation, we generated a sponge transgenic mouse model that disrupted the activities of the three-component microRNAs simultaneously and selectively in the retina. Although our morphological and functional studies showed no differences between transgenic and wild type mice under normal laboratory lighting conditions, sponge transgenic mice displayed severe retinal degeneration after 30 min of exposure to 10,000 lux light. Histological studies showed that the outer nuclear layer thickness was dramatically reduced in the superior retina of transgenic mice. Real time PCR experiments in both the sponge transgenic mouse model and different microRNA stable cell lines identified Arrdc3, Neurod4, and caspase-2 (Casp2) as probable downstream targets of this cluster, a result also supported by luciferase assay and immunoblotting analyses. Further studies indicated that expression of both the cluster and Casp2 increased in response to light exposure. Importantly, Casp2 expression was enhanced in transgenic mice, and inhibition of Casp2 partially rescued their light-induced retinal degeneration. By connecting the microRNA and apoptotic pathways, these findings imply an important role for the miR-183/96/182 cluster in acute light-induced retinal degeneration of mice. This study demonstrates a clear involvement of miRs in the physiology of postmitotic cells in vivo. PMID:21768104

  18. The tumor-suppressive microRNA-23b/27b cluster regulates the MET oncogene in oral squamous cell carcinoma.

    PubMed

    Fukumoto, Ichiro; Koshizuka, Keiichi; Hanazawa, Toyoyuki; Kikkawa, Naoko; Matsushita, Ryosuke; Kurozumi, Akira; Kato, Mayuko; Okato, Atsushi; Okamoto, Yoshitaka; Seki, Naohiko

    2016-09-01

    Our recent studies of microRNA (miRNA) expression signatures in human cancers revealed that two clustered miRNAs, microRNA-23b (miR-23b) and microRNA-27b (miR‑27b), were significantly reduced in cancer tissues. Few reports have provided functional analyses of these clustered miRNAs in oral squamous cell carcinoma (OSCC). The aim of this study was to investigate the functional significance of miR-23b and miR-27b in OSCC and to identify novel miR-23b/27b-mediated cancer pathways and target genes involved in OSCC oncogenesis and metastasis. Expression levels of miR-23b and miR-27b were significantly reduced in OSCC specimens. Restoration of miR-23b or miR-27b in cancer cells revealed that both miRNAs significantly inhibited cancer cell migration and invasion. Our in silico analyses and luciferase reporter assays showed that the receptor tyrosine kinase MET, was directly regulated by these miRNAs. Moreover, downregulating the MET gene by use of siRNA significantly inhibited cell migration and invasion by OSCC cells. The identification of novel molecular pathways regulated by miR-23b and miR-27b may lead to a better understanding of the oncogenesis and metastasis of this disease. PMID:27573718

  19. Novel and Recently Evolved MicroRNA Clusters Regulate Expansive F-BOX Gene Networks through Phased Small Interfering RNAs in Wild Diploid Strawberry1[OPEN

    PubMed Central

    Xia, Rui; Ye, Songqing; Liu, Zongrang; Meyers, Blake C.; Liu, Zhongchi

    2015-01-01

    The wild strawberry (Fragaria vesca) has recently emerged as an excellent model for cultivated strawberry (Fragaria × ananassa) as well as other Rosaceae fruit crops due to its short seed-to-fruit cycle, diploidy, and sequenced genome. Deep sequencing and parallel analysis of RNA ends were used to identify F. vesca microRNAs (miRNAs) and their target genes, respectively. Thirty-eight novel and 31 known miRNAs were identified. Many known miRNAs targeted not only conserved mRNA targets but also developed new target genes in F. vesca. Significantly, two new clusters of miRNAs were found to collectively target 94 F-BOX (FBX) genes. One of the miRNAs in the new cluster is 22 nucleotides and triggers phased small interfering RNA production from six FBX genes, which amplifies the silencing to additional FBX genes. Comparative genomics revealed that the main novel miRNA cluster evolved from duplications of FBX genes. Finally, conserved trans-acting siRNA pathways were characterized and confirmed with distinct features. Our work identified novel miRNA-FBX networks in F. vesca and shed light on the evolution of miRNAs/phased small interfering RNA networks that regulate large gene families in higher plants. PMID:26143249

  20. Novel and Recently Evolved MicroRNA Clusters Regulate Expansive F-BOX Gene Networks through Phased Small Interfering RNAs in Wild Diploid Strawberry.

    PubMed

    Xia, Rui; Ye, Songqing; Liu, Zongrang; Meyers, Blake C; Liu, Zhongchi

    2015-09-01

    The wild strawberry (Fragaria vesca) has recently emerged as an excellent model for cultivated strawberry (Fragaria × ananassa) as well as other Rosaceae fruit crops due to its short seed-to-fruit cycle, diploidy, and sequenced genome. Deep sequencing and parallel analysis of RNA ends were used to identify F. vesca microRNAs (miRNAs) and their target genes, respectively. Thirty-eight novel and 31 known miRNAs were identified. Many known miRNAs targeted not only conserved mRNA targets but also developed new target genes in F. vesca. Significantly, two new clusters of miRNAs were found to collectively target 94 F-BOX (FBX) genes. One of the miRNAs in the new cluster is 22 nucleotides and triggers phased small interfering RNA production from six FBX genes, which amplifies the silencing to additional FBX genes. Comparative genomics revealed that the main novel miRNA cluster evolved from duplications of FBX genes. Finally, conserved trans-acting siRNA pathways were characterized and confirmed with distinct features. Our work identified novel miRNA-FBX networks in F. vesca and shed light on the evolution of miRNAs/phased small interfering RNA networks that regulate large gene families in higher plants. PMID:26143249

  1. Prognostic Role of MicroRNA-200c-141 Cluster in Various Human Solid Malignant Neoplasms

    PubMed Central

    Li, Xiao-yang; Li, Hui; Bu, Jie; Xiong, Liang; Guo, Hong-bin; Liu, Li-hong; Xiao, Tao

    2015-01-01

    The miR-200 family has emerged recently as a noticeable marker for predicting cancer prognosis and tumor progression. We aimed to review the evidence of miR-200c-141 genomic cluster as prognostic biomarkers in cancers. The results suggested that high level of miR-200c had no significant impact on OS (HR = 1.14 [0.77–1.69], P = 0.501) and DFS/PFS (HR = 0.72 [0.45–1.14], P = 0.161). Stratified analyses revealed that high miR-200c expression was significantly related to poor OS in serum/plasma (HR = 2.12 [1.62–2.77], P = 0.000) but not in tissues (HR = 0.89 [0.58–1.37], P = 0.599). High miR-200c expression was significantly associated with favorable DFS/PFS in tissues (HR = 0.56 [0.43–0.73], P = 0.000) but worse DFS/PFS in serum/plasma (HR = 1.90 [1.08–3.36], P = 0.027). For miR-141, we found that high miR-141 expression predicted no significant impact on OS (HR = 1.18 [0.74–1.88], P = 0.482) but poor DFS/PFS (HR = 1.11 [1.04–1.20], P = 0.003). Similarly, subgroup analyses showed that high miR-141 expression predicted poor OS in serum/plasma (HR = 4.34 [2.30–8.21], P = 0.000) but not in tissues (HR = 1.00 [0.92–1.09], P = 0.093). High miR-141 expression was significantly associated with worse DFS/PFS in tissues (HR = 1.12 [1.04–1.20], P = 0.002) but not in serum/plasma (HR = 0.90 [0.44–1.83], P = 0.771). Our findings indicated that, compared to their tissue counterparts, the expression level of miR-200c and miR-141 in peripheral blood may be more effective for monitoring cancer prognosis. High miR-141 expression was better at predicting tumor progression than survival for malignant tumors. PMID:26556949

  2. Phenotypic MicroRNA Microarrays

    PubMed Central

    Kwon, Yong-Jun; Heo, Jin Yeong; Kim, Hi Chul; Kim, Jin Yeop; Liuzzi, Michel; Soloveva, Veronica

    2013-01-01

    Microarray technology has become a very popular approach in cases where multiple experiments need to be conducted repeatedly or done with a variety of samples. In our lab, we are applying our high density spots microarray approach to microscopy visualization of the effects of transiently introduced siRNA or cDNA on cellular morphology or phenotype. In this publication, we are discussing the possibility of using this micro-scale high throughput process to study the role of microRNAs in the biology of selected cellular models. After reverse-transfection of microRNAs and siRNA, the cellular phenotype generated by microRNAs regulated NF-κB expression comparably to the siRNA. The ability to print microRNA molecules for reverse transfection into cells is opening up the wide horizon for the phenotypic high content screening of microRNA libraries using cellular disease models.

  3. A Viral microRNA Cluster Regulates the Expression of PTEN, p27 and of a bcl-2 Homolog.

    PubMed

    Bernhardt, Katharina; Haar, Janina; Tsai, Ming-Han; Poirey, Remy; Feederle, Regina; Delecluse, Henri-Jacques

    2016-01-01

    The Epstein-Barr virus (EBV) infects and transforms B-lymphocytes with high efficiency. This process requires expression of the viral latent proteins and of the 3 miR-BHRF1 microRNAs. Here we show that B-cells infected by a virus that lacks these non-coding RNAs (Δ123) grew more slowly between day 5 and day 20, relative to wild type controls. This effect could be ascribed to a reduced S phase entry combined with a moderately increased apoptosis rate. Whilst the first phenotypic trait was consistent with an enhanced PTEN expression in B-cells infected with Δ123, the second could be explained by very low BHRF1 protein and RNA levels in the same cells. Indeed, B-cells infected either by a recombinant virus that lacks the BHRF1 protein, a viral bcl-2 homolog, or by Δ123 underwent a similar degree of apoptosis, whereas knockouts of both BHRF1 microRNAs and protein proved transformation-incompetent. We find that that the miR-BHRF1-3 seed regions, and to a lesser extent those of miR-BHRF1-2 mediate these stimulatory effects. After this critical period, B-cells infected with the Δ123 mutant recovered a normal growth rate and became more resistant to provoked apoptosis. This resulted from an enhanced BHRF1 protein expression relative to cells infected with wild type viruses and correlated with decreased p27 expression, two pro-oncogenic events. The upregulation of BHRF1 can be explained by the observation that large BHRF1 mRNAs are the source of BHRF1 protein but are destroyed following BHRF1 microRNA processing, in particular of miR-BHRF1-2. The BHRF1 microRNAs are unlikely to directly target p27 but their absence may facilitate the selection of B-cells that express low levels of this protein. Thus, the BHRF1 microRNAs allowed a time-restricted expression of the BHRF1 protein to innocuously expand the virus B-cell reservoir during the first weeks post-infection without increasing long-term immune pressure. PMID:26800049

  4. A Viral microRNA Cluster Regulates the Expression of PTEN, p27 and of a bcl-2 Homolog

    PubMed Central

    Bernhardt, Katharina; Haar, Janina; Tsai, Ming-Han; Poirey, Remy

    2016-01-01

    The Epstein-Barr virus (EBV) infects and transforms B-lymphocytes with high efficiency. This process requires expression of the viral latent proteins and of the 3 miR-BHRF1 microRNAs. Here we show that B-cells infected by a virus that lacks these non-coding RNAs (Δ123) grew more slowly between day 5 and day 20, relative to wild type controls. This effect could be ascribed to a reduced S phase entry combined with a moderately increased apoptosis rate. Whilst the first phenotypic trait was consistent with an enhanced PTEN expression in B-cells infected with Δ123, the second could be explained by very low BHRF1 protein and RNA levels in the same cells. Indeed, B-cells infected either by a recombinant virus that lacks the BHRF1 protein, a viral bcl-2 homolog, or by Δ123 underwent a similar degree of apoptosis, whereas knockouts of both BHRF1 microRNAs and protein proved transformation-incompetent. We find that that the miR-BHRF1-3 seed regions, and to a lesser extent those of miR-BHRF1-2 mediate these stimulatory effects. After this critical period, B-cells infected with the Δ123 mutant recovered a normal growth rate and became more resistant to provoked apoptosis. This resulted from an enhanced BHRF1 protein expression relative to cells infected with wild type viruses and correlated with decreased p27 expression, two pro-oncogenic events. The upregulation of BHRF1 can be explained by the observation that large BHRF1 mRNAs are the source of BHRF1 protein but are destroyed following BHRF1 microRNA processing, in particular of miR-BHRF1-2. The BHRF1 microRNAs are unlikely to directly target p27 but their absence may facilitate the selection of B-cells that express low levels of this protein. Thus, the BHRF1 microRNAs allowed a time-restricted expression of the BHRF1 protein to innocuously expand the virus B-cell reservoir during the first weeks post-infection without increasing long-term immune pressure. PMID:26800049

  5. The MicroRNA-23b/27b/24 Cluster Promotes Breast Cancer Lung Metastasis by Targeting Metastasis-suppressive Gene Prosaposin

    PubMed Central

    Ell, Brian; Qiu, Qiong; Wei, Yong; Mercatali, Laura; Ibrahim, Toni; Amadori, Dino; Kang, Yibin

    2014-01-01

    MicroRNAs (miRNAs) have been shown to function as key regulators of tumor progression and metastasis. Recent studies have indicated that the miRNAs comprising the miR-23b/27b/24 cluster might influence tumor metastasis, although the precise nature of this regulation remains unclear. Here, expression of the miR-23b/27b/24 cluster is found to correlate with metastatic potential in mouse and human breast cancer cell lines and is elevated in metastatic lung lesions in human breast cancer patients. Ectopic expression of the miRNAs in the weakly metastatic mouse 4TO7 mammary tumor cell line had no effect on proliferation or morphology of tumor cells in vitro but was found to increase lung metastasis in a mouse model of breast cancer metastasis. Furthermore, gene expression profiling analysis of miRNA overexpressing 4TO7 cells revealed the direct targeting of prosaposin (PSAP), which encodes a secreted protein found to be inversely correlated with metastatic progression in human breast cancer patients. Importantly, ectopic expression of PSAP was able to suppress the metastatic phenotype in highly metastatic 4T1 and MDA-MB-231 SCP28 cells, as well as in cells ectopically expressing miR-23b/27b/24. These findings support a metastasis-promoting function of the miR-23b/27b/24 cluster of miRNAs, which functions in part through the direct inhibition of PSAP. PMID:24966325

  6. Allergen-specific immune response suppresses interleukin 10 expression in B cells via increasing micro-RNA-17-92 cluster.

    PubMed

    Geng, Xiao-Rui; Qiu, Shu-Qi; Yang, Li-Tao; Liu, Zhi-Qiang; Yang, Gui; Liu, Jiang-Qi; Zeng, Lu; Li, Xiao-Xi; Mo, Li-Hua; Liu, Zhi-Gang; Yang, Ping-Chang

    2016-08-01

    Interleukin (IL)-10-expressing B cells play a critical role in the immune homeostasis in the body; its regulation has not been fully understood. Micro-RNA (miR)-17-92 cluster has strong regulation in the immunity. This study tests a hypothesis that miR-17-92 cluster suppresses IL-10 expression in B cells. In this study, peripheral B cells were collected from patients with allergic rhinitis (AR). The B cells were treated with specific allergens, dust mite extracts, in the culture. The expressions of miR-17-92 cluster and IL-10 in the culture were assessed by real-time quantitative reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay. The results showed that the levels of miR-19a, but not the rest of the 5 members (miR-17, miR-18a, miR-19b, miR-20a, and miR-92a), were significantly higher in peripheral B cells from AR patients as than in B cells from healthy participants. Exposure of B cells from AR patients to specific allergen, dust mite extracts, significantly increased the levels if miR-19a and suppressed the expression of IL-10 in B cells. The levels of histone deacetylase 11 and acetylated H3K9 were higher, and the RNA polymerase II and c-Maf (the IL-10 transcription factor) were lower, at the IL-10 promoter locus. In conclusion, miR-19a mediates the allergen-specific immune response-decreased IL-10 expression in B cells. PMID:27491928

  7. Modulation of MicroRNA Cluster miR-183-96-182 Expression by Epstein-Barr Virus Latent Membrane Protein 1

    PubMed Central

    Oussaief, Lassad; Fendri, Ali; Chane-Woon-Ming, Béatrice; Poirey, Remy; Delecluse, Henri-Jacques; Joab, Irène

    2015-01-01

    ABSTRACT Epstein-Barr virus (EBV) is an oncogenic human herpesvirus involved in the pathogenesis of Burkitt's lymphoma (BL) and various other lymphoproliferative disorders. In BL, EBV protein expression is restricted to EBV nuclear antigen 1 (EBNA1), but small noncoding RNAs such as EBV-encoded small RNAs (EBERs) and microRNAs (miRNAs) can also be detected. miRNAs play major roles in crucial processes such as proliferation, differentiation, and cell death. It has recently become clear that alterations in the expression profile of miRNAs contribute to the pathogenesis of a number of malignancies. During latent infection, EBV expresses 25 viral pre-miRNAs and modulates the expression of specific cellular miRNAs, such as miR-155 and miR-146, which potentially play a role in oncogenesis. Here, we established the small-RNA expression profiles of three BL cell lines. Using large-scale sequencing coupled to Northern blotting and real-time reverse transcription-PCR (RT-PCR) analysis validation, we demonstrated the differential expression of some cellular and viral miRNAs. High-level expression of the miR-183-96-182 cluster and EBV miR-BamHI A rightward transcript (miR-BART) cluster was significantly associated with EBV type I latency. This expression was not affected by viral reactivation since transforming growth factor β1 (TGF-β1) stimulation did not significantly change the miRNA profiles. However, using several approaches, including de novo infection with a mutant virus, we present evidence that the expression of latent membrane protein 1 (LMP-1) triggered downregulation of the expression of the miR-183-96-182 cluster. We further show that this effect involves the Akt signaling pathway. IMPORTANCE In addition to expressing their own miRNAs, herpesviruses also impact the expression levels of cellular miRNAs. This regulation can be either positive or negative and usually results in the perturbation of pathways to create a cellular environment that is more

  8. MicroRNA expression profiling using microarrays.

    PubMed

    Love, Cassandra; Dave, Sandeep

    2013-01-01

    MicroRNAs are small noncoding RNAs which are able to regulate gene expression at both the transcriptional and translational levels. There is a growing recognition of the role of microRNAs in nearly every tissue type and cellular process. Thus there is an increasing need for accurate quantitation of microRNA expression in a variety of tissues. Microarrays provide a robust method for the examination of microRNA expression. In this chapter, we describe detailed methods for the use of microarrays to measure microRNA expression and discuss methods for the analysis of microRNA expression data. PMID:23666707

  9. Pervasive microRNA Duplication in Chelicerates: Insights from the Embryonic microRNA Repertoire of the Spider Parasteatoda tepidariorum.

    PubMed

    Leite, Daniel J; Ninova, Maria; Hilbrant, Maarten; Arif, Saad; Griffiths-Jones, Sam; Ronshaugen, Matthew; McGregor, Alistair P

    2016-01-01

    MicroRNAs are small (∼22 nt) noncoding RNAs that repress translation and therefore regulate the production of proteins from specific target mRNAs. microRNAs have been found to function in diverse aspects of gene regulation within animal development and many other processes. Among invertebrates, both conserved and novel, lineage specific, microRNAs have been extensively studied predominantly in holometabolous insects such as Drosophila melanogaster However little is known about microRNA repertoires in other arthropod lineages such as the chelicerates. To understand the evolution of microRNAs in this poorly sampled subphylum, we characterized the microRNA repertoire expressed during embryogenesis of the common house spider Parasteatoda tepidariorum We identified a total of 148 microRNAs in P. tepidariorum representing 66 families. Approximately half of these microRNA families are conserved in other metazoans, while the remainder are specific to this spider. Of the 35 conserved microRNAs families 15 had at least two copies in the P. tepidariorum genome. A BLAST-based approach revealed a similar pattern of duplication in other spiders and a scorpion, but not among other chelicerates and arthropods, with the exception of a horseshoe crab. Among the duplicated microRNAs we found examples of lineage-specific tandem duplications, and the duplication of entire microRNA clusters in three spiders, a scorpion, and in a horseshoe crab. Furthermore, we found that paralogs of many P. tepidariorum microRNA families exhibit arm switching, which suggests that duplication was often followed by sub- or neofunctionalization. Our work shows that understanding the evolution of microRNAs in the chelicerates has great potential to provide insights into the process of microRNA duplication and divergence and the evolution of animal development. PMID:27324919

  10. Pervasive microRNA Duplication in Chelicerates: Insights from the Embryonic microRNA Repertoire of the Spider Parasteatoda tepidariorum

    PubMed Central

    Leite, Daniel J.; Ninova, Maria; Hilbrant, Maarten; Arif, Saad; Griffiths-Jones, Sam; Ronshaugen, Matthew; McGregor, Alistair P.

    2016-01-01

    MicroRNAs are small (∼22 nt) noncoding RNAs that repress translation and therefore regulate the production of proteins from specific target mRNAs. microRNAs have been found to function in diverse aspects of gene regulation within animal development and many other processes. Among invertebrates, both conserved and novel, lineage specific, microRNAs have been extensively studied predominantly in holometabolous insects such as Drosophila melanogaster. However little is known about microRNA repertoires in other arthropod lineages such as the chelicerates. To understand the evolution of microRNAs in this poorly sampled subphylum, we characterized the microRNA repertoire expressed during embryogenesis of the common house spider Parasteatoda tepidariorum. We identified a total of 148 microRNAs in P. tepidariorum representing 66 families. Approximately half of these microRNA families are conserved in other metazoans, while the remainder are specific to this spider. Of the 35 conserved microRNAs families 15 had at least two copies in the P. tepidariorum genome. A BLAST-based approach revealed a similar pattern of duplication in other spiders and a scorpion, but not among other chelicerates and arthropods, with the exception of a horseshoe crab. Among the duplicated microRNAs we found examples of lineage-specific tandem duplications, and the duplication of entire microRNA clusters in three spiders, a scorpion, and in a horseshoe crab. Furthermore, we found that paralogs of many P. tepidariorum microRNA families exhibit arm switching, which suggests that duplication was often followed by sub- or neofunctionalization. Our work shows that understanding the evolution of microRNAs in the chelicerates has great potential to provide insights into the process of microRNA duplication and divergence and the evolution of animal development. PMID:27324919

  11. Quantitative Model of microRNA-mRNA interaction

    NASA Astrophysics Data System (ADS)

    Noorbakhsh, Javad; Lang, Alex; Mehta, Pankaj

    2012-02-01

    MicroRNAs are short RNA sequences that regulate gene expression and protein translation by binding to mRNA. Experimental data reveals the existence of a threshold linear output of protein based on the expression level of microRNA. To understand this behavior, we propose a mathematical model of the chemical kinetics of the interaction between mRNA and microRNA. Using this model we have been able to quantify the threshold linear behavior. Furthermore, we have studied the effect of internal noise, showing the existence of an intermediary regime where the expression level of mRNA and microRNA has the same order of magnitude. In this crossover regime the mRNA translation becomes sensitive to small changes in the level of microRNA, resulting in large fluctuations in protein levels. Our work shows that chemical kinetics parameters can be quantified by studying protein fluctuations. In the future, studying protein levels and their fluctuations can provide a powerful tool to study the competing endogenous RNA hypothesis (ceRNA), in which mRNA crosstalk occurs due to competition over a limited pool of microRNAs.

  12. The Three Paralogous MicroRNA Clusters in Development and Disease, miR-17-92, miR-106a-363, and miR-106b-25

    PubMed Central

    Sehic, Amer

    2016-01-01

    MicroRNAs (miRNAs) form a class of noncoding RNA genes whose products are small single-stranded RNAs that are involved in the regulation of translation and degradation of mRNAs. There is a fine balance between deregulation of normal developmental programs and tumor genesis. An increasing body of evidence suggests that altered expression of miRNAs is entailed in the pathogenesis of human cancers. Studies in mouse and human cells have identified the miR-17-92 cluster as a potential oncogene. The miR-17-92 cluster is often amplified or overexpressed in human cancers and has recently emerged as the prototypical oncogenic polycistron miRNA. The functional analysis of miR-17-92 is intricate by the existence of two paralogues: miR-106a-363 and miR-106b-25. During early evolution of vertebrates, it is likely that the three clusters commenced via a series of duplication and deletion occurrences. As miR-106a-363 and miR-106b-25 contain miRNAs that are very similar, and in some cases identical, to those encoded by miR-17-92, it is feasible that they regulate a similar set of genes and have overlapping functions. Further understanding of these three clusters and their functions will increase our knowledge about cancer progression. The present review discusses the characteristics and functions of these three miRNA clusters. PMID:27127675

  13. microRNA: Diagnostic Perspective

    PubMed Central

    Faruq, Omar; Vecchione, Andrea

    2015-01-01

    Biomarkers are biological measures of a biological state. An ideal marker should be safe and easy to measure, cost efficient, modifiable with treatment, and consistent across gender and ethnic groups. To date, none of the available biomarkers satisfy all of these criteria. In addition, the major limitations of these markers are low specificity, sensitivity, and false positive results. Recently identified, microRNAs (miRNAs) are endogenous, evolutionarily conserved small non-coding RNA (about 22–25 nt long), also known as micro-coordinators of gene expression, which have been shown to be an effective tools to study the biology of diseases and to have great potential as novel diagnostic and prognostic biomarkers with high specificity and sensitivity. In fact, it has been demonstrated that miRNAs play a pivotal role in the regulation of a wide range of developmental and physiological processes and their deficiencies have been related to a number of disease. In addition, miRNAs are stable and can be easily isolated and measured from tissues and body fluids. In this review, we provide a perspective on emerging concepts and potential usefulness of miRNAs as diagnostic markers, emphasizing the involvement of specific miRNAs in particular tumor types, subtypes, cardiovascular diseases, diabetes, infectious diseases, and forensic test. PMID:26284247

  14. Identification of the miR-106b∼25 MicroRNA Cluster as a Proto-Oncogenic PTEN-Targeting Intron That Cooperates with Its Host Gene MCM7 in Transformation

    PubMed Central

    Poliseno, Laura; Salmena, Leonardo; Riccardi, Luisa; Fornari, Alessandro; Song, Min Sup; Hobbs, Robin M.; Sportoletti, Paolo; Varmeh, Shorheh; Egia, Ainara; Fedele, Giuseppe; Rameh, Lucia; Loda, Massimo; Pandolfi, Pier Paolo

    2010-01-01

    PTEN (phosphatase and tensin homolog deleted on chromosome 10) is a tumor suppressor that antagonizes signaling through the phosphatidylinositol-3-kinase–Akt pathway. We have demonstrated that subtle decreases in PTEN abundance can have critical consequences for tumorigenesis. Here, we used a computational approach to identify miR-22, miR-25, and miR-302 as three PTEN-targeting microRNA (miRNA) families found within nine genomic loci. We showed that miR-22 and the miR-106b∼25 cluster are aberrantly overexpressed in human prostate cancer, correlate with abundance of the miRNA processing enzyme DICER, and potentiate cellular transformation both in vitro and in vivo. We demonstrated that the intronic miR-106b∼25 cluster cooperates with its host gene MCM7 in cellular transformation both in vitro and in vivo, so that the concomitant overexpression of MCM7 and the miRNA cluster triggers prostatic intraepithelial neoplasia in transgenic mice. Therefore, the MCM7 gene locus delivers two simultaneous oncogenic insults when amplified or overexpressed in human cancer. Thus, we have uncovered a proto-oncogenic miRNA-dependent network for PTEN regulation and defined the MCM7 locus as a critical factor in initiating prostate tumorigenesis. PMID:20388916

  15. Endogenous MCM7 MicroRNA Cluster as a Novel Platform to Multiplex Small Interfering and Nucleolar RNAs for Combinational HIV-1 Gene Therapy

    PubMed Central

    Chung, Janet; Zhang, Jane; Li, Haitang; Ouellet, Dominique L.; DiGiusto, David L.

    2012-01-01

    Abstract Combinational therapy with small RNA inhibitory agents against multiple viral targets allows efficient inhibition of viral production by controlling gene expression at critical time points. Here we explore combinations of different classes of therapeutic anti-HIV-1 RNAs expressed from within the context of an intronic MCM7 (minichromosome maintenance complex component-7) platform that naturally harbors 3 microRNAs (miRNAs). We replaced the endogenous miRNAs with anti-HIV small RNAs, including small interfering RNAs (siRNAs) targeting HIV-1 tat and rev messages that function to induce post-transcriptional gene silencing by the RNA interference pathway, a nucleolar-localizing RNA ribozyme that targets the conserved U5 region of HIV-1 transcripts for degradation, and finally nucleolar trans-activation response (TAR) and Rev-binding element (RBE) RNA decoys designed to sequester HIV-1 Tat and Rev proteins inside the nucleolus. We demonstrate the versatility of the MCM7 platform in expressing and efficiently processing the siRNAs as miRNA mimics along with nucleolar small RNAs. Furthermore, three of the combinatorial constructs tested potently suppressed viral replication during a 1-month HIV challenge, with greater than 5-log inhibition compared with untransduced, HIV-1-infected CEM T lymphocytes. One of the most effective constructs contains an anti-HIV siRNA combined with a nucleolar-localizing U5 ribozyme and TAR decoy. This represents the first efficacious example of combining Drosha-processed siRNAs with small nucleolar ribonucleoprotein (snoRNP)-processed nucleolar RNA chimeras from a single intron platform for effective inhibition of viral replication. Moreover, we demonstrated enrichment/selection for cells expressing levels of the antiviral RNAs that provide optimal inhibition under the selective pressure of HIV. The combinations of si/snoRNAs represent a new paradigm for combinatorial RNA-based gene therapy applications. PMID:22834872

  16. MicroRNA involvement in glioblastoma pathogenesis

    SciTech Connect

    Novakova, Jana; Slaby, Ondrej; Vyzula, Rostislav; Michalek, Jaroslav

    2009-08-14

    MicroRNAs are endogenously expressed regulatory noncoding RNAs. Altered expression levels of several microRNAs have been observed in glioblastomas. Functions and direct mRNA targets for these microRNAs have been relatively well studied over the last years. According to these data, it is now evident, that impairment of microRNA regulatory network is one of the key mechanisms in glioblastoma pathogenesis. MicroRNA deregulation is involved in processes such as cell proliferation, apoptosis, cell cycle regulation, invasion, glioma stem cell behavior, and angiogenesis. In this review, we summarize the current knowledge of miRNA functions in glioblastoma with an emphasis on its significance in glioblastoma oncogenic signaling and its potential to serve as a disease biomarker and a novel therapeutic target in oncology.

  17. MicroRNA and Metastasis.

    PubMed

    Ma, L

    2016-01-01

    Noncoding RNAs are important regulatory molecules of cellular processes. MicroRNAs (miRNAs) are small noncoding RNAs that bind to complementary sequences in the 3' untranslated region of target mRNAs, leading to degradation of the target mRNAs and/or inhibition of their translation. Some miRNAs are essential for normal animal development; however, many other miRNAs are dispensable for development but play a critical role in pathological conditions, including tumorigenesis and metastasis. miRNA genes often reside at fragile chromosome sites and are deregulated in cancer. Some miRNAs function as oncogenes or tumor suppressors, collectively termed "oncomirs." Specific metastasis-regulating miRNAs, collectively termed "metastamirs," govern molecular processes and pathways in malignant progression in either a tumor cell-autonomous or a cell-nonautonomous manner. Recently, exosome-transferred miRNAs have emerged as mediators of the tumor-stroma cross talk. In this chapter, we focus on the functions, mechanisms of action, and therapeutic potential of miRNAs, particularly oncomirs and metastamirs. PMID:27613133

  18. lncRNA/MicroRNA interactions in the vasculature

    PubMed Central

    Ballantyne, MD; McDonald, RA

    2016-01-01

    MicroRNA (miRNA) have gained widespread attention for their role in diverse vascular processes including angiogenesis, apoptosis, proliferation, and migration. Despite great understanding of miRNA expression and function, knowledge of long noncoding RNA (lncRNA) molecular mechanisms still remains limited. The influence of miRNA on lncRNA function, and the converse, is now beginning to emerge. lncRNA may regulate miRNA function by acting as endogenous sponges to regulate gene expression and miRNA have been shown to bind and regulate lncRNA stability. A detailed understanding of the molecular and cellular effects of lncRNA‐miRNA‐mediated interactions in vascular pathophysiology could pave the way for new diagnostic markers and therapeutic approaches, but first there is a requirement for a more detailed understanding of the impact of such regulatory networks. PMID:26910520

  19. microRNA-17 Is the Most Up-Regulated Member of the miR-17-92 Cluster during Early Colon Cancer Evolution

    PubMed Central

    Knudsen, Kirsten Nguyen; Nielsen, Boye Schnack; Lindebjerg, Jan; Hansen, Torben Frøstrup; Holst, René; Sørensen, Flemming Brandt

    2015-01-01

    Deregulated microRNAs play a role in the development and progression of colon cancer, but little is known about their tissue and cell distribution in the continuum of normal mucosa through the premalignant adenoma to invasive adenocarcinoma. The aim of this study was to examine the expression pattern of the miR-17-92 cluster (miR-17, miR-18, miR-19, miR-20 and miR-92) as well as miR-21, miR-31, miR-135b, and miR-145 in early clinically diagnosed colon cancer. MicroRNAs were analysed by chromogenic in situ hybridisation in the normal-adenoma-adenocarcinoma sequence of nine adenocarcinomas developed in mucosal colon polyps. Subsequently, the expression of selected microRNAs was validated in 24 mucosal colon cancer polyps. Expression of miR-17 was confined to the epithelial cells, and the expression levels increased in the transitional zone from normal to adenomatous tissue. The miR-17-92 cluster members, miR-19b, miR-20a, and miR-92a, followed the same expression pattern, but miR-17 was the most predominant. An increased expression of miR-21 was found in the tumour-associated stroma with the most dramatic increase from adenoma to adenocarcinoma, while the number of positive miR-145 fibroblast-like cells in the normal lamina propria (stroma) decreased in a stepwise manner throughout the normal-adenoma-adenocarcinoma sequence. It is concluded that the expression of miR-17, miR-21, and miR-145 changes at early stages of the normal-adenoma-adenocarcinoma sequence. Thus, these microRNAs may play a role in the development of colon cancer. PMID:26465597

  20. Characteristics of microRNA co-target networks

    NASA Astrophysics Data System (ADS)

    Lee, Chang-Yong

    2011-07-01

    The database of microRNAs and their predicted target genes in humans were used to extract a microRNA co-target network. Based on the finding that more than two miRNAs can target the same gene, we constructed a microRNA co-target network and analyzed it from the perspective of the complex network. We found that a network having a positive assortative mixing can be characterized by small-world and scale-free characteristics which are found in most complex networks. The network was further analyzed by the nearest-neighbor average connectivity, and it was shown that the more assortative a microRNA network is, the wider the range of increasing average connectivity. In particular, an assortative network has a power-law relationship of the average connectivity with a positive exponent. A percolation analysis of the network showed that, although the network is diluted, there is no percolation transition in the network. From these findings, we infer that the microRNAs in the network are clustered together, forming a core group. The same analyses carried out on different species confirmed the robustness of the main results found in the microRNA networks of humans.

  1. Principles of microRNA Regulation Revealed Through Modeling microRNA Expression Quantitative Trait Loci

    PubMed Central

    Budach, Stefan; Heinig, Matthias; Marsico, Annalisa

    2016-01-01

    Extensive work has been dedicated to study mechanisms of microRNA-mediated gene regulation. However, the transcriptional regulation of microRNAs themselves is far less well understood, due to difficulties determining the transcription start sites of transient primary transcripts. This challenge can be addressed using expression quantitative trait loci (eQTLs) whose regulatory effects represent a natural source of perturbation of cis-regulatory elements. Here we used previously published cis-microRNA-eQTL data for the human GM12878 cell line, promoter predictions, and other functional annotations to determine the relationship between functional elements and microRNA regulation. We built a logistic regression model that classifies microRNA/SNP pairs into eQTLs or non-eQTLs with 85% accuracy; shows microRNA-eQTL enrichment for microRNA precursors, promoters, enhancers, and transcription factor binding sites; and depletion for repressed chromatin. Interestingly, although there is a large overlap between microRNA eQTLs and messenger RNA eQTLs of host genes, 74% of these shared eQTLs affect microRNA and host expression independently. Considering microRNA-only eQTLs we find a significant enrichment for intronic promoters, validating the existence of alternative promoters for intragenic microRNAs. Finally, in line with the GM12878 cell line derived from B cells, we find genome-wide association (GWA) variants associated to blood-related traits more likely to be microRNA eQTLs than random GWA and non-GWA variants, aiding the interpretation of GWA results. PMID:27260304

  2. Functional MicroRNA Involved in Endometriosis

    PubMed Central

    Creighton, Chad J.; Han, Derek Y.; Zariff, Azam; Anderson, Matthew L.; Gunaratne, Preethi H.; Matzuk, Martin M.

    2011-01-01

    Endometriosis is a common disease seen by gynecologists. Clinical features involve pelvic pain and unexplained infertility. Although endometriosis is pathologically characterized by endometrial tissue outside the normal uterine location, endometriosis is otherwise not easily explained. Endometriomas, endometriotic cysts of the ovary, typically cause pain and distortion of pelvic anatomy. To begin to understand the pathogenesis of endometriomas, we describe the first transcriptome-microRNAome analysis of endometriomas and eutopic endometrium using next-generation sequencing technology. Using this approach, we generated a total of more than 54 million independent small RNA reads from our 19 clinical samples. At the microRNA level, we found 10 microRNA that were up-regulated (miR-202, 193a-3p, 29c, 708, 509-3-5p, 574-3p, 193a-5p, 485-3p, 100, and 720) and 12 microRNA that were down-regulated (miR-504, 141, 429, 203, 10a, 200b, 873, 200c, 200a, 449b, 375, and 34c-5p) in endometriomas compared with endometrium. Using in silico prediction algorithms, we correlated these microRNA with their corresponding differentially expressed mRNA targets. To validate the functional roles of microRNA, we manipulated levels of miR-29c in an in vitro system of primary cultures of human endometrial stromal fibroblasts. Extracellular matrix genes that were potential targets of miR-29c in silico were significantly down-regulated using this biological in vitro system. In vitro functional studies using luciferase reporter constructs further confirmed that miR-29c directly affects specific extracellular matrix genes that are dysregulated in endometriomas. Thus, miR-29c and other abnormally regulated microRNA appear to play important roles in the pathophysiology of uterine function and dysfunction. PMID:21436257

  3. MicroRNA: Mechanism of Gene Regulation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNA (miR) are a class of small RNAs that regulate gene expression by inhibiting translation of protein encoding transcripts through activation of a specific cellular pathway. The small RNA classified as miR are short sequences of 18-26 nucleotide long, encoded by nuclear genes with distinctive...

  4. Genome-wide profiling of the microRNA-mRNA regulatory network in skeletal muscle with aging

    PubMed Central

    Kim, Ji Young; Park, Young-Kyu; Lee, Kwang-Pyo; Lee, Seung-Min; Kang, Tae-Wook; Kim, Hee-Jin; Dho, So Hee; Kim, Seon-Young; Kwon, Ki-Sun

    2014-01-01

    Skeletal muscle degenerates progressively, losing mass (sarcopenia) over time, which leads to reduced physical ability and often results in secondary diseases such as diabetes and obesity. The regulation of gene expression by microRNAs is a key event in muscle development and disease. To understand genome-wide changes in microRNAs and mRNAs during muscle aging, we sequenced microRNAs and mRNAs from mouse gastrocnemius muscles at two different ages (6 and 24 months). Thirty-four microRNAs (15 up-regulated and 19 down-regulated) were differentially expressed with age, including the microRNAs miR-206 and -434, which were differentially expressed in aged muscle in previous studies. Interestingly, eight microRNAs in a microRNA cluster at the imprinted Dlk1-Dio3 locus on chromosome 12 were coordinately down-regulated. In addition, sixteen novel microRNAs were identified. Integrative analysis of microRNA and mRNA expression revealed that microRNAs may contribute to muscle aging through the positive regulation of transcription, metabolic processes, and kinase activity. Many of the age-related microRNAs have been implicated in human muscular diseases. We suggest that genome-wide microRNA profiling will expand our knowledge of microRNA function in the muscle aging process. PMID:25063768

  5. Evaluation of microRNA alignment techniques.

    PubMed

    Ziemann, Mark; Kaspi, Antony; El-Osta, Assam

    2016-08-01

    Genomic alignment of small RNA (smRNA) sequences such as microRNAs poses considerable challenges due to their short length (∼21 nucleotides [nt]) as well as the large size and complexity of plant and animal genomes. While several tools have been developed for high-throughput mapping of longer mRNA-seq reads (>30 nt), there are few that are specifically designed for mapping of smRNA reads including microRNAs. The accuracy of these mappers has not been systematically determined in the case of smRNA-seq. In addition, it is unknown whether these aligners accurately map smRNA reads containing sequence errors and polymorphisms. By using simulated read sets, we determine the alignment sensitivity and accuracy of 16 short-read mappers and quantify their robustness to mismatches, indels, and nontemplated nucleotide additions. These were explored in the context of a plant genome (Oryza sativa, ∼500 Mbp) and a mammalian genome (Homo sapiens, ∼3.1 Gbp). Analysis of simulated and real smRNA-seq data demonstrates that mapper selection impacts differential expression results and interpretation. These results will inform on best practice for smRNA mapping and enable more accurate smRNA detection and quantification of expression and RNA editing. PMID:27284164

  6. MicroRNA biogenesis pathways in cancer

    PubMed Central

    Lin, Shuibin; Gregory, Richard I.

    2016-01-01

    MicroRNAs (miRNAs) are critical regulators of gene expression. Amplification and overexpression of individual ‘oncomiRs’ or genetic loss of tumour suppressor miRNAs are associated with human cancer and are sufficient to drive tumorigenesis in mouse models. Furthermore, global miRNA depletion caused by genetic and epigenetic alterations in components of the miRNA biogenesis machinery is oncogenic. This, together with the recent identification of novel miRNA regulatory factors and pathways, highlights the importance of miRNA dysregulation in cancer. PMID:25998712

  7. Transcriptome dynamics of the microRNA inhibition response.

    PubMed

    Wen, Jiayu; Leucci, Elenora; Vendramin, Roberto; Kauppinen, Sakari; Lund, Anders H; Krogh, Anders; Parker, Brian J

    2015-07-27

    We report a high-resolution time series study of transcriptome dynamics following antimiR-mediated inhibition of miR-9 in a Hodgkin lymphoma cell-line-the first such dynamic study of the microRNA inhibition response-revealing both general and specific aspects of the physiological response. We show miR-9 inhibition inducing a multiphasic transcriptome response, with a direct target perturbation before 4 h, earlier than previously reported, amplified by a downstream peak at ∼32 h consistent with an indirect response due to secondary coherent regulation. Predictive modelling indicates a major role for miR-9 in post-transcriptional control of RNA processing and RNA binding protein regulation. Cluster analysis identifies multiple co-regulated gene regulatory modules. Functionally, we observe a shift over time from mRNA processing at early time points to translation at later time points. We validate the key observations with independent time series qPCR and we experimentally validate key predicted miR-9 targets. Methodologically, we developed sensitive functional data analytic predictive methods to analyse the weak response inherent in microRNA inhibition experiments. The methods of this study will be applicable to similar high-resolution time series transcriptome analyses and provides the context for more accurate experimental design and interpretation of future microRNA inhibition studies. PMID:26089393

  8. Seed microRNA Research

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNAs (miRNAs) are key regulatory molecules that play critical roles in gene expression. The biological functions of miRNAs are important for developmental processes in plants and animals. Little is known about the functions of miRNAs in seeds. To gain a better understand-ing of the regulation o...

  9. Chicken rRNA Gene Cluster Structure

    PubMed Central

    Dyomin, Alexander G.; Koshel, Elena I.; Kiselev, Artem M.; Saifitdinova, Alsu F.; Galkina, Svetlana A.; Fukagawa, Tatsuo; Kostareva, Anna A.

    2016-01-01

    Ribosomal RNA (rRNA) genes, whose activity results in nucleolus formation, constitute an extremely important part of genome. Despite the extensive exploration into avian genomes, no complete description of avian rRNA gene primary structure has been offered so far. We publish a complete chicken rRNA gene cluster sequence here, including 5’ETS (1836 bp), 18S rRNA gene (1823 bp), ITS1 (2530 bp), 5.8S rRNA gene (157 bp), ITS2 (733 bp), 28S rRNA gene (4441 bp) and 3’ETS (343 bp). The rRNA gene cluster sequence of 11863 bp was assembled from raw reads and deposited to GenBank under KT445934 accession number. The assembly was validated through in situ fluorescent hybridization analysis on chicken metaphase chromosomes using computed and synthesized specific probes, as well as through the reference assembly against de novo assembled rRNA gene cluster sequence using sequenced fragments of BAC-clone containing chicken NOR (nucleolus organizer region). The results have confirmed the chicken rRNA gene cluster validity. PMID:27299357

  10. The Two Stem Cell MicroRNA Gene Clusters C19MC and miR-371-3 Are Activated by Specific Chromosomal Rearrangements in a Subgroup of Thyroid Adenomas

    PubMed Central

    Rippe, Volkhard; Dittberner, Lea; Lorenz, Verena N.; Drieschner, Norbert; Nimzyk, Rolf; Sendt, Wolfgang; Junker, Klaus; Belge, Gazanfer; Bullerdiek, Jörn

    2010-01-01

    Thyroid adenomas are common benign human tumors with a high prevalence of about 5% of the adult population even in iodine sufficient areas. Rearrangements of chromosomal band 19q13.4 represent a frequent clonal cytogenetic deviation in these tumors making them the most frequent non-random chromosomal translocations in human epithelial tumors at all. Two microRNA (miRNA) gene clusters i.e. C19MC and miR-371-3 are located in close proximity to the breakpoint region of these chromosomal rearrangements and have been checked for a possible up-regulation due to the genomic alteration. In 4/5 cell lines established from thyroid adenomas with 19q13.4 rearrangements and 5/5 primary adenomas with that type of rearrangement both the C19MC and miR-371-3 cluster were found to be significantly overexpressed compared to controls lacking that particular chromosome abnormality. In the remaining cell line qRT-PCR revealed overexpression of members of the miR-371-3 cluster only which might be due to a deletion accompanying the chromosomal rearrangement in that case. In depth molecular characterization of the breakpoint in a cell line from one adenoma of this type reveals the existence of large Pol-II mRNA fragments as the most likely source of up-regulation of the C19MC cluster. The up-regulation of the clusters is likely to be causally associated with the pathogenesis of the corresponding tumors. Of note, the expression of miRNAs miR-520c and miR-373 is known to characterize stem cells and in terms of molecular oncology has been implicated in invasive growth of epithelial cells in vitro and in vivo thus allowing to delineate a distinct molecular subtype of thyroid adenomas. Besides thyroid adenomas rearrangements of 19q13.4 are frequently found in other human neoplasias as well, suggesting that activation of both clusters might be a more general phenomenon in human neoplasias. PMID:20209130

  11. Long-term marker-free multiphoton imaging, targeted transfection, optical cleaning of stem cell clusters, and optical transport of microRNA with extreme ultrashort laser pulses

    NASA Astrophysics Data System (ADS)

    Uchugonova, Aisada; Földes-Papp, Zeno; Kostner, Gerhard M.; König, Karsten

    2010-02-01

    The novel utrashort femtosecond laser scanning microscope FemtOgene (JenLab GmbH, Germany) with 12 femtoseconds at the focal plane have been employed in marker-free imaging and optical manipulation of stem cells as well as for the non-contact introduction of microRNA in cancer cells. Human adult pancreatic stem cells, salivary gland stem cells, and human dental pulp stem cells have been investigated by femtosecond laser multiphoton microscopy. Autofluorescence based on NAD(P)H and flavoproteins and second harmonic generation due to collagen have been imaged with submicron spatial resolution, 270 ps temporal resolution, and 10 nm spectral resolution. Major emission peaks at 460 nm and 530 nm with typical mean fluorescence lifetimes of 1.8 ns and 2.0 ns, respectively, were measured in a variety of stem cells using spectral imaging and time-correlated single photon counting. During differentiation, the ratios of bound to free NAD(P)H and NAD(P)H/flavoproteins changed. In addition, the biosynthesis of lipids and collagen was detected over a long period of time of up to 5 weeks. Nanoprocessing was performed with 12 femtosecond laser pulses and low picojoule pulse energies to realize targeted transfection and optical cleaning of human adult stem cell populations. Multiphoton sub-20fs microscopes may become novel non-invasive tools for marker-free optical stem cell characterization, for on-line monitoring of differentiation within a three-dimensional microenvironment, and for optical manipulation.

  12. Fusion of TTYH1 with the C19MC microRNA cluster drives expression of a brain-specific DNMT3B isoform in the embryonal brain tumor ETMR.

    PubMed

    Kleinman, Claudia L; Gerges, Noha; Papillon-Cavanagh, Simon; Sin-Chan, Patrick; Pramatarova, Albena; Quang, Dong-Anh Khuong; Adoue, Véronique; Busche, Stephan; Caron, Maxime; Djambazian, Haig; Bemmo, Amandine; Fontebasso, Adam M; Spence, Tara; Schwartzentruber, Jeremy; Albrecht, Steffen; Hauser, Peter; Garami, Miklos; Klekner, Almos; Bognar, Laszlo; Montes, Jose-Luis; Staffa, Alfredo; Montpetit, Alexandre; Berube, Pierre; Zakrzewska, Magdalena; Zakrzewski, Krzysztof; Liberski, Pawel P; Dong, Zhifeng; Siegel, Peter M; Duchaine, Thomas; Perotti, Christian; Fleming, Adam; Faury, Damien; Remke, Marc; Gallo, Marco; Dirks, Peter; Taylor, Michael D; Sladek, Robert; Pastinen, Tomi; Chan, Jennifer A; Huang, Annie; Majewski, Jacek; Jabado, Nada

    2014-01-01

    Embryonal tumors with multilayered rosettes (ETMRs) are rare, deadly pediatric brain tumors characterized by high-level amplification of the microRNA cluster C19MC. We performed integrated genetic and epigenetic analyses of 12 ETMR samples and identified, in all cases, C19MC fusions to TTYH1 driving expression of the microRNAs. ETMR tumors, cell lines and xenografts showed a specific DNA methylation pattern distinct from those of other tumors and normal tissues. We detected extreme overexpression of a previously uncharacterized isoform of DNMT3B originating at an alternative promoter that is active only in the first weeks of neural tube development. Transcriptional and immunohistochemical analyses suggest that C19MC-dependent DNMT3B deregulation is mediated by RBL2, a known repressor of DNMT3B. Transfection with individual C19MC microRNAs resulted in DNMT3B upregulation and RBL2 downregulation in cultured cells. Our data suggest a potential oncogenic re-engagement of an early developmental program in ETMR via epigenetic alteration mediated by an embryonic, brain-specific DNMT3B isoform. PMID:24316981

  13. microRNA and Wound Healing.

    PubMed

    Banerjee, Jaideep; Sen, Chandan K

    2015-01-01

    microRNAs (miRNAs) are small noncoding RNA molecules which play pivotal roles in wound healing. The increased expression of certain genes and expression of some others represent a key component of the wound biology and are largely under the regulation of naturally occurring miRNAs. Understanding the dysregulated miRNAs in chronic wound biology will therefore enable the development of newer therapies. This chapter focuses on the miRNAs that can be potentially targeted for improving skin wound healing and the challenges in miRNA therapy, including considerations in miRNA target identification and delivery. PMID:26663189

  14. Computational prediction of microRNA genes.

    PubMed

    Hertel, Jana; Langenberger, David; Stadler, Peter F

    2014-01-01

    The computational identification of novel microRNA (miRNA) genes is a challenging task in bioinformatics. Massive amounts of data describing unknown functional RNA transcripts have to be analyzed for putative miRNA candidates with automated computational pipelines. Beyond those miRNAs that meet the classical definition, high-throughput sequencing techniques have revealed additional miRNA-like molecules that are derived by alternative biogenesis pathways. Exhaustive bioinformatics analyses on such data involve statistical issues as well as precise sequence and structure inspection not only of the functional mature part but also of the whole precursor sequence of the putative miRNA. Apart from a considerable amount of species-specific miRNAs, the majority of all those genes are conserved at least among closely related organisms. Some miRNAs, however, can be traced back to very early points in the evolution of eukaryotic species. Thus, the investigation of the conservation of newly found miRNA candidates comprises an important step in the computational annotation of miRNAs.Topics covered in this chapter include a review on the obvious problem of miRNA annotation and family definition, recommended pipelines of computational miRNA annotation or detection, and an overview of current computer tools for the prediction of miRNAs and their limitations. The chapter closes discussing how those bioinformatic approaches address the problem of faithful miRNA prediction and correct annotation. PMID:24639171

  15. MicroRNA control of ovarian function

    PubMed Central

    Christenson, L. K.

    2011-01-01

    Post-transcriptional gene regulation, a regulatory mechanism classically involved in female and male germ cell function has recently been implicated in control of somatic cells of the ovary and testis. Recent advancements in this field may be attributed primarily to the discovery and study of microRNAs (miRNA), small RNA transcripts that can influence mRNA expression via post-transcriptional gene regulatory mechanisms. In the ovary, targeted deletion of Dicer 1, a key enzyme in miRNA biogenesis, provided the first empirical evidence that miRNA/siRNA were critically involved in multiple aspects of ovarian function (folliculogenesis, oocyte maturation, ovulation, and luteal function). Functional studies of miRNA in the ovary have mostly focused on granulosa cells during the critical period of the ovarian cycle surrounding the ovulatory surge of luteinizing hormone (LH). Specific miRNA have been implicated in ovarian responses, due to their transcriptional induction by the LH surge (i.e., miR-21, -132 and -212) or through bioinformatic approaches (miR-224, -17-5p and let-7b). Numerous other miRNA are highly abundant in ovarian somatic tissues, suggesting that we have much to discover with respect to the role of miRNA and regulation of ovarian function. This review will recap the key observations of these early studies and provide insight into future experiments that might further our understanding of ovarian function. PMID:21666774

  16. MicroRNA mimicry blocks pulmonary fibrosis

    PubMed Central

    Montgomery, Rusty L; Yu, Guoying; Latimer, Paul A; Stack, Christianna; Robinson, Kathryn; Dalby, Christina M; Kaminski, Naftali; van Rooij, Eva

    2014-01-01

    Over the last decade, great enthusiasm has evolved for microRNA (miRNA) therapeutics. Part of the excitement stems from the fact that a miRNA often regulates numerous related mRNAs. As such, modulation of a single miRNA allows for parallel regulation of multiple genes involved in a particular disease. While many studies have shown therapeutic efficacy using miRNA inhibitors, efforts to restore or increase the function of a miRNA have been lagging behind. The miR-29 family has gained a lot of attention for its clear function in tissue fibrosis. This fibroblast-enriched miRNA family is downregulated in fibrotic diseases which induces a coordinate increase of many extracellular matrix genes. Here, we show that intravenous injection of synthetic RNA duplexes can increase miR-29 levels in vivo for several days. Moreover, therapeutic delivery of these miR-29 mimics during bleomycin-induced pulmonary fibrosis restores endogenous miR-29 function whereby decreasing collagen expression and blocking and reversing pulmonary fibrosis. Our data support the feasibility of using miRNA mimics to therapeutically increase miRNAs and indicate miR-29 to be a potent therapeutic miRNA for treating pulmonary fibrosis. PMID:25239947

  17. MicroRNA profiling: approaches and considerations

    PubMed Central

    Pritchard, Colin C.; Cheng, Heather H.; Tewari, Muneesh

    2015-01-01

    MicroRNAs (miRNAs) are small RNAs (~22 nt long) that post-transcriptionally regulate the expression of thousands of genes in a broad range of organisms, in both normal physiologic and disease contexts. MiRNA expression profiling is gaining popularity because miRNAs, as key regulators in gene expression networks, can influence many biological processes and have also shown promise as biomarkers for disease. Technological advances have enabled the development of various platforms for miRNA profiling, and an understanding of the strengths and pitfalls of different approaches can aid in the effective use of miRNA profiling for diverse applications. We review here the major considerations for carrying out and interpreting results of miRNA profiling studies, as well as current and emerging applications of miRNA profiling. PMID:22510765

  18. MicroRNA fingerprints during human megakaryocytopoiesis

    PubMed Central

    Garzon, Ramiro; Pichiorri, Flavia; Palumbo, Tiziana; Iuliano, Rodolfo; Cimmino, Amelia; Aqeilan, Rami; Volinia, Stefano; Bhatt, Darshna; Alder, Hansjuerg; Marcucci, Guido; Calin, George A.; Liu, Chang-Gong; Bloomfield, Clara D.; Andreeff, Michael; Croce, Carlo M.

    2006-01-01

    microRNAs are a highly conserved class of noncoding RNAs with important regulatory functions in proliferation, apoptosis, development, and differentiation. To discover novel regulatory pathways during megakaryocytic differentiation, we performed microRNA expression profiling of in vitro-differentiated megakaryocytes derived from CD34+ hematopoietic progenitors. The main finding was down-regulation of miR-10a, miR-126, miR-106, miR-10b, miR-17 and miR-20. Hypothetically, the down-regulation of microRNAs unblocks target genes involved in differentiation. We confirmed in vitro and in vivo that miR-130a targets the transcription factor MAFB, which is involved in the activation of the GPIIB promoter, a key protein for platelet physiology. In addition, we found that miR-10a expression in differentiated megakaryocytes is inverse to that of HOXA1, and we showed that HOXA1 is a direct target of miR-10a. Finally, we compared the microRNA expression of megakaryoblastic leukemic cell lines with that of in vitro differentiated megakaryocytes and CD34+ progenitors. This analysis revealed up-regulation of miR-101, miR-126, miR-99a, miR-135, and miR-20. Our data delineate the expression of microRNAs during megakaryocytopoiesis and suggest a regulatory role of microRNAs in this process by targeting megakaryocytic transcription factors. PMID:16549775

  19. MicroRNA Methylation in Colorectal Cancer.

    PubMed

    Kaur, Sippy; Lotsari-Salomaa, Johanna E; Seppänen-Kaijansinkko, Riitta; Peltomäki, Päivi

    2016-01-01

    Epigenetic alterations such as DNA methylation, histone modifications and non-coding RNA (including microRNA) associated gene silencing have been identified as a major characteristic in human cancers. These alterations may occur more frequently than genetic mutations and play a key role in silencing tumor suppressor genes or activating oncogenes, thereby affecting multiple cellular processes. In recent years, studies have shown that microRNAs, that act as posttranscriptional regulators of gene expression are frequently deregulated in colorectal cancer (CRC), via aberrant DNA methylation. Over the past decade, technological advances have revolutionized the field of epigenetics and have led to the identification of numerous epigenetically dysregulated miRNAs in CRC, which are regulated by CpG island hypermethylation and DNA hypomethylation. In addition, aberrant DNA methylation of miRNA genes holds a great promise in several clinical applications such as biomarkers for early screening, prognosis, and therapeutic applications in CRC. PMID:27573897

  20. Potential Pitfalls in microRNA Profiling

    PubMed Central

    Chugh, Pauline; Dittmer, Dirk P.

    2013-01-01

    MicroRNAs (miRNAs) are small, noncoding RNAs that post-transcriptionally influence a wide range of cellular processes such as the host response to viral infection, innate immunity, cell cycle progression, migration and apoptosis through the inhibition of target mRNA translation. Due to the growing number of microRNAs and identification of their functional roles, miRNA profiling of many different sample types has become more expansive, especially with relevance to disease signatures. Here, we address some of the advantages and potential pitfalls of the currently available methods for miRNA expression profiling. Some of the topics discussed include isomiRNAs, comparison of different profiling platforms, normalization strategies and issues with regard to sample preparation and experimental analyses. PMID:22566380

  1. MicroRNA Expression in the Glaucomatous Retina

    PubMed Central

    Jayaram, Hari; Cepurna, William O.; Johnson, Elaine C.; Morrison, John C.

    2015-01-01

    Purpose MicroRNAs are small, endogenous noncoding RNAs that modulate posttranscriptional gene expression. Although the contribution of microRNAs to the pathogenesis of glaucomatous damage is unknown, supporting evidence from central nervous system (CNS) research suggests they may play a role. It was therefore hypothesized that microRNAs known to be altered in CNS injury are also altered in experimental glaucoma. Methods Intraocular pressure (IOP) was elevated in rats by unilateral injection of hypertonic saline and IOP monitored for 5 weeks. After rats were killed, retrobulbar optic nerve sections were graded for damage. MicroRNA was extracted from whole retinae of eyes with advanced nerve damage (n = 8) and from normal, noninjected control eyes (n = 8). Quantitative PCRs were performed using a panel of 17 microRNAs, reported from CNS research to be implicated in mechanisms also linked to glaucomatous damage. Computationally and experimentally derived gene targets were identified for the differentially expressed microRNAs. These were then integrated with existing gene array data. Functional interpretation was performed using the Molecular Signatures Database and DAVID Functional Annotation Clustering. Results Eight microRNAs were significantly downregulated in glaucomatous retinae compared with controls (miR-181c, miR-497, miR-204, let-7a, miR-29b, miR-16, miR106b, and miR-25); miR-27a was significantly upregulated. Enrichment of targets associated with extracellular matrix/cell proliferation, immune system, and regulation of apoptosis were observed. Cholesterol homeostasis and mTORC-1 pathways showed reduced expression. Conclusions MicroRNAs are differentially expressed in retinae of eyes with advanced glaucomatous damage compared with normal controls. Integrating microRNA with gene expression data may improve understanding of the complex biological responses produced by chronically elevated IOP. PMID:26720444

  2. MicroRNA-106b-25 cluster targets β-TRCP2, increases the expression of Snail and enhances cell migration and invasion in H1299 (non small cell lung cancer) cells

    SciTech Connect

    Savita, Udainiya; Karunagaran, Devarajan

    2013-05-17

    Highlights: •miR-106b-25 cluster directly targets the 3′UTR of the β-TRCP2 transcript. •β-TRCP2 mRNA was lower in H1299 cells stably expressing miR-106b-25 cluster. •miR-106b-25 cluster increased the expression of Snail. •miR-106b-25 cluster promoted the migration, colony formation and invasion. •miR-106b-25 cluster enhanced endothelial tube formation. -- Abstract: Lung cancer causes high mortality without a declining trend and non small cell lung cancer represents 85% of all pulmonary carcinomas. MicroRNAs (miRNAs) serve as fine regulators of proliferation, migration, invasion/metastasis and angiogenesis of normal and cancer cells. Using TargetScan6.2, we predicted that the ubiquitin ligase, β-TRCP2, could be a target for two of the constituent miRNAs of the miR-106b-25 cluster (miR-106b and miR-93). We generated a stable clone of miR-106b-25 cluster (CL) or the empty vector (EV) in H1299 (non small cell lung cancer) cells. The expression of β-TRCP2 mRNA was significantly lower in CL than that in EV cells. Transient expression of miR-93 but not antimiR-93 decreased the expression of β-TRCP2 mRNA in H1299 cells. β-TRCP2-3′UTR reporter assay revealed that its activity in CL cells was only 60% of that in EV cells. Snail protein expression was higher in CL than that in EV cells and H1299 cells exhibited an increase in the expression of Snail upon transient transfection with miR-93. miR-106b-25 cluster-induced migration of CL measured by scratch assay was more than that in EV cells and no significant difference in migration was observed between antimiR-93-transfected H1299 cells and the corresponding control-oligo-transfected cells. miR-106b-25 cluster-induced migration of CL cells was again confirmed in a Boyden chamber assay without the matrigel. CL cells were more invasive than EV cells when assessed using Boyden chambers with matrigel but there were no significant changes in the cell viabilities between EV and CL cells. Colony formation assay

  3. MicroRNA-143/-145 in Cardiovascular Diseases

    PubMed Central

    Zhao, Wang; Zhao, Shui-Ping; Zhao, Yu-Hong

    2015-01-01

    MicroRNAs (miRNAs) play an essential role in the onset and development of many cardiovascular diseases. Increasing evidence shows that miRNAs can be used as potential diagnostic biomarkers for cardiovascular diseases, and miRNA-based therapy may be a promising therapy for the treatment of cardiovascular diseases. The microRNA-143/-145 (miR-143/-145) cluster is essential for differentiation of vascular smooth muscle cells (VSMCs) and determines VSMC phenotypic switching. In this review, we summarize the recent progress in knowledge concerning the function of miR-143/-145 in the cardiovascular system and their role in cardiovascular diseases. We discuss the potential role of miR-143/-145 as valuable biomarkers for cardiovascular diseases and explore the potential strategy of targeting miR-143 and miR-145. PMID:26221598

  4. Profiling Pre-MicroRNA and Mature MicroRNA Expressions Using a Single Microarray and Avoiding Separate Sample Preparation

    PubMed Central

    Gan, Lin; Denecke, Bernd

    2013-01-01

    Mature microRNA is a crucial component in the gene expression regulation network. At the same time, microRNA gene expression and procession is regulated in a precise and collaborated way. Pre-microRNAs mediate products during the microRNA transcription process, they can provide hints of microRNA gene expression regulation or can serve as alternative biomarkers. To date, little effort has been devoted to pre-microRNA expression profiling. In this study, three human and three mouse microRNA profile data sets, based on the Affymetrix miRNA 2.0 array, have been re-analyzed for both mature and pre-microRNA signals as a primary test of parallel mature/pre-microRNA expression profiling on a single platform. The results not only demonstrated a glimpse of pre-microRNA expression in human and mouse, but also the relationship of microRNA expressions between pre- and mature forms. The study also showed a possible application of currently available microRNA microarrays in profiling pre-microRNA expression in a time and cost effective manner.

  5. MicroRNA dysregulation in multiple sclerosis

    PubMed Central

    Jr, Omar de Faria; Moore, Craig S.; Kennedy, Timothy E.; Antel, Jack P.; Bar-Or, Amit; Dhaunchak, Ajit S.

    2013-01-01

    Multiple sclerosis (MS) is a chronic inflammatory disease characterized by central nervous system (CNS) demyelination and axonal degeneration. Although the cause of MS is still unknown, it is widely accepted that novel drug targets need to focus on both decreasing inflammation and promoting CNS repair. In MS and experimental autoimmune encephalomyelitis, non-coding small microRNAs (miRNAs) are dysregulated in the immune system and CNS. Since individual miRNAs are able to down-regulate multiple targeted mRNA transcripts, even minor changes in miRNA expression may lead to significant alterations in gene expression. Herein, we review miRNA signatures reported in CNS tissue and immune cells of MS patients and consider how altered miRNA expression may influence MS pathology. PMID:23346094

  6. MicroRNA-106b-25 cluster targets β-TRCP2, increases the expression of Snail and enhances cell migration and invasion in H1299 (non small cell lung cancer) cells.

    PubMed

    Savita, Udainiya; Karunagaran, Devarajan

    2013-05-17

    Lung cancer causes high mortality without a declining trend and non small cell lung cancer represents 85% of all pulmonary carcinomas. MicroRNAs (miRNAs) serve as fine regulators of proliferation, migration, invasion/metastasis and angiogenesis of normal and cancer cells. Using TargetScan6.2, we predicted that the ubiquitin ligase, β-TRCP2, could be a target for two of the constituent miRNAs of the miR-106b-25 cluster (miR-106b and miR-93). We generated a stable clone of miR-106b-25 cluster (CL) or the empty vector (EV) in H1299 (non small cell lung cancer) cells. The expression of β-TRCP2 mRNA was significantly lower in CL than that in EV cells. Transient expression of miR-93 but not antimiR-93 decreased the expression of β-TRCP2 mRNA in H1299 cells. β-TRCP2-3'UTR reporter assay revealed that its activity in CL cells was only 60% of that in EV cells. Snail protein expression was higher in CL than that in EV cells and H1299 cells exhibited an increase in the expression of Snail upon transient transfection with miR-93. miR-106b-25 cluster-induced migration of CL measured by scratch assay was more than that in EV cells and no significant difference in migration was observed between antimiR-93-transfected H1299 cells and the corresponding control-oligo-transfected cells. miR-106b-25 cluster-induced migration of CL cells was again confirmed in a Boyden chamber assay without the matrigel. CL cells were more invasive than EV cells when assessed using Boyden chambers with matrigel but there were no significant changes in the cell viabilities between EV and CL cells. Colony formation assay revealed that the CL cells formed more number of colonies than EV cells but they were smaller in size than those formed by EV cells. The supernatant from CL cells was more effective than that from EV cells in inducing tube formation in endothelial cells. Taken together, our data indicate that miR-106b-25 cluster may play an important role in the metastasis of human non-small cell

  7. MicroRNA Dysregulation in Cystic Fibrosis

    PubMed Central

    McKiernan, Paul J.; Greene, Catherine M.

    2015-01-01

    The cystic fibrosis lung is a complex milieu comprising multiple factors that coordinate its physiology. MicroRNAs are regulatory factors involved in most biological processes and it is becoming increasingly clear that they play a key role in the development and manifestations of CF lung disease. These small noncoding RNAs act posttranscriptionally to inhibit protein production. Their involvement in the pathogenesis of CF lung disease stems from the fact that their expression is altered in vivo in the CF lung due to intrinsic and extrinsic factors; to date defective chloride ion conductance, endoplasmic reticulum stress, inflammation, and infection have been implicated in altering endogenous miRNA expression in this setting. Here, the current state-of-the-art and biological consequences of altered microRNA expression in cystic fibrosis are reviewed. PMID:26185362

  8. Identifying survival-associated ceRNA clusters in cholangiocarcinoma.

    PubMed

    Wan, Ming; Zhang, Fu-Min; Li, Zheng-Long; Kang, Peng-Cheng; Jiang, Ping-Ming; Wang, Yi-Min; Wang, Zhi-Dong; Zhong, Xiang-Yu; Li, Chun-Long; Wang, Hao; Zhao, Shi-Yong; Cui, Yun-Fu

    2016-09-01

    Competing endogenous RNAs (ceRNAs) represent a novel layer regulations of long non-coding RNAs (lncRNAs) and genes that play important roles in cancer pathogenesis by binding microRNAs (miRNAs). However, the competition mechanism of ceRNAs in cholangiocarcinoma (CHOL) is not fully understood. In this study, we constructed a dysregulated ceRNA competitive network (CCEN) to globally characterize the competing difference between CHOL and normal tissues. Then, we integrated affinity propagation and Kaplan‑Meier (K-M) methods to identify functional clusters associated with survival. A total of 7 key ceRNA clusters were identified. Further functional annotation analyses found that Cluster23 and Cluster32 involved cell based functions, and the loss of ceRNA competitive relations in clusters may contribute to CHOL, by disturbing important biological processes, such as 'Pathway in cancer', MAPK and Neurotrophin signaling pathway. This study provides further insights into understanding the competitive mechanism of ceRNAs in CHOL. PMID:27432084

  9. An alternative mode of microRNA target recognition

    PubMed Central

    Chi, Sung Wook; Hannon, Gregory J.; Darnell, Robert B.

    2012-01-01

    MicroRNAs (miRNAs) regulate mRNA targets through perfect pairing with their seed region (position 2-7). Recently, a precise genome-wide map of miRNA interaction sites in mouse brain was generated by high-throughput sequencing of clusters of ~50 nucleotide RNA tags associated with Argonaute (Ago HITS-CLIP). By analyzing Ago HITS-CLIP “orphan clusters” – Ago binding regions from HITS-CLIP that cannot be explained by canonical seed matches – we have identified an alternative binding mode used by miRNAs. Specifically, G-bulge sites (position 5-6) are often bound and regulated by miR-124 in brain. More generally, bulged sites comprise ≥ 15% (≥ 1441 sites) of all Ago-miRNA interactions in mouse brain and are evolutionally conserved. We have termed position 6 the “pivot” nucleotide and suggest a model in which a transitional “nucleation-bulge” leads to functional bulge mRNA-miRNA interactions, expanding the number of potential miRNA regulatory sites. PMID:22343717

  10. Cotranslational microRNA mediated messenger RNA destabilization

    PubMed Central

    Tat, Trinh To; Maroney, Patricia A; Chamnongpol, Sangpen; Coller, Jeff; Nilsen, Timothy W

    2016-01-01

    MicroRNAs are small (22 nucleotide) regulatory molecules that play important roles in a wide variety of biological processes. These RNAs, which bind to targeted mRNAs via limited base pairing interactions, act to reduce protein production from those mRNAs. Considerable evidence indicates that miRNAs destabilize targeted mRNAs by recruiting enzymes that function in normal mRNA decay and mRNA degradation is widely thought to occur when mRNAs are in a ribosome free state. Nevertheless, when examined, miRNA targeted mRNAs are invariably found to be polysome associated; observations that appear to be at face value incompatible with a simple decay model. Here, we provide evidence that turnover of miRNA-targeted mRNAs occurs while they are being translated. Cotranslational mRNA degradation is initiated by decapping and proceeds 5’ to 3’ behind the last translating ribosome. These results provide an explanation for a long standing mystery in the miRNA field. DOI: http://dx.doi.org/10.7554/eLife.12880.001 PMID:27058298

  11. Bioinformatics Resources for MicroRNA Discovery

    PubMed Central

    Moore, Alyssa C.; Winkjer, Jonathan S.; Tseng, Tsai-Tien

    2015-01-01

    Biomarker identification is often associated with the diagnosis and evaluation of various diseases. Recently, the role of microRNA (miRNA) has been implicated in the development of diseases, particularly cancer. With the advent of next-generation sequencing, the amount of data on miRNA has increased tremendously in the last decade, requiring new bioinformatics approaches for processing and storing new information. New strategies have been developed in mining these sequencing datasets to allow better understanding toward the actions of miRNAs. As a result, many databases have also been established to disseminate these findings. This review focuses on several curated databases of miRNAs and their targets from both predicted and validated sources. PMID:26819547

  12. MicroRNA Processing and Human Cancer

    PubMed Central

    Ohtsuka, Masahisa; Ling, Hui; Doki, Yuichiro; Mori, Masaki; Calin, George Adrian

    2015-01-01

    MicroRNAs (miRNAs) are short non-coding RNAs of 20 to 25 nucleotides that regulate gene expression post-transcriptionally mainly by binding to a specific sequence of the 3′ end of the untranslated region (3′UTR) of target genes. Since the first report on the clinical relevance of miRNAs in cancer, many miRNAs have been demonstrated to act as oncogenes, whereas others function as tumor suppressors. Furthermore, global miRNA dysregulation, due to alterations in miRNA processing factors, has been observed in a large variety of human cancer types. As previous studies have shown, the sequential miRNA processing can be divided into three steps: processing by RNAse in the nucleus; transportation by Exportin-5 (XPO5) from the nucleus; and processing by the RNA-induced silencing complex (RISC) in the cytoplasm. Alteration in miRNA processing genes, by genomic mutations, aberrant expression or other means, could significantly affect cancer initiation, progression and metastasis. In this review, we focus on the biogenesis of miRNAs with emphasis on the potential of miRNA processing factors in human cancers. PMID:26308063

  13. miSolRNA: A tomato micro RNA relational database

    PubMed Central

    2010-01-01

    Background The economic importance of Solanaceae plant species is well documented and tomato has become a model for functional genomics studies. In plants, important processes are regulated by microRNAs (miRNA). Description We describe here a data base integrating genetic map positions of miRNA-targeted genes, their expression profiles and their relations with quantitative fruit metabolic loci and yield associated traits. miSolRNA provides a metadata source to facilitate the construction of hypothesis aimed at defining physiological modes of action of regulatory process underlying the metabolism of the tomato fruit. Conclusions The MiSolRNA database allows the simple extraction of metadata for the proposal of new hypothesis concerning possible roles of miRNAs in the regulation of tomato fruit metabolism. It permits i) to map miRNAs and their predicted target sites both on expressed (SGN-UNIGENES) and newly annotated sequences (BAC sequences released), ii) to co-locate any predicted miRNA-target interaction with metabolic QTL found in tomato fruits, iii) to retrieve expression data of target genes in tomato fruit along their developmental period and iv) to design further experiments for unresolved questions in complex trait biology based on the use of genetic materials that have been proven to be a useful tools for map-based cloning experiments in Solanaceae plant species. PMID:21059227

  14. MicroRNA targets in Drosophila

    PubMed Central

    Enright, Anton J; John, Bino; Gaul, Ulrike; Tuschl, Thomas; Sander, Chris; Marks, Debora S

    2004-01-01

    Background The recent discoveries of microRNA (miRNA) genes and characterization of the first few target genes regulated by miRNAs in Caenorhabditis elegans and Drosophila melanogaster have set the stage for elucidation of a novel network of regulatory control. We present a computational method for whole-genome prediction of miRNA target genes. The method is validated using known examples. For each miRNA, target genes are selected on the basis of three properties: sequence complementarity using a position-weighted local alignment algorithm, free energies of RNA-RNA duplexes, and conservation of target sites in related genomes. Application to the D. melanogaster, Drosophila pseudoobscura and Anopheles gambiae genomes identifies several hundred target genes potentially regulated by one or more known miRNAs. Results These potential targets are rich in genes that are expressed at specific developmental stages and that are involved in cell fate specification, morphogenesis and the coordination of developmental processes, as well as genes that are active in the mature nervous system. High-ranking target genes are enriched in transcription factors two-fold and include genes already known to be under translational regulation. Our results reaffirm the thesis that miRNAs have an important role in establishing the complex spatial and temporal patterns of gene activity necessary for the orderly progression of development and suggest additional roles in the function of the mature organism. In addition the results point the way to directed experiments to determine miRNA functions. Conclusions The emerging combinatorics of miRNA target sites in the 3' untranslated regions of messenger RNAs are reminiscent of transcriptional regulation in promoter regions of DNA, with both one-to-many and many-to-one relationships between regulator and target. Typically, more than one miRNA regulates one message, indicative of cooperative translational control. Conversely, one miRNA may have

  15. Estrogen Regulation of MicroRNA Expression

    PubMed Central

    Klinge, Carolyn M

    2009-01-01

    Women outlive men, but life expectancy is not influenced by hormone replacement (estrogen + progestin) therapy. Estrogens appear to protect brain, cardiovascular tissues, and bone from aging. Estrogens regulate genes directly through binding to estrogen receptors alpha and beta (ERα and ERβ) that are ligand-activated transcription factors and indirectly by activating plasma membrane-associated ER which, in turns, activates intracellular signaling cascades leading to altered gene expression. MicroRNAs (miRNAs) are short (19-25 nucleotides), naturally-occurring, non-coding RNA molecules that base-pair with the 3’ untranslated region of target mRNAs. This interaction either blocks translation of the mRNA or targets the mRNA transcript to be degraded. The human genome contains ~ 700-1,200 miRNAs. Aberrant patterns of miRNA expression are implicated in human diseases including breast cancer. Recent studies have identified miRNAs regulated by estrogens in human breast cancer cells, human endometrial stromal and myometrial smooth muscle cells, rat mammary gland, and mouse uterus. The decline of estradiol levels in postmenopausal women has been implicated in various age-associated disorders. The role of estrogen-regulated miRNA expression, the target genes of these miRNAs, and the role of miRNAs in aging has yet to be explored. PMID:19881910

  16. Bioinformatic tools for microRNA dissection

    PubMed Central

    Akhtar, Most Mauluda; Micolucci, Luigina; Islam, Md Soriful; Olivieri, Fabiola; Procopio, Antonio Domenico

    2016-01-01

    Recently, microRNAs (miRNAs) have emerged as important elements of gene regulatory networks. MiRNAs are endogenous single-stranded non-coding RNAs (∼22-nt long) that regulate gene expression at the post-transcriptional level. Through pairing with mRNA, miRNAs can down-regulate gene expression by inhibiting translation or stimulating mRNA degradation. In some cases they can also up-regulate the expression of a target gene. MiRNAs influence a variety of cellular pathways that range from development to carcinogenesis. The involvement of miRNAs in several human diseases, particularly cancer, makes them potential diagnostic and prognostic biomarkers. Recent technological advances, especially high-throughput sequencing, have led to an exponential growth in the generation of miRNA-related data. A number of bioinformatic tools and databases have been devised to manage this growing body of data. We analyze 129 miRNA tools that are being used in diverse areas of miRNA research, to assist investigators in choosing the most appropriate tools for their needs. PMID:26578605

  17. MicroRNA-132, -134, and -138: a microRNA troika rules in neuronal dendrites.

    PubMed

    Bicker, Silvia; Lackinger, Martin; Weiß, Kerstin; Schratt, Gerhard

    2014-10-01

    Dendritic mRNA transport and local translation in the postsynaptic compartment play an important role in synaptic plasticity, learning and memory. Local protein synthesis at the synapse has to be precisely orchestrated by a plethora of factors including RNA binding proteins as well as microRNAs, an extensive class of small non-coding RNAs. By binding to complementary sequences in target mRNAs, microRNAs fine-tune protein synthesis and thereby represent critical regulators of gene expression at the post-transcriptional level. Research over the last years identified an entire network of dendritic microRNAs that fulfills an essential role in synapse development and physiology. Recent studies provide evidence that these small regulatory molecules are highly regulated themselves, at the level of expression as well as function. The importance of microRNAs for correct function of the nervous system is reflected by an increasing number of studies linking dysregulation of microRNA pathways to neurological disorders. By focusing on three extensively studied examples (miR-132, miR-134, miR-138), this review will attempt to illustrate the complex regulatory roles of dendritic microRNAs at the synapse and their implications for pathological conditions. PMID:25008044

  18. Genetic variants in microRNA genes: impact on microRNA expression, function, and disease

    PubMed Central

    Cammaerts, Sophia; Strazisar, Mojca; De Rijk, Peter; Del Favero, Jurgen

    2015-01-01

    MicroRNAs (miRNAs) are important regulators of gene expression and like any other gene, their coding sequences are subject to genetic variation. Variants in miRNA genes can have profound effects on miRNA functionality at all levels, including miRNA transcription, maturation, and target specificity, and as such they can also contribute to disease. The impact of variants in miRNA genes is the focus of the present review. To put these effects into context, we first discuss the requirements of miRNA transcripts for maturation. In the last part an overview of available databases and tools and experimental approaches to investigate miRNA variants related to human disease is presented. PMID:26052338

  19. Dysregulated microRNA Clusters in Response to Retinoic Acid and CYP26B1 Inhibitor Induced Testicular Function in Dogs

    PubMed Central

    Kasimanickam, Vanmathy R.; Kasimanickam, Ramanathan K.; Dernell, William S.

    2014-01-01

    Spermatogenesis is a multistep synchronized process. Diploid spermatogonia differentiate into haploid spermatozoa following mitosis, meiosis and spermiogenesis. Division and differentiation of male germ cells is achieved through the sequential expression of several genes. Numerous mRNAs in the differentiating germ cells undergo post-transcriptional and translational regulation. MiRNAs are powerful negative regulators of mRNA transcription, stability, and translation and recognize their mRNA targets through base-pairing. Retinoic acid (RA) signaling is essential for spermatogenesis and testicular function. Testicular RA level is critical for RA signal transduction. This study investigated the miRNAs modulation in an RA- induced testicular environment following the administration of all-trans RA (2 µM) and CYP26B1- inhibitor (1 µM) compared to control. Eighty four canine mature miRNAs were analyzed and their expression signatures were distinguished using real-time PCR based array technology. Of the miRNAs analyzed, miRNA families such as miR-200 (cfa-miR-200a, cfa-miR-200b and cfa-miR-200c), Mirlet-7 (cfa-let-7a, cfa-let-7b, cfa-let-7c, cfa-let-7g and cfa-let-7f), miR-125 (cfa-miR-125a and cfa-miR-125b), miR-146 (cfa-miR-146a and cfa-miR-146b), miR-34 (cfa-miR-34a, cfa-miR-34b and cfa-miR-34c), miR-23 (cfa-miR-23a and cfa-miR-23b), cfa-miR-184, cfa-miR-214 and cfa-miR-141 were significantly up-regulated with testicular RA intervention via administration of CYP26B1 inhibitor and all-trans-RA and species of miRNA such as cfa-miR-19a, cfa-miR-29b, cfa-miR-29c, cfa-miR-101 and cfa-miR-137 were significantly down-regulated. This study explored information regarding chromosome distribution, human orthologous sequences and the interaction of target genes of miRNA families significantly distinguished in this study using prediction algorithms. This study importantly identified dysregulated miRNA species resulting from RA-induced spermatogenesis. The present contribution

  20. Evolution of Arabidopsis microRNA families through duplication events

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recently there has been a great interest in the identification of microRNAs and their targets as well as understanding the spatial and temporal regulation of microRNA genes. To understand how microRNA genes evolve, we looked at several rapidly evolving families in Arabidopsis thaliana, and found th...

  1. Mammalian 5′-capped microRNA precursors that generate a single microRNA

    PubMed Central

    Xie, Mingyi; Li, Mingfeng; Vilborg, Anna; Lee, Nara; Shu, Mei-Di; Yartseva, Valeria; Šestan, Nenad; Steitz, Joan A.

    2014-01-01

    Summary MicroRNAs (miRNAs) are short RNA gene regulators typically produced from primary transcripts that are cleaved by the nuclear Microprocessor complex, with the resulting precursor miRNA hairpins exported by Exportin-5 and processed by cytoplasmic Dicer to yield two (5p- and 3p-) miRNAs. Here, we document Microprocessor-independent 7-methylguanosine (m7G) capped pre-miRNAs, whose 5′ ends coincide with transcription start sites, while the 3′ ends are most likely generated by transcription termination. By establishing a small RNA Cap-seq method that employs the cap-binding protein eIF4E, we identified a group of murine m7G-capped pre-miRNAs genome-wide. The m7G-capped pre-miRNAs are exported via the PHAX-Exportin-1 pathway. After Dicer cleavage, only the 3p-miRNA is efficiently loaded onto Argonaute to form a functional microRNP. This unusual miRNA biogenesis pathway, which differs in pre-miRNA synthesis, nuclear-cytoplasmic transport and guide strand selection, enables the development of shRNA expression constructs that produce a single 3p-siRNA. PMID:24360278

  2. A probabilistic approach to microRNA-target binding

    SciTech Connect

    Ogul, Hasan; Umu, Sinan U.; Tuncel, Y. Yener; Akkaya, Mahinur S.

    2011-09-16

    Highlights: {yields} A new probabilistic model is introduced for microRNA-target binding. {yields} The new model significantly outperforms RNAHybrid and miRTif. {yields} The experiments can unveil the effects of the type and directions of distinct base pairings. -- Abstract: Elucidation of microRNA activity is a crucial step in understanding gene regulation. One key problem in this effort is how to model the pairwise interactions of microRNAs with their targets. As this interaction is strongly mediated by their sequences, it is desired to set-up a probabilistic model to explain the binding preferences between a microRNA sequence and the sequence of a putative target. To this end, we introduce a new model of microRNA-target binding, which transforms an aligned duplex to a new sequence and defines the likelihood of this sequence using a Variable Length Markov Chain. It offers a complementary representation of microRNA-mRNA pairs for microRNA target prediction tools or other probabilistic frameworks of integrative gene regulation analysis. The performance of present model is evaluated by its ability to predict microRNA-target mRNA interaction given a mature microRNA sequence and a putative mRNA binding site. In regard to classification accuracy, it outperforms two recent methods based on thermodynamic stability and sequence complementarity. The experiments can also unveil the effects of base pairing types and non-seed region in duplex formation.

  3. Modules of human micro-RNA co-target network

    NASA Astrophysics Data System (ADS)

    Basu, Mahashweta; Bhattacharyya, Nitai P.; Mohanty, P. K.

    2011-05-01

    Human micro RNAs (miRNAs) target about 90% of the coding genes and form a complex regulatory network. We study the community structure of the miRNA co-target network considering miRNAs as the nodes which are connected by weighted links. The weight of link that connects a pair of miRNAs denote the total number of common transcripts targeted by that pair. We argue that the network consists of about 74 modules, quite similar to the components (or clusters) obtained earlier [Online J Bioinformatics, 10,280], indicating that the components of the miRNA co-target network are self organized in a way to maximize the modularity.

  4. MicroRNA in United Airway Diseases

    PubMed Central

    Liu, Zheng; Zhang, Xin-Hao; Callejas-Díaz, Borja; Mullol, Joaquim

    2016-01-01

    The concept of united airway diseases (UAD) has received increasing attention in recent years. Sustained and increased inflammation is a common feature of UAD, which is inevitably accompanied with marked gene modification and tight gene regulation. However, gene regulation in the common inflammatory processes in UAD remains unclear. MicroRNA (miRNA), a novel regulator of gene expression, has been considered to be involved in many inflammatory diseases. Although there are an increasing number of studies of miRNAs in inflammatory upper and lower airway diseases, few miRNAs have been identified that directly link the upper and lower airways. In this article, therefore, we reviewed the relevant studies available in order to improve the understanding of the roles of miRNAs in the interaction and pathogenesis of UAD. PMID:27187364

  5. MicroRNA in United Airway Diseases.

    PubMed

    Liu, Zheng; Zhang, Xin-Hao; Callejas-Díaz, Borja; Mullol, Joaquim

    2016-01-01

    The concept of united airway diseases (UAD) has received increasing attention in recent years. Sustained and increased inflammation is a common feature of UAD, which is inevitably accompanied with marked gene modification and tight gene regulation. However, gene regulation in the common inflammatory processes in UAD remains unclear. MicroRNA (miRNA), a novel regulator of gene expression, has been considered to be involved in many inflammatory diseases. Although there are an increasing number of studies of miRNAs in inflammatory upper and lower airway diseases, few miRNAs have been identified that directly link the upper and lower airways. In this article, therefore, we reviewed the relevant studies available in order to improve the understanding of the roles of miRNAs in the interaction and pathogenesis of UAD. PMID:27187364

  6. MicroRNA signatures in liver diseases

    PubMed Central

    Chen, Xian-Ming

    2009-01-01

    MicroRNAs (miRNAs) are an emerging class of highly conserved non-coding small RNAs that regulate gene expression at the post-transcriptional level. It is now clear that miRNAs can potentially regulate every aspect of cellular activity, including differentiation and development, metabolism, proliferation, apoptotic cell death, viral infection and tumorigenesis. Recent studies provide clear evidence that miRNAs are abundant in the liver and modulate a diverse spectrum of liver functions. Deregulation of miRNA expression may be a key pathogenetic factor in many liver diseases including viral hepatitis, hepatocellular cancer and polycystic liver diseases. A clearer understanding of the mechanisms involved in miRNA deregulation will offer new diagnostic and therapeutic strategies to treat liver diseases. Moreover, better understanding of miRNA regulation and identification of tissue-specific miRNA targets employing transgenic/knockout models and/or modulating oligonucleotides will improve our knowledge of liver physiology and diseases. PMID:19360909

  7. MicroRNA in Teleost Fish

    PubMed Central

    Bizuayehu, Teshome Tilahun; Babiak, Igor

    2014-01-01

    MicroRNAs (miRNAs) are transcriptional and posttranscriptional regulators involved in nearly all known biological processes in distant eukaryotic clades. Their discovery and functional characterization have broadened our understanding of biological regulatory mechanisms in animals and plants. They show both evolutionary conserved and unique features across Metazoa. Here, we present the current status of the knowledge about the role of miRNA in development, growth, and physiology of teleost fishes, in comparison to other vertebrates. Infraclass Teleostei is the most abundant group among vertebrate lineage. Fish are an important component of aquatic ecosystems and human life, being the prolific source of animal proteins worldwide and a vertebrate model for biomedical research. We review miRNA biogenesis, regulation, modifications, and mechanisms of action. Specific sections are devoted to the role of miRNA in teleost development, organogenesis, tissue differentiation, growth, regeneration, reproduction, endocrine system, and responses to environmental stimuli. Each section discusses gaps in the current knowledge and pinpoints the future directions of research on miRNA in teleosts. PMID:25053657

  8. Staufen Negatively Modulates MicroRNA Activity in Caenorhabditis elegans

    PubMed Central

    Ren, Zhiji; Veksler-Lublinsky, Isana; Morrissey, David; Ambros, Victor

    2016-01-01

    The double-stranded RNA-binding protein Staufen has been implicated in various posttranscriptional gene regulatory processes. Here, we demonstrate that the Caenorhabditis elegans homolog of Staufen, STAU-1, functionally interacts with microRNAs. Loss-of-function mutations of stau-1 significantly suppress phenotypes of let-7 family microRNA mutants, a hypomorphic allele of dicer, and a lsy-6 microRNA partial loss-of-function mutant. Furthermore, STAU-1 modulates the activity of lin-14, a target of lin-4 and let-7 family microRNAs, and this modulation is abolished when the 3′ untranslated region of lin-14 is removed. Deep sequencing of small RNA cDNA libraries reveals no dramatic change in the levels of microRNAs or other small RNA populations between wild-type and stau-1 mutants, with the exception of certain endogenous siRNAs in the WAGO pathway. The modulation of microRNA activity by STAU-1 does not seem to be associated with the previously reported enhanced exogenous RNAi (Eri) phenotype of stau-1 mutants, since eri-1 exhibits the opposite effect on microRNA activity. Altogether, our results suggest that STAU-1 negatively modulates microRNA activity downstream of microRNA biogenesis, possibly by competing with microRNAs for binding on the 3′ untranslated region of target mRNAs. PMID:26921297

  9. Staufen Negatively Modulates MicroRNA Activity in Caenorhabditis elegans.

    PubMed

    Ren, Zhiji; Veksler-Lublinsky, Isana; Morrissey, David; Ambros, Victor

    2016-01-01

    The double-stranded RNA-binding protein Staufen has been implicated in various posttranscriptional gene regulatory processes. Here, we demonstrate that the Caenorhabditis elegans homolog of Staufen, STAU-1, functionally interacts with microRNAs. Loss-of-function mutations of stau-1 significantly suppress phenotypes of let-7 family microRNA mutants, a hypomorphic allele of dicer, and a lsy-6 microRNA partial loss-of-function mutant. Furthermore, STAU-1 modulates the activity of lin-14, a target of lin-4 and let-7 family microRNAs, and this modulation is abolished when the 3' untranslated region of lin-14 is removed. Deep sequencing of small RNA cDNA libraries reveals no dramatic change in the levels of microRNAs or other small RNA populations between wild-type and stau-1 mutants, with the exception of certain endogenous siRNAs in the WAGO pathway. The modulation of microRNA activity by STAU-1 does not seem to be associated with the previously reported enhanced exogenous RNAi (Eri) phenotype of stau-1 mutants, since eri-1 exhibits the opposite effect on microRNA activity. Altogether, our results suggest that STAU-1 negatively modulates microRNA activity downstream of microRNA biogenesis, possibly by competing with microRNAs for binding on the 3' untranslated region of target mRNAs. PMID:26921297

  10. Multifaceted enrichment analysis of RNA-RNA crosstalk reveals cooperating micro-societies in human colorectal cancer.

    PubMed

    Mazza, Tommaso; Mazzoccoli, Gianluigi; Fusilli, Caterina; Capocefalo, Daniele; Panza, Anna; Biagini, Tommaso; Castellana, Stefano; Gentile, Annamaria; De Cata, Angelo; Palumbo, Orazio; Stallone, Raffaella; Rubino, Rosa; Carella, Massimo; Piepoli, Ada

    2016-05-19

    Alterations in the balance of mRNA and microRNA (miRNA) expression profiles contribute to the onset and development of colorectal cancer. The regulatory functions of individual miRNA-gene pairs are widely acknowledged, but group effects are largely unexplored. We performed an integrative analysis of mRNA-miRNA and miRNA-miRNA interactions using high-throughput mRNA and miRNA expression profiles obtained from matched specimens of human colorectal cancer tissue and adjacent non-tumorous mucosa. This investigation resulted in a hypernetwork-based model, whose functional backbone was fulfilled by tight micro-societies of miRNAs. These proved to modulate several genes that are known to control a set of significantly enriched cancer-enhancer and cancer-protection biological processes, and that an array of upstream regulatory analyses demonstrated to be dependent on miR-145, a cell cycle and MAPK signaling cascade master regulator. In conclusion, we reveal miRNA-gene clusters and gene families with close functional relationships and highlight the role of miR-145 as potent upstream regulator of a complex RNA-RNA crosstalk, which mechanistically modulates several signaling pathways and regulatory circuits that when deranged are relevant to the changes occurring in colorectal carcinogenesis. PMID:27067546

  11. MicroRNA Therapeutics: the Next Magic Bullet?

    PubMed Central

    Simonson, Bridget; Das, Saumya

    2015-01-01

    MicroRNAs are short noncoding 18–25 nucleotide long RNA which bind and inhibit mRNA. Currently, there are over 1000 known human microRNAs, and microRNAs control over 50% of mammalian protein coding genes. MicroRNAs can be overexpressed or repressed in different diseases and inhibition or replacement of microRNAs is a promising area of study for therapeutics. Here we review the current knowledge of microRNA therapy, and discuss ways in which they can be utilized. We also discuss different methods of delivery of miRNA, and current clinical trials of microRNA-based therapies for disease. Finally we discuss the current limitations in the field, and how these limitations are being overcome. PMID:25807941

  12. Placental microRNA expression in pregnancies complicated by preeclampsia

    PubMed Central

    Enquobahrie, Daniel A.; Abetew, Dejene F.; Sorensen, Tanya K.; Willoughby, David; Chidambaram, Kumaravel; Williams, Michelle A.

    2010-01-01

    Objective The role of post-transcription regulation in preeclampsia is largely unknown. We investigated preeclampsia related placental microRNA (miRNA) expression using microarray and confirmatory qRT-PCR experiments. Study design Placental expressions of characterized and novel miRNAs (1,295 probes) were measured in samples collected from 20 preeclampsia cases and 20 controls. Differential expression was evaluated using Students T-test and fold change analyses. In pathway analysis, we examined functions/functional relationships of targets of differentially expressed miRNAs. Results Eight miRNAs were differentially expressed (1 up- and 7 down-regulated) among preeclampsia cases compared with controls. These included previously identified candidates (miR-210, miR-1 and a miRNA in the 14q32.31 cluster region) and others that are novel (miR- 584 and miR-34c-5p). These miRNAs target genes that participate in organ/system development (cardiovascular and reproductive system), immunologic dysfunction, cell adhesion, cell cycle and signaling. Conclusion Expression of microRNAs that target genes in diverse pathophysiological processes is altered in the setting of preeclampsia. PMID:21093846

  13. PARma: identification of microRNA target sites in AGO-PAR-CLIP data.

    PubMed

    Erhard, Florian; Dölken, Lars; Jaskiewicz, Lukasz; Zimmer, Ralf

    2013-01-01

    PARma is a complete data analysis software for AGO-PAR-CLIP experiments to identify target sites of microRNAs as well as the microRNA binding to these sites. It integrates specific characteristics of the experiments into a generative model. The model and a novel pattern discovery tool are iteratively applied to data to estimate seed activity probabilities, cluster confidence scores and to assign the most probable microRNA. Based on differential PAR-CLIP analysis and comparison to RIP-Chip data, we show that PARma is more accurate than existing approaches. PARma is available from http://www.bio.ifi.lmu.de/PARma. PMID:23895117

  14. PARma: identification of microRNA target sites in AGO-PAR-CLIP data

    PubMed Central

    2013-01-01

    PARma is a complete data analysis software for AGO-PAR-CLIP experiments to identify target sites of microRNAs as well as the microRNA binding to these sites. It integrates specific characteristics of the experiments into a generative model. The model and a novel pattern discovery tool are iteratively applied to data to estimate seed activity probabilities, cluster confidence scores and to assign the most probable microRNA. Based on differential PAR-CLIP analysis and comparison to RIP-Chip data, we show that PARma is more accurate than existing approaches. PARma is available from http://www.bio.ifi.lmu.de/PARma PMID:23895117

  15. MicroRNA expression profiling reveals miRNA families regulating specific biological pathways in mouse frontal cortex and hippocampus.

    PubMed

    Juhila, Juuso; Sipilä, Tessa; Icay, Katherine; Nicorici, Daniel; Ellonen, Pekka; Kallio, Aleksi; Korpelainen, Eija; Greco, Dario; Hovatta, Iiris

    2011-01-01

    MicroRNAs (miRNAs) are small regulatory molecules that cause post-transcriptional gene silencing. Although some miRNAs are known to have region-specific expression patterns in the adult brain, the functional consequences of the region-specificity to the gene regulatory networks of the brain nuclei are not clear. Therefore, we studied miRNA expression patterns by miRNA-Seq and microarrays in two brain regions, frontal cortex (FCx) and hippocampus (HP), which have separate biological functions. We identified 354 miRNAs from FCx and 408 from HP using miRNA-Seq, and 245 from FCx and 238 from HP with microarrays. Several miRNA families and clusters were differentially expressed between FCx and HP, including the miR-8 family, miR-182|miR-96|miR-183 cluster, and miR-212|miR-312 cluster overexpressed in FCx and miR-34 family overexpressed in HP. To visualize the clusters, we developed support for viewing genomic alignments of miRNA-Seq reads in the Chipster genome browser. We carried out pathway analysis of the predicted target genes of differentially expressed miRNA families and clusters to assess their putative biological functions. Interestingly, several miRNAs from the same family/cluster were predicted to regulate specific biological pathways. We have developed a miRNA-Seq approach with a bioinformatic analysis workflow that is suitable for studying miRNA expression patterns from specific brain nuclei. FCx and HP were shown to have distinct miRNA expression patterns which were reflected in the predicted gene regulatory pathways. This methodology can be applied for the identification of brain region-specific and phenotype-specific miRNA-mRNA-regulatory networks from the adult and developing rodent brain. PMID:21731767

  16. MicroRNA Expression Profiling Reveals MiRNA Families Regulating Specific Biological Pathways in Mouse Frontal Cortex and Hippocampus

    PubMed Central

    Juhila, Juuso; Sipilä, Tessa; Icay, Katherine; Nicorici, Daniel; Ellonen, Pekka; Kallio, Aleksi; Korpelainen, Eija; Greco, Dario; Hovatta, Iiris

    2011-01-01

    MicroRNAs (miRNAs) are small regulatory molecules that cause post-transcriptional gene silencing. Although some miRNAs are known to have region-specific expression patterns in the adult brain, the functional consequences of the region-specificity to the gene regulatory networks of the brain nuclei are not clear. Therefore, we studied miRNA expression patterns by miRNA-Seq and microarrays in two brain regions, frontal cortex (FCx) and hippocampus (HP), which have separate biological functions. We identified 354 miRNAs from FCx and 408 from HP using miRNA-Seq, and 245 from FCx and 238 from HP with microarrays. Several miRNA families and clusters were differentially expressed between FCx and HP, including the miR-8 family, miR-182|miR-96|miR-183 cluster, and miR-212|miR-312 cluster overexpressed in FCx and miR-34 family overexpressed in HP. To visualize the clusters, we developed support for viewing genomic alignments of miRNA-Seq reads in the Chipster genome browser. We carried out pathway analysis of the predicted target genes of differentially expressed miRNA families and clusters to assess their putative biological functions. Interestingly, several miRNAs from the same family/cluster were predicted to regulate specific biological pathways. We have developed a miRNA-Seq approach with a bioinformatic analysis workflow that is suitable for studying miRNA expression patterns from specific brain nuclei. FCx and HP were shown to have distinct miRNA expression patterns which were reflected in the predicted gene regulatory pathways. This methodology can be applied for the identification of brain region-specific and phenotype-specific miRNA-mRNA-regulatory networks from the adult and developing rodent brain. PMID:21731767

  17. An Integrative Analysis of microRNA and mRNA Profiling in CML Stem Cells.

    PubMed

    Nassar, Farah J; El Eit, Rabab; Nasr, Rihab

    2016-01-01

    Integrative analysis of microRNA (miRNA) and messenger RNA (mRNA) in Chronic Myeloid leukemia (CML) stem cells is an important technique to study the involvement of miRNA and their targets in CML stem cells self-renewal, maintenance, and therapeutic resistance. Here, we describe a simplified integrative analysis using Ingenuity Pathway Analysis software after performing proper RNA extraction, miRNA and mRNA microarray and data analysis. PMID:27581151

  18. The microRNA cluster miR-183/96/182 contributes to long-term memory in a protein phosphatase 1-dependent manner

    PubMed Central

    Woldemichael, Bisrat T.; Jawaid, Ali; Kremer, Eloïse A.; Gaur, Niharika; Krol, Jacek; Marchais, Antonin; Mansuy, Isabelle M.

    2016-01-01

    Memory formation is a complex cognitive function regulated by coordinated synaptic and nuclear processes in neurons. In mammals, it is controlled by multiple molecular activators and suppressors, including the key signalling regulator, protein phosphatase 1 (PP1). Here, we show that memory control by PP1 involves the miR-183/96/182 cluster and its selective regulation during memory formation. Inhibiting nuclear PP1 in the mouse brain, or training on an object recognition task similarly increases miR-183/96/182 expression in the hippocampus. Mimicking this increase by miR-183/96/182 overexpression enhances object memory, while knocking-down endogenous miR-183/96/182 impairs it. This effect involves the modulation of several plasticity-related genes, with HDAC9 identified as an important functional target. Further, PP1 controls miR-183/96/182 in a transcription-independent manner through the processing of their precursors. These findings provide novel evidence for a role of miRNAs in memory formation and suggest the implication of PP1 in miRNAs processing in the adult brain. PMID:27558292

  19. The microRNA cluster miR-183/96/182 contributes to long-term memory in a protein phosphatase 1-dependent manner.

    PubMed

    Woldemichael, Bisrat T; Jawaid, Ali; Kremer, Eloïse A; Gaur, Niharika; Krol, Jacek; Marchais, Antonin; Mansuy, Isabelle M

    2016-01-01

    Memory formation is a complex cognitive function regulated by coordinated synaptic and nuclear processes in neurons. In mammals, it is controlled by multiple molecular activators and suppressors, including the key signalling regulator, protein phosphatase 1 (PP1). Here, we show that memory control by PP1 involves the miR-183/96/182 cluster and its selective regulation during memory formation. Inhibiting nuclear PP1 in the mouse brain, or training on an object recognition task similarly increases miR-183/96/182 expression in the hippocampus. Mimicking this increase by miR-183/96/182 overexpression enhances object memory, while knocking-down endogenous miR-183/96/182 impairs it. This effect involves the modulation of several plasticity-related genes, with HDAC9 identified as an important functional target. Further, PP1 controls miR-183/96/182 in a transcription-independent manner through the processing of their precursors. These findings provide novel evidence for a role of miRNAs in memory formation and suggest the implication of PP1 in miRNAs processing in the adult brain. PMID:27558292

  20. MicroRNA Regulation of Human Breast Cancer Stem Cells

    PubMed Central

    Shimono, Yohei; Mukohyama, Junko; Nakamura, Shun-ichi; Minami, Hironobu

    2015-01-01

    MicroRNAs (miRNAs) are involved in virtually all biological processes, including stem cell maintenance, differentiation, and development. The dysregulation of miRNAs is associated with many human diseases including cancer. We have identified a set of miRNAs differentially expressed between human breast cancer stem cells (CSCs) and non-tumorigenic cancer cells. In addition, these miRNAs are similarly upregulated or downregulated in normal mammary stem/progenitor cells. In this review, we mainly describe the miRNAs that are dysregulated in human breast CSCs directly isolated from clinical specimens. The miRNAs and their clusters, such as the miR-200 clusters, miR-183 cluster, miR-221-222 cluster, let-7, miR-142 and miR-214, target the genes and pathways important for stem cell maintenance, such as the self-renewal gene BMI1, apoptosis, Wnt signaling, Notch signaling, and epithelial-to-mesenchymal transition. In addition, the current evidence shows that metastatic breast CSCs acquire a phenotype that is different from the CSCs in a primary site. Thus, clarifying the miRNA regulation of the metastatic breast CSCs will further advance our understanding of the roles of human breast CSCs in tumor progression. PMID:26712794

  1. MicroRNA miR-125a controls hematopoietic stem cell number

    PubMed Central

    Guo, Shangqin; Lu, Jun; Schlanger, Rita; Zhang, Hao; Wang, Judy Y.; Fox, Michelle C.; Purton, Louise E.; Fleming, Heather H.; Cobb, Bradley; Merkenschlager, Matthias; Golub, Todd R.; Scadden, David T.

    2010-01-01

    MicroRNAs influence hematopoietic differentiation, but little is known about their effects on the stem cell state. Here, we report that the microRNA processing enzyme Dicer is essential for stem cell persistence in vivo and a specific microRNA, miR-125a, controls the size of the stem cell population by regulating hematopoietic stem/progenitor cell (HSPC) apoptosis. Conditional deletion of Dicer revealed an absolute dependence for the multipotent HSPC population in a cell-autonomous manner, with increased HSPC apoptosis in mutant animals. An evolutionarily conserved microRNA cluster containing miR-99b, let-7e, and miR-125a was preferentially expressed in long-term hematopoietic stem cells. MicroRNA miR-125a alone was capable of increasing the number of hematopoietic stem cells in vivo by more than 8-fold. This result was accomplished through a differentiation stage-specific reduction of apoptosis in immature hematopoietic progenitors, possibly through targeting multiple proapoptotic genes. Bak1 was directly down-regulated by miR-125a and expression of a 3′UTR-less Bak1 blocked miR-125a-induced hematopoietic expansion in vivo. These data demonstrate cell-state-specific regulation by microRNA and identify a unique microRNA functioning to regulate the stem cell pool size. PMID:20616003

  2. MicroRNA target prediction in glaucoma.

    PubMed

    Romano, Giovanni Luca; Platania, Chiara Bianca Maria; Forte, Stefano; Salomone, Salvatore; Drago, Filippo; Bucolo, Claudio

    2015-01-01

    Glaucoma is a progressive optic neuropathy and is one of the leading causes of blindness in the industrialized countries. The aim of this study is to investigate microRNA (miRNA) regulation in glaucoma and other neurodegenerative diseases, that share similar pathways, by means of in silico approaches such as bibliographic search and access to bioinformatic resources. First of all, data mining was carried out on Human miRNA Disease Database (HMDD) and miR2Disease databases. Then, predictions of deregulated miRNAs were carried out accessing to microrna.org database. Finally, the potential combinatorial effect of miRNAs, on regulation of biochemical pathways, was studied by an enrichment analysis performed by DIANA-miRPath v.2.0. We found, from literature search, 8 deregulated miRNAs in glaucoma and 9 and 23 in age-related macular degeneration (AMD) and Alzheimer's disease (AD), respectively. One miRNA is commonly deregulated in glaucoma and AMD (miR-23a). Two miRNAs (miR-29a, miR-29b) are common to glaucoma and AD, and four miRNAs were identified to be commonly deregulated in AMD and AD (miR-9, miR-21, miR-34a, miR-146a). The match of the miRNA common to glaucoma and the other two neurodegenerative diseases (AMD and AD) did not generate any output. Enrichment of information has been reached through miRNAs prediction: 88 predicted miRNAs are common to glaucoma and AMD, 19 are common to glaucoma and AD, and 9 are common to AMD and AD. Indeed, predicted miRNAs common to the three neurodegenerative diseases are nine (miR-107, miR-137, miR-146a, miR-181c, miR-197, miR-21, miR-22, miR-590, miR-9). DIANA-miRPath predicted that those nine miRNAs might regulate pathways involved in inflammation. The findings hereby obtained provide a valuable hint to assess deregulation of specific miRNA, as potential biomarkers and therapeutic targets, in glaucoma and other neurodegenerative diseases by means of preclinical and clinical studies. PMID:26497793

  3. MicroRNA-155 Reinforces HIV Latency.

    PubMed

    Ruelas, Debbie S; Chan, Jonathan K; Oh, Eugene; Heidersbach, Amy J; Hebbeler, Andrew M; Chavez, Leonard; Verdin, Eric; Rape, Michael; Greene, Warner C

    2015-05-29

    The presence of a small number of infected but transcriptionally dormant cells currently thwarts a cure for the more than 35 million individuals infected with HIV. Reactivation of these latently infected cells may result in three fates: 1) cell death due to a viral cytopathic effect, 2) cell death due to immune clearance, or 3) a retreat into latency. Uncovering the dynamics of HIV gene expression and silencing in the latent reservoir will be crucial for developing an HIV-1 cure. Here we identify and characterize an intracellular circuit involving TRIM32, an HIV activator, and miR-155, a microRNA that may promote a return to latency in these transiently activated reservoir cells. Notably, we demonstrate that TRIM32, an E3 ubiquitin ligase, promotes reactivation from latency by directly modifying IκBα, leading to a novel mechanism of NF-κB induction not involving IκB kinase activation. PMID:25873391

  4. MicroRNA-155 Reinforces HIV Latency*

    PubMed Central

    Ruelas, Debbie S.; Chan, Jonathan K.; Oh, Eugene; Heidersbach, Amy J.; Hebbeler, Andrew M.; Chavez, Leonard; Verdin, Eric; Rape, Michael; Greene, Warner C.

    2015-01-01

    The presence of a small number of infected but transcriptionally dormant cells currently thwarts a cure for the more than 35 million individuals infected with HIV. Reactivation of these latently infected cells may result in three fates: 1) cell death due to a viral cytopathic effect, 2) cell death due to immune clearance, or 3) a retreat into latency. Uncovering the dynamics of HIV gene expression and silencing in the latent reservoir will be crucial for developing an HIV-1 cure. Here we identify and characterize an intracellular circuit involving TRIM32, an HIV activator, and miR-155, a microRNA that may promote a return to latency in these transiently activated reservoir cells. Notably, we demonstrate that TRIM32, an E3 ubiquitin ligase, promotes reactivation from latency by directly modifying IκBα, leading to a novel mechanism of NF-κB induction not involving IκB kinase activation. PMID:25873391

  5. MicroRNA response to DNA damage

    PubMed Central

    Wan, Guohui; Mathur, Rohit; Hu, Xiaoxiao; Zhang, Xinna; Lu, Xiongbin

    2011-01-01

    Faithful transmission of genetic material in eukaryotic cells requires not only accurate DNA replication and chromosome distribution, but also the ability to sense and repair spontaneous and induced DNA damage. To maintain genomic integrity, cells undergo a DNA damage response using a complex network of signaling pathways, composed of coordinate sensors, transducers and effectors in cell cycle arrest, apoptosis and DNA repair. Emerging evidence has suggested that microRNAs (miRNAs) play a critical role in regulation of DNA damage response. Here, we discuss the recent findings on how miRNAs interact with the canonical DNA damage response and how miRNA expression is regulated after DNA damage. PMID:21741842

  6. Epigenetics, microRNA, and addiction

    PubMed Central

    Kenny, Paul J.

    2014-01-01

    Drug addiction is characterized by uncontrolled drug consumption and high rates of relapse to drug taking during periods of attempted abstinence. Addiction is now largely considered a disorder of experience-dependent neuroplasticity, driven by remodeling of synapses in reward and motivation relevant brain circuits in response to a history of prolonged drug intake. Alterations in gene expression play a central role in addiction-relevant neuroplasticity, but the mechanisms by which additive drugs remodel brain motivation circuits remains unclear. MicroRNAs (miRNAs) are a class of noncoding RNA that can regulate the expression of large numbers of protein-coding mRNA transcripts by binding to the 3' untranslated region (3' UTR) of target transcripts and blocking their translation into the encoded protein or triggering their destabilization and degradation. Emerging evidence has implicated miRNAs in regulating addiction-relevant neuroplasticity in the brain, and in controlling the motivational properties of cocaine and other drugs of abuse. Here, the role for miRNAs in regulating basic aspects of neuronal function is reviewed. The involvement of miRNAs in controlling the motivational properties of addictive drugs is also summarized. Finally, mechanisms by which miRNAs exert their actions on drug intake, when known, are considered. PMID:25364284

  7. Insight into temperature-dependent microRNA function in mammalian hibernators

    PubMed Central

    Biggar, Kyle K; Storey, Kenneth B

    2014-01-01

    Mammalian hibernation involves re-programming of metabolic functions, in part, facilitated by microRNA. Although much is known about microRNA function, we lack knowledge on low temperature microRNA target selection. It is possible that the thermodynamics of microRNA target selection could dictate unique temperature-dependent sets of microRNA targets for hibernators.

  8. MicroRNA modulation in obesity and periodontitis.

    PubMed

    Perri, R; Nares, S; Zhang, S; Barros, S P; Offenbacher, S

    2012-01-01

    The aim of this pilot investigation was to determine if microRNA expression differed in the presence or absence of obesity, comparing gingival biopsies obtained from patients with or without periodontal disease. Total RNA was extracted from gingival biopsy samples collected from 20 patients: 10 non-obese patients (BMI < 30 kg/m(2)) and 10 obese patients (BMI > 30 kg/m(2)), each group with 5 periodontally healthy sites and 5 chronic periodontitis sites. MicroRNA expression patterns were assessed with a quantitative microRNA PCR array to survey 88 candidate microRNA species. Four microRNA databases were used to identify potential relevant mRNA target genes of differentially expressed microRNAs. Two microRNA species (miR-18a, miR-30e) were up-regulated among obese individuals with a healthy periodontium. Two microRNA species (miR-30e, miR-106b) were up-regulated in non-obese individuals with periodontal disease. In the presence of periodontal disease and obesity, 9 of 11 listed microRNAs were significantly up-regulated (miR-15a, miR-18a, miR-22, miR-30d, miR-30e, miR-103, miR-106b, miR-130a, miR-142-3p, miR-185, and miR-210). Predicted targets include 69 different mRNAs from genes that comprise cytokines, chemokines, specific collagens, and regulators of glucose and lipid metabolism. The expression of specific microRNA species in obesity, which could also target and post-transcriptionally modulate cytokine mRNA, provides new insight into possible mechanisms of how risk factors might modify periodontal inflammation and may represent novel therapeutic targets. PMID:22043006

  9. MicroRNA Transcriptome Profiles During Swine Skeletal Muscle Development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNA (miR) are a class of small RNAs that regulate gene expression by inhibiting translation of protein encoding transcripts. To evaluate the role of miR in skeletal muscle of swine, global microRNA abundance was measured at specific developmental stages including proliferating satellite cells,...

  10. MicroRNA in rectal cancer

    PubMed Central

    Azizian, Azadeh; Gruber, Jens; Ghadimi, B Michael; Gaedcke, Jochen

    2016-01-01

    In rectal cancer, one of the most common cancers worldwide, the proper staging of the disease determines the subsequent therapy. For those with locally advanced rectal cancer, a neoadjuvant chemoradiotherapy (CRT) is recommended before any surgery. However, response to CRT ranges from complete response (responders) to complete resistance (non-responders). To date we are not able to separate in advance the first group from the second, due to the absence of a valid biomarker. Therefore all patients receive the same therapy regardless of whether they reap benefits. On the other hand almost all patients receive a surgical resection after the CRT, although a watch-and-wait procedure or an endoscopic resection might be sufficient for those who responded well to the CRT. Being highly conserved regulators of gene expression, microRNAs (miRNAs) seem to be promising candidates for biomarkers. Many studies have been analyzing the miRNAs expressed in rectal cancer tissue to determine a specific miRNA profile for the ailment. Unfortunately, there is only a small overlap of identified miRNAs between different studies, posing the question as to whether different methods or differences in tissue storage may contribute to that fact or if the results simply are not reproducible, due to unknown factors with undetected influences on miRNA expression. Other studies sought to find miRNAs which correlate to clinical parameters (tumor grade, nodal stage, metastasis, survival) and therapy response. Although several miRNAs seem to have an impact on the response to CRT or might predict nodal stage, there is still only little overlap between different studies. We here aimed to summarize the current literature on rectal cancer and miRNA expression with respect to the different relevant clinical parameters. PMID:27190581

  11. MicroRNA profiles in various hepatocellular carcinoma cell lines

    PubMed Central

    Morishita, Asahiro; Iwama, Hisakazu; Fujihara, Shintaro; Sakamoto, Teppei; Fujita, Koji; Tani, Joji; Miyoshi, Hisaaki; Yoneyama, Hirohito; Himoto, Takashi; Masaki, Tsutomu

    2016-01-01

    Hepatocellular carcinoma (HCC) is one of the most common causes of cancer-associated mortality worldwide. Although surgery is considered the most effective treatment for patients with HCC, its indication is restricted by limited criteria and a high relapse rate following surgery; therefore, systemic chemotherapy is required for patients with advanced-stage HCC to prolong their survival. MicroRNAs (miRNAs) are endogenous non-coding RNAs of 18–22 nucleotides in length. It has been reported that aberrant expression of miRNAs is a feature shared by various types of human cancer. Previous studies have indicated that the modulation of non-coding RNAs, particularly miRNAs, may be a valuable therapeutic target for HCC. The aim of the present study was to elucidate the miRNA profiles associated with differentiation and hepatitis B virus (HBV) infection observed in HCC cell lines. The human Alex, Hep3B, HepG2, HuH1, HuH7, JHH1, JHH2, JHH5, JHH6, HLE, HLF and Li-7 HCC cell lines were used for an miRNA array. Replicate data were analyzed following their classification into: i) Poorly- and well-differentiated human HCC cells and ii) HBV-positive and -negative human HCC cells. Out of the 1,719 miRNAs, 4 were found to be significantly upregulated and 52 significantly downregulated in the poorly-differentiated cells, as compared with the well-differentiated cells. Conversely, in the HBV-positive cells 125 miRNAs were found to be upregulated and 2 downregulated, as compared with the HBV-negative cells. Unsupervised hierarchical clustering analysis with Pearson's correlation revealed that the miRNA expression levels were clustered both together and separately in each group. In conclusion, miRNA profile characterization based on various parameters may be a novel approach to determine the etiology of HCC.

  12. MicroRNA 665 Regulates Dentinogenesis through MicroRNA-Mediated Silencing and Epigenetic Mechanisms

    PubMed Central

    Heair, Hannah M.; Kemper, Austin G.; Roy, Bhaskar; Lopes, Helena B.; Rashid, Harunur; Clarke, John C.; Afreen, Lubana K.; Ferraz, Emanuela P.; Kim, Eddy; Javed, Amjad; Beloti, Marcio M.; MacDougall, Mary

    2015-01-01

    Studies of proteins involved in microRNA (miRNA) processing, maturation, and silencing have indicated the importance of miRNAs in skeletogenesis, but the specific miRNAs involved in this process are incompletely defined. Here, we identified miRNA 665 (miR-665) as a potential repressor of odontoblast maturation. Studies with cultured cell lines and primary embryonic cells showed that miR-665 represses the expression of early and late odontoblast marker genes and stage-specific proteases involved in dentin maturation. Notably, miR-665 directly targeted Dlx3 mRNA and decreased Dlx3 expression. Furthermore, RNA-induced silencing complex (RISC) immunoprecipitation and biotin-labeled miR-665 pulldown studies identified Kat6a as another potential target of miR-665. KAT6A interacted physically and functionally with RUNX2, activating tissue-specific promoter activity and prompting odontoblast differentiation. Overexpression of miR-665 reduced the recruitment of KAT6A to Dspp and Dmp1 promoters and prevented KAT6A-induced chromatin remodeling, repressing gene transcription. Taken together, our results provide novel molecular evidence that miR-665 functions in an miRNA-epigenetic regulatory network to control dentinogenesis. PMID:26124283

  13. MicroRNA 665 Regulates Dentinogenesis through MicroRNA-Mediated Silencing and Epigenetic Mechanisms.

    PubMed

    Heair, Hannah M; Kemper, Austin G; Roy, Bhaskar; Lopes, Helena B; Rashid, Harunur; Clarke, John C; Afreen, Lubana K; Ferraz, Emanuela P; Kim, Eddy; Javed, Amjad; Beloti, Marcio M; MacDougall, Mary; Hassan, Mohammad Q

    2015-09-01

    Studies of proteins involved in microRNA (miRNA) processing, maturation, and silencing have indicated the importance of miRNAs in skeletogenesis, but the specific miRNAs involved in this process are incompletely defined. Here, we identified miRNA 665 (miR-665) as a potential repressor of odontoblast maturation. Studies with cultured cell lines and primary embryonic cells showed that miR-665 represses the expression of early and late odontoblast marker genes and stage-specific proteases involved in dentin maturation. Notably, miR-665 directly targeted Dlx3 mRNA and decreased Dlx3 expression. Furthermore, RNA-induced silencing complex (RISC) immunoprecipitation and biotin-labeled miR-665 pulldown studies identified Kat6a as another potential target of miR-665. KAT6A interacted physically and functionally with RUNX2, activating tissue-specific promoter activity and prompting odontoblast differentiation. Overexpression of miR-665 reduced the recruitment of KAT6A to Dspp and Dmp1 promoters and prevented KAT6A-induced chromatin remodeling, repressing gene transcription. Taken together, our results provide novel molecular evidence that miR-665 functions in an miRNA-epigenetic regulatory network to control dentinogenesis. PMID:26124283

  14. Roquin binds microRNA-146a and Argonaute2 to regulate microRNA homeostasis

    PubMed Central

    Srivastava, Monika; Duan, Guowen; Kershaw, Nadia J.; Athanasopoulos, Vicki; Yeo, Janet H. C.; Ose, Toyoyuki; Hu, Desheng; Brown, Simon H. J.; Jergic, Slobodan; Patel, Hardip R.; Pratama, Alvin; Richards, Sashika; Verma, Anil; Jones, E. Yvonne; Heissmeyer, Vigo; Preiss, Thomas; Dixon, Nicholas E.; Chong, Mark M. W.; Babon, Jeffrey J.; Vinuesa, Carola G.

    2015-01-01

    Roquin is an RNA-binding protein that prevents autoimmunity and inflammation via repression of bound target mRNAs such as inducible costimulator (Icos). When Roquin is absent or mutated (Roquinsan), Icos is overexpressed in T cells. Here we show that Roquin enhances Dicer-mediated processing of pre-miR-146a. Roquin also directly binds Argonaute2, a central component of the RNA-induced silencing complex, and miR-146a, a microRNA that targets Icos mRNA. In the absence of functional Roquin, miR-146a accumulates in T cells. Its accumulation is not due to increased transcription or processing, rather due to enhanced stability of mature miR-146a. This is associated with decreased 3′ end uridylation of the miRNA. Crystallographic studies reveal that Roquin contains a unique HEPN domain and identify the structural basis of the ‘san’ mutation and Roquin’s ability to bind multiple RNAs. Roquin emerges as a protein that can bind Ago2, miRNAs and target mRNAs, to control homeostasis of both RNA species. PMID:25697406

  15. RNA Secondary Structure Modulates FMRP's Bi-Functional Role in the MicroRNA Pathway.

    PubMed

    Kenny, Phillip; Ceman, Stephanie

    2016-01-01

    MicroRNAs act by post-transcriptionally regulating the gene expression of 30%-60% of mammalian genomes. MicroRNAs are key regulators in all cellular processes, though the mechanism by which the cell activates or represses microRNA-mediated translational regulation is poorly understood. In this review, we discuss the RNA binding protein Fragile X Mental Retardation Protein (FMRP) and its role in microRNA-mediated translational regulation. Historically, FMRP is known to function as a translational suppressor. However, emerging data suggests that FMRP has both an agonistic and antagonistic role in regulating microRNA-mediated translational suppression. This bi-functional role is dependent on FMRP's interaction with the RNA helicase Moloney leukemia virus 10 (MOV10), which modifies the structural landscape of bound mRNA, therefore facilitating or inhibiting its association with the RNA-Induced Silencing Complex. PMID:27338369

  16. RNA editing of microRNA prevents RNA-induced silencing complex recognition of target mRNA

    PubMed Central

    Cui, Yalei; Huang, Tianzhi; Zhang, Xiaobo

    2015-01-01

    MicroRNAs (miRNAs) integrate with Argonaut (Ago) to create the RNA-induced silencing complex, and regulate gene expression by silencing target mRNAs. RNA editing of miRNA may affect miRNA processing, assembly of the Ago complex and target mRNA binding. However, the function of edited miRNA, assembled within the Ago complex, has not been extensively investigated. In this study, sequence analysis of the Ago complex of Marsupenaeus japonicus shrimp infected with white spot syndrome virus (WSSV) revealed that host ADAR (adenosine deaminase acting on RNA) catalysed A-to-I RNA editing of a viral miRNA (WSSV-miR-N12) at the +16 site. This editing of the non-seed sequence did not affect association of the edited miRNA with the Ago protein, but inhibited interaction between the miRNA and its target gene (wsv399). The WSSV early gene wsv399 inhibited WSSV infection. As a result, the RNA editing of miRNA caused virus latency. Our results highlight a novel example of miRNA editing in the miRNA-induced silencing complex. PMID:26674414

  17. High-Throughput Functional MicroRNA Profiling Using Recombinant AAV-Based MicroRNA Sensor Arrays

    PubMed Central

    Tian, Wenhong; Dong, Xiaoyan; Wu, Xiaobing; Wu, Zhijian

    2014-01-01

    There is a lack of methods for high-throughput functional microRNA (miRNA) profiling. In this chapter, we describe a recombinant adeno-associated virus-based miRNA sensor array (miRNA Asensor array), which is able to profile functional miRNAs in cultured cells. The preparation of an miRNA Asensor array and its usage are discussed. PMID:24026702

  18. Integrated microRNA-mRNA analyses reveal OPLL specific microRNA regulatory network using high-throughput sequencing.

    PubMed

    Xu, Chen; Chen, Yu; Zhang, Hao; Chen, Yuanyuan; Shen, Xiaolong; Shi, Changgui; Liu, Yang; Yuan, Wen

    2016-01-01

    Ossification of the posterior longitudinal ligament (OPLL) is a genetic disorder which involves pathological heterotopic ossification of the spinal ligaments. Although studies have identified several genes that correlated with OPLL, the underlying regulation network is far from clear. Through small RNA sequencing, we compared the microRNA expressions of primary posterior longitudinal ligament cells form OPLL patients with normal patients (PLL) and identified 218 dysregulated miRNAs (FDR < 0.01). Furthermore, assessing the miRNA profiling data of multiple cell types, we found these dysregulated miRNAs were mostly OPLL specific. In order to decipher the regulation network of these OPLL specific miRNAs, we integrated mRNA expression profiling data with miRNA sequencing data. Through computational approaches, we showed the pivotal roles of these OPLL specific miRNAs in heterotopic ossification of longitudinal ligament by discovering highly correlated miRNA/mRNA pairs that associated with skeletal system development, collagen fibril organization, and extracellular matrix organization. The results of which provide strong evidence that the miRNA regulatory networks we established may indeed play vital roles in OPLL onset and progression. To date, this is the first systematic analysis of the micronome in OPLL, and thus may provide valuable resources in finding novel treatment and diagnostic targets of OPLL. PMID:26868491

  19. Profile of microRNA in Blood Plasma of Healthy Humans.

    PubMed

    Skurnikov, M Yu; Makarova, Yu A; Knyazev, E N; Fomicheva, K A; Nyushko, K M; Saribekyan, E K; Alekseev, B Ya; Kaprin, A D

    2016-03-01

    Analysis of the plasma microRNA profile can be used for the diagnosis of various pathological and physiological conditions. Complete microRNA microprofiling is an extremely important task. Here we used microarray analysis allowing measurement of the expression of 2500 microRNA (MirBase, version 20). About 10% known microRNA were found in the plasma. Most of the detected microRNA (69 microRNA; ~30%) were encoded by mirtrons. PMID:27021098

  20. Bio-barcode gel assay for microRNA

    NASA Astrophysics Data System (ADS)

    Lee, Hyojin; Park, Jeong-Eun; Nam, Jwa-Min

    2014-02-01

    MicroRNA has been identified as a potential biomarker because expression level of microRNA is correlated with various cancers. Its detection at low concentrations would be highly beneficial for cancer diagnosis. Here, we develop a new type of a DNA-modified gold nanoparticle-based bio-barcode assay that uses a conventional gel electrophoresis platform and potassium cyanide chemistry and show this assay can detect microRNA at aM levels without enzymatic amplification. It is also shown that single-base-mismatched microRNA can be differentiated from perfectly matched microRNA and the multiplexed detection of various combinations of microRNA sequences is possible with this approach. Finally, differently expressed microRNA levels are selectively detected from cancer cells using the bio-barcode gel assay, and the results are compared with conventional polymerase chain reaction-based results. The method and results shown herein pave the way for practical use of a conventional gel electrophoresis for detecting biomolecules of interest even at aM level without polymerase chain reaction amplification.

  1. MicroRNA dysregulation in melanoma.

    PubMed

    Latchana, Nicholas; Ganju, Akaansha; Howard, J Harrison; Carson, William E

    2016-09-01

    Melanoma is the deadliest form of skin cancer. Current challenges facing the management of melanoma include accurate prediction of individuals who will respond to adjuvant therapies as well as early detection of recurrences. These and other challenges have prompted investigation into biomarkers that could be used as diagnostic, prognostic and therapeutic aids. MicroRNAs (miRs) are small 19-22 nucleotide RNA inhibitors of protein translation. Over 800 different miRs are present within cells and importantly miR expression profiles may vary across different cells types and stages of malignancy. Unique expression profiles have been described for malignant melanoma; however, this work has yet to be translated into routine clinical practice. We highlight pertinent studies involving common miRs implicated in the oncogenesis of melanoma including miR-21, miR-125b, miR-150, miR-155, miR-205, and miR-211. In particular, emphasis is placed upon differential expression across different stages of melanoma progression, prognostic implications and potential mechanistic involvement. Focused efforts on inhibition of these miRs could be the most efficient method of translating preclinical endeavors into clinically meaningful applications. PMID:27566021

  2. MicroRNA and Heart Failure.

    PubMed

    Wong, Lee Lee; Wang, Juan; Liew, Oi Wah; Richards, Arthur Mark; Chen, Yei-Tsung

    2016-01-01

    Heart failure (HF) imposes significant economic and public health burdens upon modern society. It is known that disturbances in neurohormonal status play an important role in the pathogenesis of HF. Therapeutics that antagonize selected neurohormonal pathways, specifically the renin-angiotensin-aldosterone and sympathetic nervous systems, have significantly improved patient outcomes in HF. Nevertheless, mortality remains high with about 50% of HF patients dying within five years of diagnosis thus mandating ongoing efforts to improve HF management. The discovery of short noncoding microRNAs (miRNAs) and our increasing understanding of their functions, has presented potential therapeutic applications in complex diseases, including HF. Results from several genome-wide miRNA studies have identified miRNAs differentially expressed in HF cohorts suggesting their possible involvement in the pathogenesis of HF and their potential as both biomarkers and as therapeutic targets. Unravelling the functional relevance of miRNAs within pathogenic pathways is a major challenge in cardiovascular research. In this article, we provide an overview of the role of miRNAs in the cardiovascular system. We highlight several HF-related miRNAs reported from selected cohorts and review their putative roles in neurohormonal signaling. PMID:27058529

  3. MicroRNA and Heart Failure

    PubMed Central

    Wong, Lee Lee; Wang, Juan; Liew, Oi Wah; Richards, Arthur Mark; Chen, Yei-Tsung

    2016-01-01

    Heart failure (HF) imposes significant economic and public health burdens upon modern society. It is known that disturbances in neurohormonal status play an important role in the pathogenesis of HF. Therapeutics that antagonize selected neurohormonal pathways, specifically the renin-angiotensin-aldosterone and sympathetic nervous systems, have significantly improved patient outcomes in HF. Nevertheless, mortality remains high with about 50% of HF patients dying within five years of diagnosis thus mandating ongoing efforts to improve HF management. The discovery of short noncoding microRNAs (miRNAs) and our increasing understanding of their functions, has presented potential therapeutic applications in complex diseases, including HF. Results from several genome-wide miRNA studies have identified miRNAs differentially expressed in HF cohorts suggesting their possible involvement in the pathogenesis of HF and their potential as both biomarkers and as therapeutic targets. Unravelling the functional relevance of miRNAs within pathogenic pathways is a major challenge in cardiovascular research. In this article, we provide an overview of the role of miRNAs in the cardiovascular system. We highlight several HF-related miRNAs reported from selected cohorts and review their putative roles in neurohormonal signaling. PMID:27058529

  4. MicroRNA processing without Dicer

    PubMed Central

    2010-01-01

    The canonical processing of precursor microRNAs requires the endonuclease Dicer. A recent study shows that microRNAs can be processed independently of Dicer but instead require Argonaute 2. PMID:20565849

  5. MicroRNA in oral cancer research: future prospects.

    PubMed

    Sarode, Sachin C; Sarode, Gargi S; Patil, Shankargouda

    2014-01-01

    MicroRNA (miRNA) and related therapeutic approaches hold great promise in the field of cancer managements. Various studies on epithelial malignancies have shown encouraging results on various fronts. Its association with invasion, tumor growth, epithelial mesenchymal transition (EMT), angiogenesis, cancer stem cells (CSCs), metastasis and refects the diversified role of miRNA. Moreover, miRNA plays an important role in determining the prognosis of the patients. MicroRNAs interactions with each other and with external factors [human papilloma virus (HPV) (like oncoproteins)] intrigue us to explore more deep into this fascinating world.(1.) PMID:25707845

  6. microRNAs and microRNA Targets Involved in Alfalfa Stem Development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To examine the possible involvement of microRNAs in alfalfa stem development, we hybridized 32P-labled total microRNAs purified from elongating and post-elongation stem internodes (ES and PES, respectively) of the alfalfa Clone 252 to a microRNA dot blot that contains a total of 70 reference anti-mi...

  7. NPK macronutrients and microRNA homeostasis

    PubMed Central

    Kulcheski, Franceli R.; Côrrea, Régis; Gomes, Igor A.; de Lima, Júlio C.; Margis, Rogerio

    2015-01-01

    Macronutrients are essential elements for plant growth and development. In natural, non-cultivated systems, the availability of macronutrients is not a limiting factor of growth, due to fast recycling mechanisms. However, their availability might be an issue in modern agricultural practices, since soil has been frequently over exploited. From a crop management perspective, the nitrogen (N), phosphorus (P), and potassium (K) are three important limiting factors and therefore frequently added as fertilizers. NPK are among the nutrients that have been reported to alter post-embryonic root developmental processes and consequently, impairs crop yield. To cope with nutrients scarcity, plants have evolved several mechanisms involved in metabolic, physiological, and developmental adaptations. In this scenario, microRNAs (miRNAs) have emerged as additional key regulators of nutrients uptake and assimilation. Some studies have demonstrated the intrinsic relation between miRNAs and their targets, and how they can modulate plants to deal with the NPK availability. In this review, we focus on miRNAs and their regulation of targets involved in NPK metabolism. In general, NPK starvation is related with miRNAs that are involved in root-architectural changes and uptake activity modulation. We further show that several miRNAs were discovered to be involved in plant–microbe symbiosis during N and P uptake, and in this way we present a global view of some studies that were conducted in the last years. The integration of current knowledge about miRNA-NPK signaling may help future studies to focus in good candidates genes for the development of important tools for plant nutritional breeding. PMID:26136763

  8. Combination of microRNA expression profiling with genome-wide SNP genotyping to construct a coronary artery disease-related miRNA-miRNA synergistic network.

    PubMed

    Hua, Lin; Xia, Hong; Zhou, Ping; Li, Dongguo; Li, Lin

    2014-12-01

    In recent years, microRNAs (miRNAs) were found to play critical roles in many important biological processes. On the other hand, the rapid development of genome-wide association studies (GWAS) help identify potential genetic variants associated with the disease phenotypic variance. Therefore, we suggested a combined analysis of microRNA expression profiling with genome-wide Single Nucleotide Polymorphism (SNP) genotyping to identify potential disease-related biomarkers. Considering functional SNPs in miRNA genes or target sites might be important signals associated with human complex diseases, we constructed a miRNA-miRNA synergistic network related to coronary artery disease (CAD) by performing a genome-wide scan for SNPs in human miRNA 3' -untranslated regions (UTRs) target sites and computed potential SNP cooperation effects contributing to disease based on potential miRNA-SNP interactions reported recently. Furthermore, we identified some potential CAD-related miRNAs by analyzing the constructed miRNAmiRNA synergistic network. As a result, the predicted miRNA-miRNA network and miRNA clusters were validated by significantly high interaction effects of CAD-related miRNAs. Accurate classification performances were obtained for all of the identified miRNA clusters, and the sensitivity and specificity were all more than 90%. The network topological analysis confirmed some novel CAD-related miRNAs identified recently by experiments. Our method might help to understand miRNA function and CAD disease, as well as to explore the novel mechanisms involved. PMID:25641175

  9. MicroRNA-145 Regulates Human Corneal Epithelial Differentiation

    PubMed Central

    Ng, Tsz-Kin; Huang, Li; Lei, Peng; Choy, Kwong-Wai; Liu, Yingpeng; Zhang, Mingzhi; Lam, Dennis Shun-Chiu; Yam, Gary Hin-Fai; Pang, Chi-Pui

    2011-01-01

    Background Epigenetic factors, such as microRNAs, are important regulators in the self-renewal and differentiation of stem cells and progenies. Here we investigated the microRNAs expressed in human limbal-peripheral corneal (LPC) epithelia containing corneal epithelial progenitor cells (CEPCs) and early transit amplifying cells, and their role in corneal epithelium. Methodology/Principal Findings Human LPC epithelia was extracted for small RNAs or dissociated for CEPC culture. By Agilent Human microRNA Microarray V2 platform and GeneSpring GX11.0 analysis, we found differential expression of 18 microRNAs against central corneal (CC) epithelia, which were devoid of CEPCs. Among them, miR-184 was up-regulated in CC epithelia, similar to reported finding. Cluster miR-143/145 was expressed strongly in LPC but weakly in CC epithelia (P = 0.0004, Mann-Whitney U-test). This was validated by quantitative polymerase chain reaction (qPCR). Locked nucleic acid-based in situ hybridization on corneal rim cryosections showed miR-143/145 presence localized to the parabasal cells of limbal epithelium but negligible in basal and superficial epithelia. With holoclone forming ability, CEPCs transfected with lentiviral plasmid containing mature miR-145 sequence gave rise to defective epithelium in organotypic culture and had increased cytokeratin-3/12 and connexin-43 expressions and decreased ABCG2 and p63 compared with cells transfected with scrambled sequences. Global gene expression was analyzed using Agilent Whole Human Genome Oligo Microarray and GeneSpring GX11.0. With a 5-fold difference compared to cells with scrambled sequences, miR-145 up-regulated 324 genes (containing genes for immune response) and down-regulated 277 genes (containing genes for epithelial development and stem cell maintenance). As validated by qPCR and luciferase reporter assay, our results showed miR-145 suppressed integrin β8 (ITGB8) expression in both human corneal epithelial cells and primary

  10. Robust and Adaptive MicroRNA-Mediated Incoherent Feedforward Motifs

    NASA Astrophysics Data System (ADS)

    Xu, Feng-Dan; Liu, Zeng-Rong; Zhang, Zhi-Yong; Shen, Jian-Wei

    2009-02-01

    We integrate transcriptional and post-transcriptional regulation into microRNA-mediated incoherent feedforward motifs and analyse their dynamical behaviour and functions. The analysis show that the behaviour of the system is almost uninfluenced by the varying input in certain ranges and by introducing of delay and noise. The results indicate that microRNA-mediated incoherent feedforward motifs greatly enhance the robustness of gene regulation.

  11. Epigenetic and microRNA regulation during osteoarthritis development

    PubMed Central

    Chen, Di; Shen, Jie; Hui, Tianqian

    2015-01-01

    Osteoarthritis (OA) is a common degenerative joint disease, the pathological mechanism of which is currently unknown. Genetic alteration is one of the key contributing factors for OA pathology. Recent evidence suggests that epigenetic and microRNA regulation of critical genes may contribute to OA development. In this article, we review the epigenetic and microRNA regulations of genes related to OA development. Potential therapeutic strategies may be developed on the basis of novel findings.

  12. MicroRNA and Transcriptional Crosstalk in Myelinating Glia

    PubMed Central

    Svaren, John

    2014-01-01

    Several recent studies have addressed the important role of microRNA in regulation of differentiation of myelinating glia. While Schwann cells and oligodendrocytes in the peripheral and central nervous systems, respectively, exhibit significant morphological and regulatory differences, some aspects of transcriptional and microRNA regulation are shared between these two cell types. This review focuses on the intersection of microRNAs with transcriptional regulation in Schwann cell and oligodendrocyte differentiation. In particular, several microRNAs have been shown to modulate expression of critical transcription factors, and in turn, the regulation of microRNA expression is enmeshed within transcriptional networks that coordinate both coding gene and noncoding RNA profiles of myelinating cells. These hubs of regulation control both myelin gene expression as well as the cell cycle transitions of Schwann cells and oligodendrocytes as they terminally differentiate. In addition, some studies have begin to highlight the combinatorial effects of different microRNAs that establish the narrow range of gene regulation required for efficient and stable myelin formation. Overall, the integration of microRNA and transcriptional aspects will help elucidate mechanistic control of the myelination process. PMID:24979526

  13. piRNA cluster database: a web resource for piRNA producing loci

    PubMed Central

    Rosenkranz, David

    2016-01-01

    Piwi proteins and their guiding small RNAs, termed Piwi-interacting (pi-) RNAs, are essential for silencing of transposons in the germline of animals. A substantial fraction of piRNAs originates from genomic loci termed piRNA clusters and sequences encoded in these piRNA clusters determine putative targets for the Piwi/piRNA system. In the past decade, studies of piRNA transcriptomes in different species revealed additional roles for piRNAs beyond transposon silencing, reflecting the astonishing plasticity of the Piwi/piRNA system along different phylogenetic branches. Moreover, piRNA transcriptomes can change drastically during development and vary across different tissues. Since piRNA clusters crucially shape piRNA profiles, analysis of these loci is imperative for a thorough understanding of functional and evolutionary aspects of the piRNA pathway. But despite the ever-growing amount of available piRNA sequence data, we know little about the factors that determine differential regulation of piRNA clusters, nor the evolutionary events that cause their gain or loss. In order to facilitate addressing these subjects, we established a user-friendly piRNA cluster database (http://www.smallrnagroup-mainz.de/piRNAclusterDB.html) that provides comprehensive data on piRNA clusters in multiple species, tissues and developmental stages based on small RNA sequence data deposited at NCBI's Sequence Read Archive (SRA). PMID:26582915

  14. piRNA cluster database: a web resource for piRNA producing loci.

    PubMed

    Rosenkranz, David

    2016-01-01

    Piwi proteins and their guiding small RNAs, termed Piwi-interacting (pi-) RNAs, are essential for silencing of transposons in the germline of animals. A substantial fraction of piRNAs originates from genomic loci termed piRNA clusters and sequences encoded in these piRNA clusters determine putative targets for the Piwi/piRNA system. In the past decade, studies of piRNA transcriptomes in different species revealed additional roles for piRNAs beyond transposon silencing, reflecting the astonishing plasticity of the Piwi/piRNA system along different phylogenetic branches. Moreover, piRNA transcriptomes can change drastically during development and vary across different tissues.Since piRNA clusters crucially shape piRNA profiles, analysis of these loci is imperative for a thorough understanding of functional and evolutionary aspects of the piRNA pathway. But despite the ever-growing amount of available piRNA sequence data, we know little about the factors that determine differential regulation of piRNA clusters, nor the evolutionary events that cause their gain or loss.In order to facilitate addressing these subjects, we established a user-friendly piRNA cluster database (http://www.smallrnagroup-mainz.de/piRNAclusterDB.html) that provides comprehensive data on piRNA clusters in multiple species, tissues and developmental stages based on small RNA sequence data deposited at NCBI's Sequence Read Archive (SRA). PMID:26582915

  15. A MicroRNA precursor surveillance system in quality control of MicroRNA synthesis.

    PubMed

    Liu, Xuhang; Zheng, Qi; Vrettos, Nicholas; Maragkakis, Manolis; Alexiou, Panagiotis; Gregory, Brian D; Mourelatos, Zissimos

    2014-09-18

    MicroRNAs (miRNAs) are essential for regulation of gene expression. Though numerous miRNAs have been identified by high-throughput sequencing, few precursor miRNAs (pre-miRNAs) are experimentally validated. Here we report a strategy for constructing high-throughput sequencing libraries enriched for full-length pre-miRNAs. We find widespread and extensive uridylation of Argonaute (Ago)-bound pre-miRNAs, which is primarily catalyzed by two terminal uridylyltransferases: TUT7 and TUT4. Uridylation by TUT7/4 not only polishes pre-miRNA 3' ends, but also facilitates their degradation by the exosome, preventing clogging of Ago with defective species. We show that the exosome exploits distinct substrate preferences of DIS3 and RRP6, its two catalytic subunits, to distinguish productive from defective pre-miRNAs. Furthermore, we identify a positive feedback loop formed by the exosome and TUT7/4 in triggering uridylation and degradation of Ago-bound pre-miRNAs. Our study reveals a pre-miRNA surveillance system that comprises TUT7, TUT4, and the exosome in quality control of miRNA synthesis. PMID:25175028

  16. MicroRNA-21 in glomerular injury.

    PubMed

    Lai, Jennifer Y; Luo, Jinghui; O'Connor, Christopher; Jing, Xiaohong; Nair, Viji; Ju, Wenjun; Randolph, Ann; Ben-Dov, Iddo Z; Matar, Regina N; Briskin, Daniel; Zavadil, Jiri; Nelson, Robert G; Tuschl, Thomas; Brosius, Frank C; Kretzler, Matthias; Bitzer, Markus

    2015-04-01

    TGF-β(1) is a pleotropic growth factor that mediates glomerulosclerosis and podocyte apoptosis, hallmarks of glomerular diseases. The expression of microRNA-21 (miR-21) is regulated by TGF-β(1), and miR-21 inhibits apoptosis in cancer cells. TGF-β(1)-transgenic mice exhibit accelerated podocyte loss and glomerulosclerosis. We determined that miR-21 expression increases rapidly in cultured murine podocytes after exposure to TGF-β(1) and is higher in kidneys of TGF-β(1)-transgenic mice than wild-type mice. miR-21-deficient TGF-β(1)-transgenic mice showed increased proteinuria and glomerular extracellular matrix deposition and fewer podocytes per glomerular tuft compared with miR-21 wild-type TGF-β(1)-transgenic littermates. Similarly, miR-21 expression was increased in streptozotocin-induced diabetic mice, and loss of miR-21 in these mice was associated with increased albuminuria, podocyte depletion, and mesangial expansion. In cultured podocytes, inhibition of miR-21 was accompanied by increases in the rate of cell death, TGF-β/Smad3-signaling activity, and expression of known proapoptotic miR-21 target genes p53, Pdcd4, Smad7, Tgfbr2, and Timp3. In American-Indian patients with diabetic nephropathy (n=48), albumin-to-creatinine ratio was positively associated with miR-21 expression in glomerular fractions (r=0.6; P<0.001) but not tubulointerstitial fractions (P=0.80). These findings suggest that miR-21 ameliorates TGF-β(1) and hyperglycemia-induced glomerular injury through repression of proapoptotic signals, thereby inhibiting podocyte loss. This finding is in contrast to observations in murine models of tubulointerstitial kidney injury but consistent with findings in cancer models. The aggravation of glomerular disease in miR-21-deficient mice and the positive association with albumin-to-creatinine ratio in patients with diabetic nephropathy support miR-21 as a feedback inhibitor of TGF-β signaling and functions. PMID:25145934

  17. MicroRNA-21 in Glomerular Injury

    PubMed Central

    Lai, Jennifer Y.; Luo, Jinghui; O’Connor, Christopher; Jing, Xiaohong; Nair, Viji; Ju, Wenjun; Randolph, Ann; Ben-Dov, Iddo Z.; Matar, Regina N.; Briskin, Daniel; Zavadil, Jiri; Nelson, Robert G.; Tuschl, Thomas; Brosius, Frank C.; Kretzler, Matthias

    2015-01-01

    TGF-β1 is a pleotropic growth factor that mediates glomerulosclerosis and podocyte apoptosis, hallmarks of glomerular diseases. The expression of microRNA-21 (miR-21) is regulated by TGF-β1, and miR-21 inhibits apoptosis in cancer cells. TGF-β1–transgenic mice exhibit accelerated podocyte loss and glomerulosclerosis. We determined that miR-21 expression increases rapidly in cultured murine podocytes after exposure to TGF-β1 and is higher in kidneys of TGF-β1–transgenic mice than wild-type mice. miR-21–deficient TGF-β1–transgenic mice showed increased proteinuria and glomerular extracellular matrix deposition and fewer podocytes per glomerular tuft compared with miR-21 wild-type TGF-β1–transgenic littermates. Similarly, miR-21 expression was increased in streptozotocin-induced diabetic mice, and loss of miR-21 in these mice was associated with increased albuminuria, podocyte depletion, and mesangial expansion. In cultured podocytes, inhibition of miR-21 was accompanied by increases in the rate of cell death, TGF-β/Smad3-signaling activity, and expression of known proapoptotic miR-21 target genes p53, Pdcd4, Smad7, Tgfbr2, and Timp3. In American-Indian patients with diabetic nephropathy (n=48), albumin-to-creatinine ratio was positively associated with miR-21 expression in glomerular fractions (r=0.6; P<0.001) but not tubulointerstitial fractions (P=0.80). These findings suggest that miR-21 ameliorates TGF-β1 and hyperglycemia-induced glomerular injury through repression of proapoptotic signals, thereby inhibiting podocyte loss. This finding is in contrast to observations in murine models of tubulointerstitial kidney injury but consistent with findings in cancer models. The aggravation of glomerular disease in miR-21–deficient mice and the positive association with albumin-to-creatinine ratio in patients with diabetic nephropathy support miR-21 as a feedback inhibitor of TGF-β signaling and functions. PMID:25145934

  18. Characterization of the rainbow trout oocyte microRNA transcriptome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNAs (miRNAs) are a class of endogenous small non-coding RNA molecules that regulate post-transcriptional expression of target genes and play important roles in animal development. The objectives of this study were to characterize the egg miRNA transcriptome and identify novel egg-specific miRN...

  19. Ontogenic expression of microRNA in bovine mammary gland

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNAs (miR) are small RNA molecules (~22 nucleotides) that are important regulators of numerous biological processes, including organ and tissue morphogenesis and function. In this capacity, most miR inhibit protein synthesis by binding to the 3’-untranslated region of targeted mRNA species. H...

  20. A 22-nt artificial microRNA mediates widespread RNA silencing in Arabidopsis

    PubMed Central

    McHale, Marcus; Eamens, Andrew L; Finnegan, E Jean; Waterhouse, Peter M

    2013-01-01

    It is known that 22-nucleotide (nt) microRNAs (miRNAs) derived from asymmetric duplexes trigger phased small-interfering RNA (phasiRNA) production from complementary targets. Here we investigate the efficacy of 22-nt artificial miRNA (amiRNA)-mediated RNA silencing relative to conventional hairpin RNA (hpRNA) and 21-nt amiRNA-mediated RNA silencing. CHALCONE SYNTHASE (CHS) was selected as a target in Arabidopsis thaliana due to the obvious and non-lethal loss of anthocyanin accumulation upon widespread RNA silencing. Over-expression of CHS in the pap1-D background facilitated visual detection of both local and systemic RNA silencing. RNA silencing was initiated in leaf tissues from hpRNA and amiRNA plant expression vectors under the control of an Arabidopsis RuBisCo small subunit 1A promoter (SSU). In this system, hpRNA expression triggered CHS silencing in most leaf tissues but not in roots or seed coats. Similarly, 21-nt amiRNA expression from symmetric miRNA/miRNA* duplexes triggered CHS silencing in all leaf tissues but not in roots or seed coats. However, 22-nt amiRNA expression from an asymmetric duplex triggered CHS silencing in all tissues, including roots and seed coats, in the majority of plant lines. This widespread CHS silencing required RNA-DEPENDENT RNA POLYMERASE6-mediated accumulation of phasiRNAs from the endogenous CHS transcript. These results demonstrate the efficacy of asymmetric 22-nt amiRNA-directed RNA silencing and associated phasiRNA production and activity, in mediating widespread RNA silencing of an endogenous target gene. Asymmetric 22-nt amiRNA-directed RNA silencing requires little modification of existing amiRNA technology and is expected to be effective in suppressing other genes and/or members of gene families. PMID:23937661

  1. MicroRNA Profiling of Primary Cutaneous Large B-Cell Lymphomas

    PubMed Central

    Koens, Lianne; Qin, Yongjun; Leung, Wai Y.; Corver, Willem E.; Jansen, Patty M.; Willemze, Rein; Vermeer, Maarten H.; Tensen, Cornelis P.

    2013-01-01

    Aberrant expression of microRNAs is widely accepted to be pathogenetically involved in nodal diffuse large B-cell lymphomas (DLBCLs). However, the microRNAs profiles of primary cutaneous large B-cell lymphomas (PCLBCLs) are not yet described. Its two main subtypes, i.e., primary cutaneous diffuse large B-cell lymphoma, leg type (PCLBCL-LT) and primary cutaneous follicle center lymphoma (PCFCL) are characterized by an activated B-cell (ABC)-genotype and a germinal center B-cell (GCB)-genotype, respectively. We performed high-throughput sequencing analysis on frozen tumor biopsies from 19 cases of PCFCL and PCLBCL-LT to establish microRNA profiles. Cluster analysis of the complete microRNome could not distinguish between the two subtypes, but 16 single microRNAs were found to be differentially expressed. Single microRNA RT-qPCR was conducted on formalin-fixed paraffin-embedded tumor biopsies of 20 additional cases, confirming higher expression of miR-9-5p, miR-31-5p, miR-129-2-3p and miR-214-3p in PCFCL as compared to PCLBCL-LT. MicroRNAs previously described to be higher expressed in ABC-type as compared to GCB-type nodal DLBCL were not differentially expressed between PCFCL and PCLBCL-LT. In conclusion, PCFCL and PCLBCL-LT differ in their microRNA profiles. In contrast to their gene expression profile, they only show slight resemblance with the microRNA profiles found in GCB- and ABC-type nodal DLBCL. PMID:24358187

  2. MicroRNA profiling of primary cutaneous large B-cell lymphomas.

    PubMed

    Koens, Lianne; Qin, Yongjun; Leung, Wai Y; Corver, Willem E; Jansen, Patty M; Willemze, Rein; Vermeer, Maarten H; Tensen, Cornelis P

    2013-01-01

    Aberrant expression of microRNAs is widely accepted to be pathogenetically involved in nodal diffuse large B-cell lymphomas (DLBCLs). However, the microRNAs profiles of primary cutaneous large B-cell lymphomas (PCLBCLs) are not yet described. Its two main subtypes, i.e., primary cutaneous diffuse large B-cell lymphoma, leg type (PCLBCL-LT) and primary cutaneous follicle center lymphoma (PCFCL) are characterized by an activated B-cell (ABC)-genotype and a germinal center B-cell (GCB)-genotype, respectively. We performed high-throughput sequencing analysis on frozen tumor biopsies from 19 cases of PCFCL and PCLBCL-LT to establish microRNA profiles. Cluster analysis of the complete microRNome could not distinguish between the two subtypes, but 16 single microRNAs were found to be differentially expressed. Single microRNA RT-qPCR was conducted on formalin-fixed paraffin-embedded tumor biopsies of 20 additional cases, confirming higher expression of miR-9-5p, miR-31-5p, miR-129-2-3p and miR-214-3p in PCFCL as compared to PCLBCL-LT. MicroRNAs previously described to be higher expressed in ABC-type as compared to GCB-type nodal DLBCL were not differentially expressed between PCFCL and PCLBCL-LT. In conclusion, PCFCL and PCLBCL-LT differ in their microRNA profiles. In contrast to their gene expression profile, they only show slight resemblance with the microRNA profiles found in GCB- and ABC-type nodal DLBCL. PMID:24358187

  3. MicroRNA in intervertebral disc degeneration.

    PubMed

    Li, Zheng; Yu, Xin; Shen, Jianxiong; Chan, Matthew T V; Wu, William Ka Kei

    2015-06-01

    Aetiology of intervertebral disc degeneration (IDD) is complex, with genetic, developmental, biochemical and biomechanical factors contributing to the disease process. It is becoming obvious that epigenetic processes influence evolution of IDD as strongly as the genetic background. Deregulated phenotypes of nucleus pulposus cells, including differentiation, migration, proliferation and apoptosis, are involved in all stages of progression of human IDD. Non-coding RNAs, including microRNAs, have recently been recognized as important regulators of gene expression. Research into roles of microRNAs in IDD has been very active over the past 5 years. Our review summarizes current research enlightenment towards understanding roles of microRNAs in regulating nucleus pulposus cell functions in IDD. These exciting findings support the notion that specific modulation of microRNAs may represent an attractive approach for management of IDD. PMID:25736871

  4. Clamping of RNA with PNA enables targeting of microRNA.

    PubMed

    Ghidini, Alice; Bergquist, Helen; Murtola, Merita; Punga, Tanel; Zain, Rula; Strömberg, Roger

    2016-06-21

    To be able to target microRNAs also at stages where these are in a double stranded or hairpin form we have studied BisPNA designed to clamp the target and give sufficient affinity to allow for strand invasion. We show that BisPNA complexes are more stable with RNA than with DNA. In addition, 24-mer BisPNA (AntimiR) constructs form complexes with a hairpin RNA that is a model of the microRNA miR-376b, suggesting that PNA-clamping may be an effective way of targeting microRNAs. PMID:27203783

  5. MicroRNA regulation of airway smooth muscle function.

    PubMed

    Sun, Maoyun; Lu, Quan

    2016-06-01

    Airway smooth muscle (ASM) controls airway narrowing and plays a pivotal role in the pathogenesis of asthma. MicroRNAs are small yet powerful gene tuners that regulate diverse cellular processes. Recent studies have demonstrated the versatile role of microRNAs in regulating multiple ASM phenotypes that are critically involved in asthma pathogenesis. These ASM phenotypes include proliferation, cell size, chemokine secretion, and contractility. Here we review microRNA-mediated regulation of ASM functions and discuss the potential of microRNAs as a novel class of therapeutic targets to improve ASM function for asthma therapy. PMID:26812790

  6. microRNAs in the Same Clusters Evolve to Coordinately Regulate Functionally Related Genes

    PubMed Central

    Wang, Yirong; Luo, Junjie; Zhang, Hong; Lu, Jian

    2016-01-01

    MicroRNAs (miRNAs) are endogenously expressed small noncoding RNAs. The genomic locations of animal miRNAs are significantly clustered in discrete loci. We found duplication and de novo formation were important mechanisms to create miRNA clusters and the clustered miRNAs tend to be evolutionarily conserved. We proposed a “functional co-adaptation” model to explain how clustering helps newly emerged miRNAs survive and develop functions. We presented evidence that abundance of miRNAs in the same clusters were highly correlated and those miRNAs exerted cooperative repressive effects on target genes in human tissues. By transfecting miRNAs into human and fly cells and extensively profiling the transcriptome alteration with deep-sequencing, we further demonstrated the functional co-adaptation between new and old miRNAs in the miR-17–92 cluster. Our population genomic analysis suggest that positive Darwinian selection might be the driving force underlying the formation and evolution of miRNA clustering. Our model provided novel insights into mechanisms and evolutionary significance of miRNA clustering. PMID:27189568

  7. microRNAs in the Same Clusters Evolve to Coordinately Regulate Functionally Related Genes.

    PubMed

    Wang, Yirong; Luo, Junjie; Zhang, Hong; Lu, Jian

    2016-09-01

    MicroRNAs (miRNAs) are endogenously expressed small noncoding RNAs. The genomic locations of animal miRNAs are significantly clustered in discrete loci. We found duplication and de novo formation were important mechanisms to create miRNA clusters and the clustered miRNAs tend to be evolutionarily conserved. We proposed a "functional co-adaptation" model to explain how clustering helps newly emerged miRNAs survive and develop functions. We presented evidence that abundance of miRNAs in the same clusters were highly correlated and those miRNAs exerted cooperative repressive effects on target genes in human tissues. By transfecting miRNAs into human and fly cells and extensively profiling the transcriptome alteration with deep-sequencing, we further demonstrated the functional co-adaptation between new and old miRNAs in the miR-17-92 cluster. Our population genomic analysis suggest that positive Darwinian selection might be the driving force underlying the formation and evolution of miRNA clustering. Our model provided novel insights into mechanisms and evolutionary significance of miRNA clustering. PMID:27189568

  8. Biogenesis of Y RNA-derived small RNAs is independent of the microRNA pathway.

    PubMed

    Nicolas, Francisco Esteban; Hall, Adam E; Csorba, Tibor; Turnbull, Carly; Dalmay, Tamas

    2012-04-24

    Y RNAs are approximately 100 nucleotide long conserved cytoplasmic non-coding RNAs, which produce smaller RNA fragments during apoptosis. Here we show that these smaller RNA molecules are also produced in non-stressed cells and in a range of human cancerous and non-cancerous cell types. Recent reports have speculated that the cleavage products of Y RNAs enter the microRNA pathway. We tested this hypothesis and found that Y5 and Y3 RNA fragments are Dicer independent, they are in different complexes than microRNAs and that they are not co-immunoprecipitated with Ago2. Therefore we conclude that Y RNA fragments do not enter the microRNA pathway. PMID:22575660

  9. A Complex Genome-MicroRNA Interplay in Human Mitochondria

    PubMed Central

    Shinde, Santosh; Bhadra, Utpal

    2015-01-01

    Small noncoding regulatory RNA exist in wide spectrum of organisms ranging from prokaryote bacteria to humans. In human, a systematic search for noncoding RNA is mainly limited to the nuclear and cytosolic compartments. To investigate whether endogenous small regulatory RNA are present in cell organelles, human mitochondrial genome was also explored for prediction of precursor microRNA (pre-miRNA) and mature miRNA (miRNA) sequences. Six novel miRNA were predicted from the organelle genome by bioinformatics analysis. The structures are conserved in other five mammals including chimp, orangutan, mouse, rat, and rhesus genome. Experimentally, six human miRNA are well accumulated or deposited in human mitochondria. Three of them are expressed less prominently in Northern analysis. To ascertain their presence in human skeletal muscles, total RNA was extracted from enriched mitochondria by an immunomagnetic method. The expression of six novel pre-miRNA and miRNA was confirmed by Northern blot analysis; however, low level of remaining miRNA was found by sensitive Northern analysis. Their presence is further confirmed by real time RT-PCR. The six miRNA find their multiple targets throughout the human genome in three different types of software. The luciferase assay was used to confirm that MT-RNR2 gene was the potential target of hsa-miR-mit3 and hsa-miR-mit4. PMID:25695052

  10. DIANA-microT web server: elucidating microRNA functions through target prediction.

    PubMed

    Maragkakis, M; Reczko, M; Simossis, V A; Alexiou, P; Papadopoulos, G L; Dalamagas, T; Giannopoulos, G; Goumas, G; Koukis, E; Kourtis, K; Vergoulis, T; Koziris, N; Sellis, T; Tsanakas, P; Hatzigeorgiou, A G

    2009-07-01

    Computational microRNA (miRNA) target prediction is one of the key means for deciphering the role of miRNAs in development and disease. Here, we present the DIANA-microT web server as the user interface to the DIANA-microT 3.0 miRNA target prediction algorithm. The web server provides extensive information for predicted miRNA:target gene interactions with a user-friendly interface, providing extensive connectivity to online biological resources. Target gene and miRNA functions may be elucidated through automated bibliographic searches and functional information is accessible through Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. The web server offers links to nomenclature, sequence and protein databases, and users are facilitated by being able to search for targeted genes using different nomenclatures or functional features, such as the genes possible involvement in biological pathways. The target prediction algorithm supports parameters calculated individually for each miRNA:target gene interaction and provides a signal-to-noise ratio and a precision score that helps in the evaluation of the significance of the predicted results. Using a set of miRNA targets recently identified through the pSILAC method, the performance of several computational target prediction programs was assessed. DIANA-microT 3.0 achieved there with 66% the highest ratio of correctly predicted targets over all predicted targets. The DIANA-microT web server is freely available at www.microrna.gr/microT. PMID:19406924

  11. Identification of microRNA Genes in Three Opisthorchiids

    PubMed Central

    Ovchinnikov, Vladimir Y.; Afonnikov, Dmitry A.; Vasiliev, Gennady V.; Kashina, Elena V.; Sripa, Banchob; Mordvinov, Viacheslav A.; Katokhin, Alexey V.

    2015-01-01

    Background Opisthorchis felineus, O. viverrini, and Clonorchis sinensis (family Opisthorchiidae) are parasitic flatworms that pose a serious threat to humans in some countries and cause opisthorchiasis/clonorchiasis. Chronic disease may lead to a risk of carcinogenesis in the biliary ducts. MicroRNAs (miRNAs) are small noncoding RNAs that control gene expression at post-transcriptional level and are implicated in the regulation of various cellular processes during the parasite- host interplay. However, to date, the miRNAs of opisthorchiid flukes, in particular those essential for maintaining their complex biology and parasitic mode of existence, have not been satisfactorily described. Methodology/Principal Findings Using a SOLiD deep sequencing-bioinformatic approach, we identified 43 novel and 18 conserved miRNAs for O. felineus (miracidia, metacercariae and adult worms), 20 novel and 16 conserved miRNAs for O. viverrini (adult worms), and 33 novel and 18 conserved miRNAs for C. sinensis (adult worms). The analysis of the data revealed differences in the expression level of conserved miRNAs among the three species and among three the developmental stages of O. felineus. Analysis of miRNA genes revealed two gene clusters, one cluster-like region and one intronic miRNA in the genome. The presence and structure of the two gene clusters were validated using a PCR-based approach in the three flukes. Conclusions This study represents a comprehensive description of miRNAs in three members of the family Opistorchiidae, significantly expands our knowledge of miRNAs in multicellular parasites and provides a basis for understanding the structural and functional evolution of miRNAs in these metazoan parasites. Results of this study also provides novel resources for deeper understanding the complex parasite biology, for further research on the pathogenesis and molecular events of disease induced by the liver flukes. The present data may also facilitate the development of novel

  12. Heart Structure-Specific Transcriptomic Atlas Reveals Conserved microRNA-mRNA Interactions

    PubMed Central

    Vacchi-Suzzi, Caterina; Hahne, Florian; Scheubel, Philippe; Marcellin, Magali; Dubost, Valerie; Westphal, Magdalena; Boeglen, Catherine; Büchmann-Møller, Stine; Cheung, Ming Sin; Cordier, André; De Benedetto, Christopher; Deurinck, Mark; Frei, Moritz; Moulin, Pierre; Oakeley, Edward; Grenet, Olivier; Grevot, Armelle; Stull, Robert; Theil, Diethilde; Moggs, Jonathan G.; Marrer, Estelle; Couttet, Philippe

    2013-01-01

    MicroRNAs are short non-coding RNAs that regulate gene expression at the post-transcriptional level and play key roles in heart development and cardiovascular diseases. Here, we have characterized the expression and distribution of microRNAs across eight cardiac structures (left and right ventricles, apex, papillary muscle, septum, left and right atrium and valves) in rat, Beagle dog and cynomolgus monkey using microRNA sequencing. Conserved microRNA signatures enriched in specific heart structures across these species were identified for cardiac valve (miR-let-7c, miR-125b, miR-127, miR-199a-3p, miR-204, miR-320, miR-99b, miR-328 and miR-744) and myocardium (miR-1, miR-133b, miR-133a, miR-208b, miR-30e, miR-499-5p, miR-30e*). The relative abundance of myocardium-enriched (miR-1) and valve-enriched (miR-125b-5p and miR-204) microRNAs was confirmed using in situ hybridization. MicroRNA-mRNA interactions potentially relevant for cardiac functions were explored using anti-correlation expression analysis and microRNA target prediction algorithms. Interactions between miR-1/Timp3, miR-125b/Rbm24, miR-204/Tgfbr2 and miR-208b/Csnk2a2 were identified and experimentally investigated in human pulmonary smooth muscle cells and luciferase reporter assays. In conclusion, we have generated a high-resolution heart structure-specific mRNA/microRNA expression atlas for three mammalian species that provides a novel resource for investigating novel microRNA regulatory circuits involved in cardiac molecular physiopathology. PMID:23300973

  13. Viral microRNA genomics and target validation

    PubMed Central

    Ziegelbauer, Joseph M.

    2014-01-01

    A subset of viruses express their own microRNAs (miRNAs) and one way to understand the functions of these microRNAs is to identify the targets of these miRNAs. Sequence analysis and mRNA expression profiling were some of the first techniques to identify targets of viral miRNAs. More recently, proteomics and sequencing of RNA by crosslinking and immunoprecipitation (CLIP) methods have been insightful and discovered many miRNA targets that may be missed using other methods. We are now at a point where numerous validated miRNA targets have been described and integration of these genomic datasets will provide a richer understanding of miRNA targeting and viral infection, persistence, and pathogenesis. PMID:24763063

  14. MicroRNA-regulated viral vectors for gene therapy.

    PubMed

    Geisler, Anja; Fechner, Henry

    2016-05-20

    Safe and effective gene therapy approaches require targeted tissue-specific transfer of a therapeutic transgene. Besides traditional approaches, such as transcriptional and transductional targeting, microRNA-dependent post-transcriptional suppression of transgene expression has been emerging as powerful new technology to increase the specificity of vector-mediated transgene expression. MicroRNAs are small non-coding RNAs and often expressed in a tissue-, lineage-, activation- or differentiation-specific pattern. They typically regulate gene expression by binding to imperfectly complementary sequences in the 3' untranslated region (UTR) of the mRNA. To control exogenous transgene expression, tandem repeats of artificial microRNA target sites are usually incorporated into the 3' UTR of the transgene expression cassette, leading to subsequent degradation of transgene mRNA in cells expressing the corresponding microRNA. This targeting strategy, first shown for lentiviral vectors in antigen presenting cells, has now been used for tissue-specific expression of vector-encoded therapeutic transgenes, to reduce immune response against the transgene, to control virus tropism for oncolytic virotherapy, to increase safety of live attenuated virus vaccines and to identify and select cell subsets for pluripotent stem cell therapies, respectively. This review provides an introduction into the technical mechanism underlying microRNA-regulation, highlights new developments in this field and gives an overview of applications of microRNA-regulated viral vectors for cardiac, suicide gene cancer and hematopoietic stem cell therapy, as well as for treatment of neurological and eye diseases. PMID:27226955

  15. MicroRNA-regulated viral vectors for gene therapy

    PubMed Central

    Geisler, Anja; Fechner, Henry

    2016-01-01

    Safe and effective gene therapy approaches require targeted tissue-specific transfer of a therapeutic transgene. Besides traditional approaches, such as transcriptional and transductional targeting, microRNA-dependent post-transcriptional suppression of transgene expression has been emerging as powerful new technology to increase the specificity of vector-mediated transgene expression. MicroRNAs are small non-coding RNAs and often expressed in a tissue-, lineage-, activation- or differentiation-specific pattern. They typically regulate gene expression by binding to imperfectly complementary sequences in the 3’ untranslated region (UTR) of the mRNA. To control exogenous transgene expression, tandem repeats of artificial microRNA target sites are usually incorporated into the 3’ UTR of the transgene expression cassette, leading to subsequent degradation of transgene mRNA in cells expressing the corresponding microRNA. This targeting strategy, first shown for lentiviral vectors in antigen presenting cells, has now been used for tissue-specific expression of vector-encoded therapeutic transgenes, to reduce immune response against the transgene, to control virus tropism for oncolytic virotherapy, to increase safety of live attenuated virus vaccines and to identify and select cell subsets for pluripotent stem cell therapies, respectively. This review provides an introduction into the technical mechanism underlying microRNA-regulation, highlights new developments in this field and gives an overview of applications of microRNA-regulated viral vectors for cardiac, suicide gene cancer and hematopoietic stem cell therapy, as well as for treatment of neurological and eye diseases. PMID:27226955

  16. Assessing an improved protocol for plasma microRNA extraction.

    PubMed

    Moret, Inés; Sánchez-Izquierdo, Dolors; Iborra, Marisa; Tortosa, Luis; Navarro-Puche, Ana; Nos, Pilar; Cervera, José; Beltrán, Belén

    2013-01-01

    The first step in biomarkers discovery is to identify the best protocols for their purification and analysis. This issue is critical when considering peripheral blood samples (plasma and serum) that are clinically interesting but meet several methodological problems, mainly complexity and low biomarker concentration. Analysis of small molecules, such as circulating microRNAs, should overcome these disadvantages. The present study describes an optimal RNA extraction method of microRNAs from human plasma samples. Different reagents and commercially available kits have been analyzed, identifying also the best pre-analytical conditions for plasma isolation. Between all of them, the column-based approaches were shown to be the most effective. In this context, miRNeasy Serum/Plasma Kit (from Qiagen) rendered more concentrated RNA, that was better suited for microarrays studies and did not require extra purification steps for sample concentration and purification than phenol based extraction methods. We also present evidences that the addition of low doses of an RNA carrier before starting the extraction process improves microRNA purification while an already published carrier dose can result in significant bias over microRNA profiles. Quality controls for best protocol selection were developed by spectrophotometry measurement of contaminants and microfluidics electrophoresis (Agilent 2100 Bioanalyzer) for RNA integrity. Selected donor and patient plasma samples and matched biopsies were tested by Affymetrix microarray technology to compare differentially expressed microRNAs. In summary, this study defines an optimized protocol for microRNA purification from human blood samples, increasing the performance of assays and shedding light over the best way to discover and use these biomarkers in clinical practice. PMID:24376572

  17. A conserved RNA polymerase III promoter required for gammaherpesvirus TMER transcription and microRNA processing

    PubMed Central

    Diebel, Kevin W.; Claypool, David J.; van Dyk, Linda F.

    2014-01-01

    Canonical RNA polymerase III (pol III) type 2 promoters contain a single A and B box and are well documented for their role in tRNA and SINE transcription in eukaryotic cells. The genome of Murid herpesvirus 4 (MuHV-4) contains eight polycistronic tRNA-microRNA encoded RNA (TMER) genes that are transcribed from a RNA pol III type 2-like promoter containing triplicated A box elements. Here, we demonstrate that the triplicated A box sequences are required in their entirety to produce functional MuHV-4 miRNAs. We also identify that these RNA pol III type 2-like promoters are conserved in eukaryotic genomes. Human and mouse predicted tRNA genes containing these promoters also show enrichment of alternative RNA pol III transcription termination sequences and are predicted to give rise to longer tRNA primary transcripts. PMID:24747015

  18. Early lethality of shRNA-transgenic pigs due to saturation of microRNA pathways* #

    PubMed Central

    Dai, Zhen; Wu, Rong; Zhao, Yi-cheng; Wang, Kan-kan; Huang, Yong-ye; Yang, Xin; Xie, Zi-cong; Tu, Chang-chun; Ouyang, Hong-sheng; Wang, Tie-dong; Pang, Da-xin

    2014-01-01

    RNA interference (RNAi) is considered as a potential modality for clinical treatment and anti-virus animal breeding. Here, we investigate the feasibility of inhibiting classical swine fever virus (CSFV) replication by short hairpin RNA (shRNA) in vitro and in vivo. We generate four different shRNA-positive clonal cells and two types of shRNA-transgenic pigs. CSFV could be effectively inhibited in shRNA-positive clonal cells and tail tip fibroblasts of shRNA-transgenic pigs. Unexpectedly, an early lethality due to shRNA is observed in these shRNA-transgenic pigs. With further research on shRNA-positive clonal cells and transgenic pigs, we report a great induction of interferon (IFN)-responsive genes in shRNA-positive clonal cells, altered levels of endogenous microRNAs (miRNA), and their processing enzymes in shRNA-positive cells. What is more, abnormal expressions of miRNAs and their processing enzymes are also observed in the livers of shRNA-transgenic pigs, indicating saturation of miRNA/shRNA pathways induced by shRNA. In addition, we investigate the effects of shRNAs on the development of somatic cell nuclear transfer (SCNT) embryos. These results show that shRNA causes adverse effects in vitro and in vivo and shRNA-induced disruption of the endogenous miRNA pathway may lead to the early lethality of shRNA-transgenic pigs. We firstly report abnormalities of the miRNA pathway in shRNA-transgenic animals, which may explain the early lethality of shRNA-transgenic pigs and has important implications for shRNA-transgenic animal preparation. PMID:24793764

  19. High-Throughput Functional MicroRNAs Profiling by Recombinant AAV-Based MicroRNA Sensor Arrays

    PubMed Central

    Liu, Xuerong; Wang, Gang; Dong, Zheyue; Shen, Wei; Zheng, Gang; Lu, Jianxin; Chen, Jinzhong; Wang, Yue; Wu, Zhijian; Wu, Xiaobing

    2012-01-01

    Background microRNAs (miRNAs) are small and non-coding RNAs which play critical roles in physiological and pathological processes. A number of methods have been established to detect and quantify miRNA expression. However, method for high-throughput miRNA function detection is still lacking. Principal Findings We describe an adeno-associated virus (AAV) vector-based microRNA (miRNA) sensor (Asensor) array for high-throughput functional miRNA profiling. Each Asensor contains a Gaussia luciferase (Gluc) and a firefly luciferase (Fluc) expression cassette to sense functional miRNA and to serve as an internal control respectively. Using this array, we acquired functional profiles of 115 miRNAs for 12 cell lines and found “functional miRNA signatures” for several specific cell lines. The activities of specific miRNAs including the let-7 family, miR-17-92 cluster, miR-221, and miR-222 in HEK 293 cells were compared with their expression levels determined by quantitative reverse transcriptase polymerase chain reaction (QRT-PCR). We also demonstrate two other practical applications of the array, including a comparison of the miRNA activity between HEK293 and HEK293T cells and the ability to monitor miRNA activity changes in K562 cells treated with 12-O-tetradecanoylphorbol-13-acetate (TPA). Conclusions/Significance Our approach has potential applications in the identification of cell types, the characterization of biological and pathological processes, and the evaluation of responses to interventions. PMID:22242174

  20. MicroRNA profiling in the malignant progression of gliomas

    NASA Astrophysics Data System (ADS)

    Stupak, E. V.; Veryaskina, Yu. A.; Titov, S. E.; Achmerova, L. G.; Stupak, V. V.; Ivanov, M. K.; Zhimulev, I. F.; Kolesnikov, N. N.

    2016-08-01

    Wealth of data indicates that microRNAs (miRNAs) are directly involved in carcinogenesis and that miRNA can, on their own, act as diagnostic and prognostic markers for various types of cancers, including gliomas. The aim of this study was to conduct a comparative analysis of expression profile for 10 microRNAs (miR-124, -125b, -16, -181b, -191, -21, -221, -223, -31, and -451) in surgical specimens of various hystotypes of glioimatissues vs adjacent normal tissues from the same patient (n = 77). The study identified specific microRNA expression profiles for different histotypes of tumors that are related to their degree of malignancy. We have outlined approaches to development of miRNA-based diagnostic and prognostic panel, which may be used to compensate for the lack of appropriate screening methods.

  1. Analysis of Plasma microRNA Associated with Hemolysis.

    PubMed

    Shkurnikov, M Yu; Knyazev, E N; Fomicheva, K A; Mikhailenko, D S; Nyushko, K M; Saribekyan, E K; Samatov, T R; Alekseev, B Ya

    2016-04-01

    We analyzed the effect of hemolysis on microRNA profi le of blood plasma. It was found that hemolysis of ~0.05% erythrocytes in a sample signifi cantly affected the concentration of 9 microRNA: hsa-miR-486-5p, hsa-miR-16-5p, hsa-miR-451a, hsa-miR-106a-5p, hsa-miR-17-5p, hsa-miR-93-5p, hsa-miR-20a-5p, hsa-miR-107, and hsa-miR-20b-5p. The effect of hemolysis on plasma content of miR-17 family microRNA was demonstrated. PMID:27165077

  2. The locus of microRNA-10b

    PubMed Central

    Biagioni, Francesca; Bossel Ben-Moshe, Noa; Fontemaggi, Giulia; Yarden, Yosef; Domany, Eytan; Blandino, Giovanni

    2013-01-01

    Contemporary microRNA research has led to significant advances in our understanding of the process of tumorigenesis. MicroRNAs participate in different events of a cancer cell’s life, through their ability to target hundreds of putative transcripts involved in almost every cellular function, including cell cycle, apoptosis, and differentiation. The relevance of these small molecules is even more evident in light of the emerging linkage between their expression and both prognosis and clinical outcome of many types of human cancers. This identifies microRNAs as potential therapeutic modifiers of cancer phenotypes. From this perspective, we overview here the miR-10b locus and its involvement in cancer, focusing on its role in the establishment (miR-10b*) and spreading (miR-10b) of breast cancer. We conclude that targeting the locus of microRNA 10b holds great potential for cancer treatment. PMID:23839045

  3. MicroRNA, Nutrition, and Cancer Prevention1

    PubMed Central

    Ross, Sharon A.; Davis, Cindy D.

    2011-01-01

    MicroRNA (miRNA) are small noncoding RNA molecules that are involved in post-transcriptional gene silencing. Alterations in miRNA expression are observed in and may underlie many different human diseases, including cancer. In fact, miRNA have been shown to affect the hallmarks of cancer, including sustaining proliferative signaling, evading growth suppressors, resisting cell death, enabling replicative immortality, inducing angiogenesis, and activating invasion and metastasis. Genetic and epigenetic alterations may explain aberrant miRNA expression in cancer cells and may also contribute to cancer risk. It is now thought that by circulating through the bloodstream, miRNA can exert their effects at distant sites as well as within the cells of origin. Recent evidence suggests that nutrients and other bioactive food components protect against cancer through modulation of miRNA expression. Moreover, dietary factors have been shown to modify miRNA expression and their mRNA targets in various cancer processes, including apoptosis, cell cycle regulation, differentiation, inflammation, angiogenesis, and metastasis as well as pathways in stress response. Herein, we provide a brief overview of dietary modulation of miRNA expression and its potential role in cancer prevention. Understanding the affect of dietary factors on miRNA expression and function may provide insight on prevention strategies to reduce the burden of cancer. PMID:22332090

  4. Changes of microRNA profile and microRNA-mRNA regulatory network in bones of ovariectomized mice.

    PubMed

    An, Jee Hyun; Ohn, Jung Hun; Song, Jung Ah; Yang, Jae-Yeon; Park, Hyojung; Choi, Hyung Jin; Kim, Sang Wan; Kim, Seong Yeon; Park, Woog-Yang; Shin, Chan Soo

    2014-03-01

    Growing evidence shows the possibility of a role of microRNAs (miRNA) in regulating bone mass. We investigated the change of miRNAs and mRNA expression profiles in bone tissue in an ovariectomized mice model and evaluated the regulatory mechanism of bone mass mediated by miRNAs in an estrogen-deficiency state. Eight-week-old female C3H/HeJ mice underwent ovariectomy (OVX) or sham operation (Sham-op), and their femur and tibia were harvested to extract total bone RNAs after 4 weeks for microarray analysis. Eight miRNAs (miR-127, -133a, -133a*, -133b, -136, -206, -378, -378*) were identified to be upregulated after OVX, whereas one miRNA (miR-204) was downregulated. Concomitant analysis of mRNA microarray revealed that 658 genes were differentially expressed between OVX and Sham-op mice. Target prediction of differentially expressed miRNAs identified potential targets, and integrative analysis using the mRNA microarray results showed that PPARγ and CREB pathways are activated in skeletal tissues after ovariectomy. Among the potential candidates of miRNA, we further studied the role of miR-127 in vitro, which exhibited the greatest changes after OVX. We also studied the effects of miR-136, which has not been studied in the context of bone mass regulation. Transfection of miR-127 inhibitor has enhanced osteoblastic differentiation in UAMS-32 cells as measured by alkaline phosphatase activities and mRNA expression of osteoblast-specific genes, whereas miR-136 precursor has inhibited osteoblastic differentiation. Furthermore, transfection of both miR-127 and miR-136 inhibitors enhanced the osteocyte-like morphological changes and survival in MLO-Y4 cells, whereas precursors of miR-127 and -136 have aggravated dexamethasone-induced cell death. Both of the precursors enhanced osteoclastic differentiation in bone marrow macrophages, indicating that both miR-127 and -136 are negatively regulating bone mass. Taken together, these results suggest a novel insight into the

  5. RNA Polymerase II cluster dynamics predict mRNA output in living cells

    PubMed Central

    Cho, Won-Ki; Jayanth, Namrata; English, Brian P; Inoue, Takuma; Andrews, J Owen; Conway, William; Grimm, Jonathan B; Spille, Jan-Hendrik; Lavis, Luke D; Lionnet, Timothée; Cisse, Ibrahim I

    2016-01-01

    Protein clustering is a hallmark of genome regulation in mammalian cells. However, the dynamic molecular processes involved make it difficult to correlate clustering with functional consequences in vivo. We developed a live-cell super-resolution approach to uncover the correlation between mRNA synthesis and the dynamics of RNA Polymerase II (Pol II) clusters at a gene locus. For endogenous β-actin genes in mouse embryonic fibroblasts, we observe that short-lived (~8 s) Pol II clusters correlate with basal mRNA output. During serum stimulation, a stereotyped increase in Pol II cluster lifetime correlates with a proportionate increase in the number of mRNAs synthesized. Our findings suggest that transient clustering of Pol II may constitute a pre-transcriptional regulatory event that predictably modulates nascent mRNA output. DOI: http://dx.doi.org/10.7554/eLife.13617.001 PMID:27138339

  6. microRNAs and microRNA Targets Involved in Alfalfa Stem Development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To examine the possible involvement of microRNAs (miRNAs) in alfalfa stem development, we hybridized 32P-labeled total miRNA purified from elongating and post-elongation stem internodes (ES and PES, respectively) of alfalfa clone 252 to a miRNA-macroarray that contained a total of 70 reference anti-...

  7. Practical Aspects of microRNA Target Prediction

    PubMed Central

    Witkos, T.M; Koscianska, E; Krzyzosiak, W.J

    2011-01-01

    microRNAs (miRNAs) are endogenous non-coding RNAs that control gene expression at the posttranscriptional level. These small regulatory molecules play a key role in the majority of biological processes and their expression is also tightly regulated. Both the deregulation of genes controlled by miRNAs and the altered miRNA expression have been linked to many disorders, including cancer, cardiovascular, metabolic and neurodegenerative diseases. Therefore, it is of particular interest to reliably predict potential miRNA targets which might be involved in these diseases. However, interactions between miRNAs and their targets are complex and very often there are numerous putative miRNA recognition sites in mRNAs. Many miRNA targets have been computationally predicted but only a limited number of these were experimentally validated. Although a variety of miRNA target prediction algorithms are available, results of their application are often inconsistent. Hence, finding a functional miRNA target is still a challenging task. In this review, currently available and frequently used computational tools for miRNA target prediction, i.e., PicTar, TargetScan, DIANA-microT, miRanda, rna22 and PITA are outlined and various practical aspects of miRNA target analysis are extensively discussed. Moreover, the performance of three algorithms (PicTar, TargetScan and DIANA-microT) is both demonstrated and evaluated by performing an in-depth analysis of miRNA interactions with mRNAs derived from genes triggering hereditary neurological disorders known as trinucleotide repeat expansion diseases (TREDs), such as Huntington’s disease (HD), a number of spinocerebellar ataxias (SCAs), and myotonic dystrophy type 1 (DM1). PMID:21342132

  8. Small Molecule Chemical Probes of MicroRNA Function

    PubMed Central

    Velagapudi, Sai Pradeep; Vummidi, Balayeshwanth R.; Disney, Matthew D.

    2015-01-01

    MicroRNAs (miRNAs) are small, non-coding RNAs that control protein expression. Aberrant miRNA expression has been linked to various human diseases, and thus miRNAs have been explored as diagnostic markers and therapeutic targets. Although it is challenging to target RNA with small molecules in general, there have been successful campaigns that have identified small molecule modulators of miRNA function by targeting various pathways. For example, small molecules that modulate transcription and target nuclease processing sites in miRNA precursors have been identified. Herein, we describe challenges in developing chemical probes that target miRNAs and highlight aspects of miRNA cellular biology elucidated by using small molecule chemical probes. We expect that this area will expand dramatically in the near future as strides are made to understand small molecule recognition of RNA from a fundamental perspective. PMID:25500006

  9. Small molecule chemical probes of microRNA function.

    PubMed

    Velagapudi, Sai Pradeep; Vummidi, Balayeshwanth R; Disney, Matthew D

    2015-02-01

    MicroRNAs (miRNAs) are small, non-coding RNAs that control protein expression. Aberrant miRNA expression has been linked to various human diseases, and thus miRNAs have been explored as diagnostic markers and therapeutic targets. Although it is challenging to target RNA with small molecules in general, there have been successful campaigns that have identified small molecule modulators of miRNA function by targeting various pathways. For example, small molecules that modulate transcription and target nuclease processing sites in miRNA precursors have been identified. Herein, we describe challenges in developing chemical probes that target miRNAs and highlight aspects of miRNA cellular biology elucidated by using small molecule chemical probes. We expect that this area will expand dramatically in the near future as progress is made in understanding small molecule recognition of RNA. PMID:25500006

  10. The Whereabouts of microRNA Actions: Cytoplasm and Beyond.

    PubMed

    Leung, Anthony K L

    2015-10-01

    MicroRNAs (miRNAs) are a conserved class of approximately 22 nucleotide (nt) short noncoding RNAs that normally silence gene expression via translational repression and/or degradation of targeted mRNAs in plants and animals. Identifying the whereabouts of miRNAs potentially informs miRNA functions, some of which are perhaps specialized to specific cellular compartments. In this review, the significance of miRNA localizations in the cytoplasm, including those at RNA granules and endomembranes, and the export of miRNAs to extracellular space will be discussed. How miRNA localizations and functions are regulated by protein modifications on the core miRNA-binding protein Argonaute (AGO) during normal and stress conditions will be explored, and in conclusion new AGO partners, non-AGO miRNA-binding proteins, and the emergent understanding of miRNAs found in the nucleoplasm, nucleoli, and mitochondria will be discussed. PMID:26410406

  11. Therapeutic evaluation of microRNA-15a and microRNA-16 in ovarian cancer

    PubMed Central

    Dhar Dwivedi, Shailendra Kumar; Mustafi, Soumyajit Banerjee; Mangala, Lingegowda S.; Jiang, Dahai; Pradeep, Sunila; Rodriguez-Aguayo, Cristian; Ling, Hui; Ivan, Cristina; Mukherjee, Priyabrata; Calin, George A.; Lopez-Berestein, Gabriel; Sood, Anil K.; Bhattacharya, Resham

    2016-01-01

    Treatment of chemo-resistant ovarian cancer (OvCa) remains clinically challenging and there is a pressing need to identify novel therapeutic strategies. Here we report that multiple mechanisms that promote OvCa progression and chemo-resistance could be inhibited by ectopic expression of miR-15a and miR-16. Significant correlations between low expression of miR-16, high expression of BMI1 and shortened overall survival (OS) were noted in high grade serous (HGS) OvCa patients upon analysis of The Cancer Genome Atlas (TCGA). Targeting BMI1, in vitro with either microRNA reduced clonal growth of OvCa cells. Additionally, epithelial to mesenchymal transition (EMT) as well as expression of the cisplatin transporter ATP7B were inhibited by miR-15a and miR-16 resulting in decreased degradation of the extra-cellular matrix and enhanced sensitization of OvCa cells to cisplatin. Nanoliposomal delivery of the miR-15a and miR-16 combination, in a pre-clinical chemo-resistant orthotopic mouse model of OvCa, demonstrated striking reduction in tumor burden compared to cisplatin alone. Thus, with the advent of miR replacement therapy some of which are in Phase 2 clinical trials, miR-15a and miR-16 represent novel ammunition in the anti-OvCa arsenal. PMID:26918603

  12. Common features of microRNA target prediction tools.

    PubMed

    Peterson, Sarah M; Thompson, Jeffrey A; Ufkin, Melanie L; Sathyanarayana, Pradeep; Liaw, Lucy; Congdon, Clare Bates

    2014-01-01

    The human genome encodes for over 1800 microRNAs (miRNAs), which are short non-coding RNA molecules that function to regulate gene expression post-transcriptionally. Due to the potential for one miRNA to target multiple gene transcripts, miRNAs are recognized as a major mechanism to regulate gene expression and mRNA translation. Computational prediction of miRNA targets is a critical initial step in identifying miRNA:mRNA target interactions for experimental validation. The available tools for miRNA target prediction encompass a range of different computational approaches, from the modeling of physical interactions to the incorporation of machine learning. This review provides an overview of the major computational approaches to miRNA target prediction. Our discussion highlights three tools for their ease of use, reliance on relatively updated versions of miRBase, and range of capabilities, and these are DIANA-microT-CDS, miRanda-mirSVR, and TargetScan. In comparison across all miRNA target prediction tools, four main aspects of the miRNA:mRNA target interaction emerge as common features on which most target prediction is based: seed match, conservation, free energy, and site accessibility. This review explains these features and identifies how they are incorporated into currently available target prediction tools. MiRNA target prediction is a dynamic field with increasing attention on development of new analysis tools. This review attempts to provide a comprehensive assessment of these tools in a manner that is accessible across disciplines. Understanding the basis of these prediction methodologies will aid in user selection of the appropriate tools and interpretation of the tool output. PMID:24600468

  13. Identification of microRNA-mRNA interactions in atrial fibrillation using microarray expression profiles and bioinformatics analysis

    PubMed Central

    WANG, TAO; WANG, BIN

    2016-01-01

    The present study integrated microRNA (miRNA) and mRNA expression data obtained from atrial fibrillation (AF) tissues and healthy tissues, in order to identify miRNAs and target genes that may be important in the development of AF. The GSE28954 miRNA expression profile and GSE2240 mRNA gene expression profile were downloaded from the Gene Expression Omnibus. Differentially expressed miRNAs and genes (DEGs) in AF tissues, compared with in control samples, were identified and hierarchically clustered. Subsequently, differentially expressed miRNAs and DEGs were searched for in the miRecords database and TarBase, and were used to construct a regulatory network using Cytoscape. Finally, functional analysis of the miRNA-targeted genes was conducted. After data processing, 71 differentially expressed miRNAs and 390 DEGs were identified between AF and normal tissues. A total of 3,506 miRNA-mRNA pairs were selected, of which 372 were simultaneously predicted by both miRecords and TarBase, and were therefore used to construct the miRNA-mRNA regulatory network. Furthermore, 10 miRNAs and 12 targeted mRNAs were detected, which formed 14 interactive pairs. The miRNA-targeted genes were significantly enriched into 14 Gene Ontology (GO) categories, of which the most significant was gene expression regulation (GO 10468), which was associated with 7 miRNAs and 8 target genes. These results suggest that the screened miRNAs and target genes may be target molecules in AF development, and may be beneficial for the early diagnosis and future treatment of AF. PMID:27082053

  14. MicroRNA-mediated target mRNA cleavage and 3'-uridylation in human cells.

    PubMed

    Xu, Kai; Lin, Jing; Zandi, Roza; Roth, Jack A; Ji, Lin

    2016-01-01

    MicroRNAs (miRNAs) play an important role in targeted gene silencing by facilitating posttranscriptional and translational repression. However, the precise mechanism of mammalian miRNA-mediated gene silencing remains to be elucidated. Here, we used a stem-loop array reverse-transcription polymerase chain reaction assay to analyse miRNA-induced mRNA recognition, cleavage, posttranscriptional modification, and degradation. We detected endogenous let-7 miRNA-induced and Argonaute-catalysed endonucleolytic cleavage on target mRNAs at various sites within partially paired miRNA:mRNA sequences. Most of the cleaved mRNA 5'-fragments were 3'-oligouridylated by activities of terminal uridylyl transferases (TUTases) in miRNA-induced silencing complexes and temporarily accumulated in the cytosol for 5'-3' degradation or other molecular fates. Some 3'-5' decayed mRNA fragments could also be captured by the miRNA-induced silencing complex stationed at the specific miRNA:mRNA target site and oligouridylated by other TUTases at its proximity without involving Argonaute-mediated RNA cleavage. Our findings provide new insights into the molecular mechanics of mammalian miRNA-mediated gene silencing by coordinated target mRNA recognition, cleavage, uridylation and degradation. PMID:27440378

  15. Control of Metastatic Progression by microRNA Regulatory Networks

    PubMed Central

    Pencheva, Nora; Tavazoie, Sohail F.

    2015-01-01

    Aberrant microRNA (miRNA) expression is a defining feature of human malignancy. Specific miRNAs have been identified as promoters or suppressors of metastatic progression. These miRNAs control metastasis through divergent or convergent regulation of metastatic gene pathways. Some miRNA regulatory networks govern cell-autonomous cancer phenotypes, while others modulate the cell-extrinsic composition of the metastatic microenvironment. The use of small RNAs as probes into the molecular and cellular underpinnings of metastasis holds promise for the identification of candidate genes for potential therapeutic intervention. PMID:23728460

  16. Control of metastatic progression by microRNA regulatory networks.

    PubMed

    Pencheva, Nora; Tavazoie, Sohail F

    2013-06-01

    Aberrant microRNA (miRNA) expression is a defining feature of human malignancy. Specific miRNAs have been identified as promoters or suppressors of metastatic progression. miRNAs control metastasis through divergent or convergent regulation of metastatic gene pathways. Some miRNA regulatory networks govern cell-autonomous cancer phenotypes, whereas others modulate the cell-extrinsic composition of the metastatic microenvironment. The use of small RNAs as probes into the molecular and cellular underpinnings of metastasis holds promise for the identification of candidate genes for potential therapeutic intervention. PMID:23728460

  17. The microRNA-argonaute complex: a platform for mRNA modulation.

    PubMed

    Hammell, Christopher M

    2008-01-01

    With the cloning the lin-4 gene in 1993, the possibility of an approximately 21-nucleotide RNA functioning as a regulatory molecule intrigued a relatively small number of scientists. This idea appeared to be a peculiarity of C. elegans as it was not until seven years later that the second, more conserved small RNA, let-7 was cloned. A spate of papers in 2000 and 2001 revealed that the underlying properties of the lin-4 and let-7 genes were a common facet of animal genomes and the absolute number and potential of this new class of gene products requires us to integrate them with other aspects of gene expression and evolution.(1-3) A wealth of information has accumulated in the intervening years that outline, in general, how these small RNAs are expressed and processed into a functional form. Contemporaneous to these studies, experiments also identified a cadre of evolutionarily conserved proteins, the Argonautes (Agos) that directly associate with and are required for microRNA function. Computational and experimental methods have led the identification of many functional mRNA targets. In the last few years, a significant body of work has focused on resolving two key issues: How do microRNAs function in particular genetic contexts (i.e., as "molecular switches" or "fine-tuners" of gene expression) and secondly, what facet/s of mRNA metabolism do microRNAs modulate in their role(s) as a regulatory molecule? The primary objective here is not to comprehensively compare the competing models of microRNA function (reviewed in refs. 4-6) but to frame a potential solution to these two fundamental questions by suggesting that the core microRNA-Ribonucleic-Protein Complex (microRNP), composed of the microRNA and an Ago protein, functions as a highly modifiable scaffold that associates with specific mRNAs via the bound microRNA and facilitates the localized activity of a variety of accessory proteins. The resulting composite mechanism could account for the apparent complexities

  18. MicroRNA in myogenesis and muscle atrophy

    PubMed Central

    Wang, Xiaonan H.

    2014-01-01

    Purpose of review To understand the impact of microRNA on myogenesis and muscle wasting in order to provide valuable information for clinical investigation. Recent findings Muscle wasting increases the risk of morbidity/mortality in primary muscle diseases, secondary muscle disorders and elderly population. Muscle mass is controlled by several different signalling pathways. Insulin-like growth factor/PI3K/Akt is a positive signalling pathway, as it increases muscle mass by increasing protein synthesis and decreasing protein degradation. This pathway is directly and/or indirectly downregulated by miR-1, miR-133, miR-206 or miR-125b, and upregulated by miR-23a or miR-486. Myostatin and the transforming growth factor-β signalling pathway are negative regulators that cause muscle wasting. An increase of miR-27 reduces myostatin and increases muscle cell proliferation. Muscle regeneration capacity also plays a significant role in the regulation of muscle mass. This review comprehensively describes the effect of microRNA on myoblasts proliferation and differentiation, and summarizes the varied influences of microRNA on different muscle atrophy. Summary Growing evidence indicates that microRNAs significantly impact muscle growth, regeneration and metabolism. MicroRNAs have a great potential to become diagnostic and/or prognostic markers, therapeutic agents and therapeutic targets. PMID:23449000

  19. Strategies to identify microRNA targets: New advances

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNAs (miRNAs) are small regulatory RNA molecules functioning to modulate gene expression at the post-transcriptional level, and playing an important role in many developmental and physiological processes. Ten thousand miRNAs have been discovered in various organisms. Although considerable progr...

  20. MicroRNA polymorphisms: a giant leap towards personalized medicine

    PubMed Central

    Mishra, Prasun J

    2010-01-01

    “An individual’s genetic inheritance of microRNA polymorphisms associated with disease progression, prognosis and treatment holds the key to create safer and more personalized drugs and can be a giant leap towards personalized medicine.” PMID:20428464

  1. MicroRNA in Human Glioma

    PubMed Central

    Li, Mengfeng; Li, Jun; Liu, Lei; Li, Wei; Yang, Yi; Yuan, Jie

    2013-01-01

    Glioma represents a serious health problem worldwide. Despite advances in surgery, radiotherapy, chemotherapy, and targeting therapy, the disease remains one of the most lethal malignancies in humans, and new approaches to improvement of the efficacy of anti-glioma treatments are urgently needed. Thus, new therapeutic targets and tools should be developed based on a better understanding of the molecular pathogenesis of glioma. In this context, microRNAs (miRNAs), a class of small, non-coding RNAs, play a pivotal role in the development of the malignant phenotype of glioma cells, including cell survival, proliferation, differentiation, tumor angiogenesis, and stem cell generation. This review will discuss the biological functions of miRNAs in human glioma and their implications in improving clinical diagnosis, prediction of prognosis, and anti-glioma therapy. PMID:24202447

  2. MicroRNA regulation of lymphocyte tolerance and autoimmunity

    PubMed Central

    Simpson, Laura J.; Ansel, K. Mark

    2015-01-01

    Understanding the cell-intrinsic cues that permit self-reactivity in lymphocytes, and therefore autoimmunity, requires an understanding of the transcriptional and posttranscriptional regulation of gene expression in these cells. In this Review, we address seminal and recent research on microRNA (miRNA) regulation of central and peripheral tolerance. Human and mouse studies demonstrate that the PI3K pathway is a critical point of miRNA regulation of immune cell development and function that affects the development of autoimmunity. We also discuss how miRNA expression profiling in human autoimmune diseases has inspired mechanistic studies of miRNA function in the pathogenesis of multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, type 1 diabetes, and asthma. PMID:26030228

  3. MetaMirClust: Discovery and Exploration of Evolutionarily Conserved miRNA Clusters.

    PubMed

    Chan, Wen-Ching; Lin, Wen-Chang

    2016-01-01

    Recent emerging studies suggest that a substantial fraction of microRNA (miRNA) genes is likely to form clusters in terms of evolutionary conservation and biological implications, posing a significant challenge for the research community and shifting the bottleneck of scientific discovery from miRNA singletons to miRNA clusters. In addition, the advance in molecular sequencing technique such as next-generation sequencing (NGS) has facilitated researchers to comprehensively characterize miRNAs with low abundance on genome-wide scale in multiple species. Taken together, a large scale, cross-species survey of grouped miRNAs based on genomic location would be valuable for investigating their biological functions and regulations in an evolutionary perspective. In the present chapter, we describe the application of effective and efficient bioinformatics tools on the identification of clustered miRNAs and illustrate how to use the recently developed Web-based database, MetaMirClust ( http://fgfr.ibms.sinic.aedu.tw/MetaMirClust ) to discover evolutionarily conserved pattern of miRNA clusters across metazoans. PMID:25861770

  4. microRNAs and RNA-binding proteins

    PubMed Central

    Ciafrè, Silvia Anna; Galardi, Silvia

    2013-01-01

    In the last decade, an ever-growing number of connections between microRNAs (miRNAs) and RNA-binding proteins (RBPs) have uncovered a new level of complexity of gene expression regulation in cancer. In this review, we examine several aspects of the functional interactions between miRNAs and RBPs in cancer models. We will provide examples of reciprocal regulation: miRNAs regulating the expression of RBPs, or the converse, where an RNA-binding protein specifically regulates the expression of a specific miRNA, or when an RBP can exert a widespread effect on miRNAs via the modulation of a key protein for miRNA production or function. Moreover, we will focus on the ever-growing number of functional interactions that have been discovered in the last few years: RBPs that were shown to cooperate with microRNAs in the downregulation of shared target mRNAs or, on the contrary, that inhibit microRNA action, thus resulting in a protection of the specific target mRNAs. We surely need to obtain a deeper comprehension of such intricate networks to have a chance of understanding and, thus, fighting cancer. PMID:23696003

  5. Plant-based microRNA presences in mice and human sera to breast milk

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plant foods contain hundreds of thousands of different small RNAs, including microRNAs (miRNAs). A microRNA (miRNA) is a tiny (19-24 nucleotide) piece of RNA that attaches to a specific protein-making mRNA, thus inhibiting protein production. A recent finding shows that a miRNA in rice survives dige...

  6. MicroRNA variants as genetic determinants of bone mass.

    PubMed

    Dole, Neha S; Delany, Anne M

    2016-03-01

    Single nucleotide polymorphisms (SNPs) are the most abundant genetic variants that contribute to the heritability of bone mass. MicroRNAs (miRNAs, miRs) are key post-transcriptional regulators that modulate the differentiation and function of skeletal cells by targeting multiple genes in the same or distinct signaling pathways. SNPs in miRNA genes and miRNA binding sites can alter miRNA abundance and mRNA targeting. This review describes the potential impact of miRNA-related SNPs on skeletal phenotype. Although many associations between SNPs and bone mass have been described, this review is limited to gene variants for which a function has been experimentally validated. SNPs in miRNA genes (miR-SNPs) that impair miRNA processing and alter the abundance of mature miRNA are discussed for miR-146a, miR-125a, miR-196a, miR-149 and miR-27a. SNPs in miRNA targeting sites (miR-TS-SNPs) that alter miRNA binding are described for the bone remodeling genes bone morphogenetic protein receptor 1 (Bmpr1), fibroblast growth factor 2 (Fgf2), osteonectin (Sparc) and histone deacetylase 5 (Hdac5). The review highlights two aspects of miRNA-associated SNPs: the mechanism for altering miRNA mediated gene regulation and the potential of miR-associated SNPs to alter osteoblast, osteoclast or chondrocyte differentiation and function. Given the polygenic nature of skeletal diseases like osteoporosis and osteoarthritis, validating the function of additional miRNA-associated SNPs has the potential to enhance our understanding of the genetic determinants of bone mass and predisposition to selected skeletal diseases. PMID:26723575

  7. A structural view of microRNA-target recognition.

    PubMed

    Leoni, Guido; Tramontano, Anna

    2016-05-19

    It is well established that the correct identification of the messenger RNA targeted by a given microRNA (miRNA) is a difficult problem, and that available methods all suffer from low specificity. We hypothesize that the correct identification of the pairing should take into account the effect of the Argonaute protein (AGO), an essential catalyst of the recognition process. Therefore, we developed a strategy named MiREN for building and scoring three-dimensional models of the ternary complex formed by AGO, a miRNA and 22 nt of a target mRNA that putatively interacts with it. We show here that MiREN can be used to assess the likelihood that an RNA molecule is the target of a given miRNA and that this approach is more accurate than other existing methods, usually based on sequence or sequence-related features. Our results also suggest that AGO plays a relevant role in the selection of the miRNA targets. Our method can represent an additional step for refining predictions made by faster but less accurate classical methods for the identification of miRNA targets. PMID:26825463

  8. Intratumoral Heterogeneity of MicroRNA Expression in Rectal Cancer

    PubMed Central

    Andersen, Rikke Fredslund; Nielsen, Boye Schnack; Sørensen, Flemming Brandt; Appelt, Ane Lindegaard; Jakobsen, Anders; Hansen, Torben Frøstrup

    2016-01-01

    Introduction An increasing number of studies have investigated microRNAs (miRNAs) as potential markers of diagnosis, treatment and prognosis. So far, agreement between studies has been minimal, which may in part be explained by intratumoral heterogeneity of miRNA expression. The aim of the present study was to assess the heterogeneity of a panel of selected miRNAs in rectal cancer, using two different technical approaches. Materials and Methods The expression of the investigated miRNAs was analysed by real-time quantitative polymerase chain reaction (RT-qPCR) and in situ hybridization (ISH) in tumour specimens from 27 patients with T3-4 rectal cancer. From each tumour, tissue from three different luminal localisations was examined. Inter- and intra-patient variability was assessed by calculating intraclass correlation coefficients (ICCs). Correlations between RT-qPCR and ISH were evaluated using Spearman’s correlation. Results ICCsingle (one sample from each patient) was higher than 50% for miRNA-21 and miRNA-31. For miRNA-125b, miRNA-145, and miRNA-630, ICCsingle was lower than 50%. The ICCmean (mean of three samples from each patient) was higher than 50% for miRNA-21(RT-qPCR and ISH), miRNA-125b (RT-qPCR and ISH), miRNA-145 (ISH), miRNA-630 (RT-qPCR), and miRNA-31 (RT-qPCR). For miRNA-145 (RT-qPCR) and miRNA-630 (ISH), ICCmean was lower than 50%. Spearman correlation coefficients, comparing results obtained by RT-qPCR and ISH, respectively, ranged from 0.084 to 0.325 for the mean value from each patient, and from -0.085 to 0.515 in the section including the deepest part of the tumour. Conclusion Intratumoral heterogeneity may influence the measurement of miRNA expression and consequently the number of samples needed for representative estimates. Our findings with two different methods suggest that one sample is sufficient for adequate assessment of miRNA-21 and miRNA-31, whereas more samples would improve the assessment of miRNA-125b, miRNA-145, and miRNA-630

  9. The microRNA156 and microRNA172 gene regulation cascades at post-germinative stages in Arabidopsis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNAs (miRNAs) are involved in developmental programs of plants including seed germination and post-germination. Here, we provide evidence that two different miRNA pathways, miR156 and miR172, interact during the post-germination stages in Arabidopsis. Mutant seedlings expressing miR156resistant...

  10. piRNA clusters and open chromatin structure

    PubMed Central

    2014-01-01

    Transposable elements (TEs) are major structural components of eukaryotic genomes; however, mobilization of TEs generally has negative effects on the host genome. To counteract this threat, host cells have evolved genetic and epigenetic mechanisms that keep TEs silenced. One such mechanism involves the Piwi-piRNA complex, which represses TEs in animal gonads either by cleaving TE transcripts in the cytoplasm or by directing specific chromatin modifications at TE loci in the nucleus. Most Piwi-interacting RNAs (piRNAs) are derived from genomic piRNA clusters. There has been remarkable progress in our understanding of the mechanisms underlying piRNA biogenesis. However, little is known about how a specific locus in the genome is converted into a piRNA-producing site. In this review, we will discuss a possible link between chromatin boundaries and piRNA cluster formation. PMID:25126116

  11. MicroRNA 33 Regulates Glucose Metabolism

    PubMed Central

    Ramírez, Cristina M.; Goedeke, Leigh; Rotllan, Noemi; Yoon, Je-Hyun; Cirera-Salinas, Daniel; Mattison, Julie A.; Suárez, Yajaira; de Cabo, Rafael; Gorospe, Myriam

    2013-01-01

    Metabolic diseases are characterized by the failure of regulatory genes or proteins to effectively orchestrate specific pathways involved in the control of many biological processes. In addition to the classical regulators, recent discoveries have shown the remarkable role of small noncoding RNAs (microRNAs [miRNAs]) in the posttranscriptional regulation of gene expression. In this regard, we have recently demonstrated that miR-33a and miR33b, intronic miRNAs located within the sterol regulatory element-binding protein (SREBP) genes, regulate lipid metabolism in concert with their host genes. Here, we show that miR-33b also cooperates with SREBP1 in regulating glucose metabolism by targeting phosphoenolpyruvate carboxykinase (PCK1) and glucose-6-phosphatase (G6PC), key regulatory enzymes of hepatic gluconeogenesis. Overexpression of miR-33b in human hepatic cells inhibits PCK1 and G6PC expression, leading to a significant reduction of glucose production. Importantly, hepatic SREBP1c/miR-33b levels correlate inversely with the expression of PCK1 and G6PC upon glucose infusion in rhesus monkeys. Taken together, these results suggest that miR-33b works in concert with its host gene to ensure a fine-tuned regulation of lipid and glucose homeostasis, highlighting the clinical potential of miR-33a/b as novel therapeutic targets for a range of metabolic diseases. PMID:23716591

  12. Discovery of MicroRNA169 Gene Copies in Genomes of Flowering Plants through Positional Information

    PubMed Central

    Calviño, Martín; Messing, Joachim

    2013-01-01

    Expansion and contraction of microRNA (miRNA) families can be studied in sequenced plant genomes through sequence alignments. Here, we focused on miR169 in sorghum because of its implications in drought tolerance and stem-sugar content. We were able to discover many miR169 copies that have escaped standard genome annotation methods. A new miR169 cluster was found on sorghum chromosome 1. This cluster is composed of the previously annotated sbi-MIR169o together with two newly found MIR169 copies, named sbi-MIR169t and sbi-MIR169u. We also found that a miR169 cluster on sorghum chr7 consisting of sbi-MIR169l, sbi-MIR169m, and sbi-MIR169n is contained within a chromosomal inversion of at least 500 kb that occurred in sorghum relative to Brachypodium, rice, foxtail millet, and maize. Surprisingly, synteny of chromosomal segments containing MIR169 copies with linked bHLH and CONSTANS-LIKE genes extended from Brachypodium to dictotyledonous species such as grapevine, soybean, and cassava, indicating a strong conservation of linkages of certain flowering and/or plant height genes and microRNAs, which may explain linkage drag of drought and flowering traits and would have consequences for breeding new varieties. Furthermore, alignment of rice and sorghum orthologous regions revealed the presence of two additional miR169 gene copies (miR169r and miR169s) on sorghum chr7 that formed an antisense miRNA gene pair. Both copies are expressed and target different set of genes. Synteny-based analysis of microRNAs among different plant species should lead to the discovery of new microRNAs in general and contribute to our understanding of their evolution. PMID:23348041

  13. Discovery of MicroRNA169 gene copies in genomes of flowering plants through positional information.

    PubMed

    Calviño, Martín; Messing, Joachim

    2013-01-01

    Expansion and contraction of microRNA (miRNA) families can be studied in sequenced plant genomes through sequence alignments. Here, we focused on miR169 in sorghum because of its implications in drought tolerance and stem-sugar content. We were able to discover many miR169 copies that have escaped standard genome annotation methods. A new miR169 cluster was found on sorghum chromosome 1. This cluster is composed of the previously annotated sbi-MIR169o together with two newly found MIR169 copies, named sbi-MIR169t and sbi-MIR169u. We also found that a miR169 cluster on sorghum chr7 consisting of sbi-MIR169l, sbi-MIR169m, and sbi-MIR169n is contained within a chromosomal inversion of at least 500 kb that occurred in sorghum relative to Brachypodium, rice, foxtail millet, and maize. Surprisingly, synteny of chromosomal segments containing MIR169 copies with linked bHLH and CONSTANS-LIKE genes extended from Brachypodium to dictotyledonous species such as grapevine, soybean, and cassava, indicating a strong conservation of linkages of certain flowering and/or plant height genes and microRNAs, which may explain linkage drag of drought and flowering traits and would have consequences for breeding new varieties. Furthermore, alignment of rice and sorghum orthologous regions revealed the presence of two additional miR169 gene copies (miR169r and miR169s) on sorghum chr7 that formed an antisense miRNA gene pair. Both copies are expressed and target different set of genes. Synteny-based analysis of microRNAs among different plant species should lead to the discovery of new microRNAs in general and contribute to our understanding of their evolution. PMID:23348041

  14. MicroRNA-338 and microRNA-21 co-transfection for the treatment of rat sciatic nerve injury.

    PubMed

    Wang, Jianyong; Muheremu, Aikeremujiang; Zhang, Ming; Gong, Kai; Huang, Chuyi; Ji, Yuchen; Wei, Yujun; Ao, Qiang

    2016-06-01

    The objective of this study is to find if co-transfecting microRNA-338 and microRNA-21 into the neurons in the spinal cord can promote functional recovery after peripheral nerve injury in rats. Animals were divided into three groups: 20 animals in the GFP control vector group (group A), 20 animals in the GFP experimental vector group (group B) and ten animals in the normal control group. Right sciatic nerves of animals in groups A and B were transected and were bridged with collagen nerve conduits with 10 mm distance between the stumps. 3 µl GFP control vector or 3 µl lentiviral vectors encoding the sequence of microRNA-338 and microRNA-21 were injected in the conduit. 8 weeks after the surgery, the treatment effect was evaluated by functional analysis, electrophysiological analysis, immunohistochemical analysis as well as transmitting electronic microscope observations in all the rats. Animals treated with microRNA-338 and microRNA-21 showed significantly better recovery than GFP control group animals by means of functional analysis (Sciatic nerve index -47.7 ± 2.5 vs -59.4 ± 3.7), electrophysiological analysis (Conduction velocity 20.5 ± 2.8 vs 10.5 ± 1.4 m/s), ratio of wet weight of the gastrocnemius muscles (0.83 ± 0.03 vs 0.55 ± 0.06), axon diameter (5.0 ± 1.8 µm vs 4.0 ± 2.2), myelin sheath thickness (1.4 ± 0.43 vs 0.80 ± 0.31 µm) and G-ratio (0.80 ± 0.06 vs 0.75 ± 0.04). Lentiviral vectors encoding microRNA 338 and 21 might be explored in the future as potential therapeutic intervention to promote nerve regeneration. PMID:26909749

  15. Accurate microRNA target prediction correlates with protein repression levels

    PubMed Central

    Maragkakis, Manolis; Alexiou, Panagiotis; Papadopoulos, Giorgio L; Reczko, Martin; Dalamagas, Theodore; Giannopoulos, George; Goumas, George; Koukis, Evangelos; Kourtis, Kornilios; Simossis, Victor A; Sethupathy, Praveen; Vergoulis, Thanasis; Koziris, Nectarios; Sellis, Timos; Tsanakas, Panagiotis; Hatzigeorgiou, Artemis G

    2009-01-01

    Background MicroRNAs are small endogenously expressed non-coding RNA molecules that regulate target gene expression through translation repression or messenger RNA degradation. MicroRNA regulation is performed through pairing of the microRNA to sites in the messenger RNA of protein coding genes. Since experimental identification of miRNA target genes poses difficulties, computational microRNA target prediction is one of the key means in deciphering the role of microRNAs in development and disease. Results DIANA-microT 3.0 is an algorithm for microRNA target prediction which is based on several parameters calculated individually for each microRNA and combines conserved and non-conserved microRNA recognition elements into a final prediction score, which correlates with protein production fold change. Specifically, for each predicted interaction the program reports a signal to noise ratio and a precision score which can be used as an indication of the false positive rate of the prediction. Conclusion Recently, several computational target prediction programs were benchmarked based on a set of microRNA target genes identified by the pSILAC method. In this assessment DIANA-microT 3.0 was found to achieve the highest precision among the most widely used microRNA target prediction programs reaching approximately 66%. The DIANA-microT 3.0 prediction results are available online in a user friendly web server at PMID:19765283

  16. Clinical Potential of microRNA-7 in Cancer

    PubMed Central

    Horsham, Jessica L.; Kalinowski, Felicity C.; Epis, Michael R.; Ganda, Clarissa; Brown, Rikki A. M.; Leedman, Peter J.

    2015-01-01

    microRNAs (miRNAs) are a family of short, non-coding RNA molecules that drive a complex network of post-transcriptional gene regulation by enhancing target mRNA decay and/or inhibiting protein synthesis from mRNA transcripts. They regulate genes involved in key aspects of normal cell growth, development and the maintenance of body homeostasis and have been closely linked to the development and progression of human disease, in particular cancer. Over recent years there has been much interest regarding their potential as biomarkers and as therapeutic agents or targets. microRNA-7 (miR-7) is a 23 nucleotide (nt) miRNA known primarily to act as a tumour suppressor. miR-7 directly inhibits a number of oncogenic targets and impedes various aspects of cancer progression in vitro and in vivo, however, some studies have also implicated miR-7 in oncogenic roles. This review summarises the role of miR-7 in cancer, its potential in miRNA-based replacement therapy and its capacity as both a diagnostic and prognostic biomarker. PMID:26308064

  17. New wheat microRNA using whole-genome sequence.

    PubMed

    Kurtoglu, Kuaybe Yucebilgili; Kantar, Melda; Budak, Hikmet

    2014-06-01

    MicroRNAs are post-transcriptional regulators of gene expression, taking roles in a variety of fundamental biological processes. Hence, their identification, annotation and characterization are of great significance, especially in bread wheat, one of the main food sources for humans. The recent availability of 5× coverage Triticum aestivum L. whole-genome sequence provided us with the opportunity to perform a systematic prediction of a complete catalogue of wheat microRNAs. Using an in silico homology-based approach, stem-loop coding regions were derived from two assemblies, constructed from wheat 454 reads. To avoid the presence of pseudo-microRNAs in the final data set, transposable element related stem-loops were eliminated by repeat analysis. Overall, 52 putative wheat microRNAs were predicted, including seven, which have not been previously published. Moreover, with distinct analysis of the two different assemblies, both variety and representation of putative microRNA-coding stem-loops were found to be predominant in the intergenic regions. By searching available expressed sequences and small RNA library databases, expression evidence for 39 (out of 52) putative wheat microRNAs was provided. Expression of three of the predicted microRNAs (miR166, miR396 and miR528) was also comparatively quantified with real-time quantitative reverse transcription PCR. This is the first report on in silico prediction of a whole repertoire of bread wheat microRNAs, supported by the wet-lab validation. PMID:24395439

  18. Nonconventional chemical inhibitors of microRNA: therapeutic scope.

    PubMed

    Jayaraj, Gopal Gunanathan; Nahar, Smita; Maiti, Souvik

    2015-01-18

    MicroRNAs (miRNAs) are a class of genomically encoded small RNA molecules (∼22nts in length), which regulate gene expression post transcriptionally. The term microRNA or miRNA was coined in 2001, and research in the past decade has shed light on their widespread occurrence, evolutionary conservation and tissue specific functions. It is estimated that they modulate the gene expression of approximately 60% of the mammalian genes by regulating the levels of target mRNAs to which they can bind on the basis of sequence complementarities. miRNAs are produced in a well coordinated series of steps from being transcribed in the nucleus to exerting their function in the cytoplasm. miRNAs are now implicated in diverse biological phenomena ranging from development to stress response which makes miRNAs one of the central regulatory molecules which modulate information flow along the central dogma of gene expression. More importantly, like any regulatory molecule, deregulation of miRNAs is causally associated with several diseases (mainly cancer) and is directly involved in a variety of pathophysiologies owing to their aberrant expression. Thus, modulation of miRNA levels is of prime therapeutic importance. Conventional methods of miRNA knockdown using chemically modified antisense-oligonucleotides have been explored extensively but face the challenges of modes of delivery, biostability and biodistribution. This calls for the development of more alternative and non-conventional methods to target miRNA. Small molecules targeting RNA chemical and structural space provide one such timely opportunity. In this article we first provide a brief overview of miRNA biogenesis and its disease associations. We then summarize the major developments in conventional oligonucleotide based approaches to miRNA knockdown and its status. We then focus on the more non-conventional methods like oligonucleotide enzymes and small molecules and provide an outlook on the future of such methods. PMID

  19. Molecular Beacon-Based MicroRNA Imaging During Neurogenesis.

    PubMed

    Lee, Jonghwan; Kim, Soonhag

    2016-01-01

    The fluorescence monitoring system for examining endogenous microRNA (miRNA) activity in cellular level provides crucial information on not only understanding a critical role of miRNA involving a variety of biological processes, but also evaluating miRNA expression patterns in a noninvasive manner. In this protocol, we report the details of a new procedure for a molecular beacon-based miRNA monitoring system, which includes the illustration scheme for miRNA detection strategy, exogenous miRNA detection, and measurement of endogenous miRNA expression level during neurogenesis. The fluorescence signal of miR-124a beacon quenched by BHQ2 was gradually recovered as increasing concentration of the miR-124a in tube. The functional work of miR-124a beacon was examined in intracellular environment, allowing for the internalization of the miR-124a beacon by lipofectamine, which resulted in activated fluorescent signals of the miR-124a beacon in the HeLa cells after the addition of synthetic miR-124a. The endogenous miR-124a expression level was detected by miR-124a beacon system during neurogenesis, showing brighter fluorescence intensity in cytoplasmic area of P19 cells after induction of neuronal differentiation by retinoic acid. The molecular beacon based-miRNA detection technique could be applicable to the simultaneous visualization of a variety of miRNA expression patterns using different fluorescence dyes. For the study of examining endogenous miRNA expression level using miRNA-beacon system, if cellular differentiation step is already prepared, transfection step of miR-124a beacon into P19 cells, and acquisition of activated fluorescence signal measured by confocal microscope can be conducted approximately within 6 h. PMID:26530921

  20. The Expansion of Animal MicroRNA Families Revisited

    PubMed Central

    Hertel, Jana; Stadler, Peter F.

    2015-01-01

    MicroRNAs are important regulatory small RNAs in many eukaryotes. Due to their small size and simple structure, they are readily innovated de novo. Throughout the evolution of animals, the emergence of novel microRNA families traces key morphological innovations. Here, we use a computational approach based on homology search and parsimony-based presence/absence analysis to draw a comprehensive picture of microRNA evolution in 159 animal species. We confirm previous observations regarding bursts of innovations accompanying the three rounds of genome duplications in vertebrate evolution and in the early evolution of placental mammals. With a much better resolution for the invertebrate lineage compared to large-scale studies, we observe additional bursts of innovation, e.g., in Rhabditoidea. More importantly, we see clear evidence that loss of microRNA families is not an uncommon phenomenon. The Enoplea may serve as a second dramatic example beyond the tunicates. The large-scale analysis presented here also highlights several generic technical issues in the analysis of very large gene families that will require further research. PMID:25780960

  1. miRNA-dis: microRNA precursor identification based on distance structure status pairs.

    PubMed

    Liu, Bin; Fang, Longyun; Chen, Junjie; Liu, Fule; Wang, Xiaolong

    2015-04-01

    MicroRNA precursor identification is an important task in bioinformatics. Support Vector Machine (SVM) is one of the most effective machine learning methods used in this field. The performance of SVM-based methods depends on the vector representations of RNAs. However, the discriminative power of the existing feature vectors is limited, and many methods lack an interpretable model for analysis of characteristic sequence features. Prior studies have demonstrated that sequence or structure order effects were relevant for discrimination, but little work has explored how to use this kind of information for human pre-microRNA identification. In this study, in order to incorporate the structure-order information into the prediction, a method called "miRNA-dis" was proposed, in which the feature vector was constructed by the occurrence frequency of the "distance structure status pair" or just the "distance-pair". Rigorous cross-validations on a much larger and more stringent newly constructed benchmark dataset showed that the miRNA-dis outperformed some state-of-the-art predictors in this area. Remarkably, miRNA-dis trained with human data can correctly predict 87.02% of the 4022 pre-miRNAs from 11 different species ranging from animals, plants and viruses. miRNA-dis would be a useful high throughput tool for large-scale analysis of microRNA precursors. In addition, the learnt model can be easily analyzed in terms of discriminative features, and some interesting patterns were discovered, which could reflect the characteristics of microRNAs. A user-friendly web-server of miRNA-dis was constructed, which is freely accessible to the public at the web-site on http://bioinformatics.hitsz.edu.cn/miRNA-dis/. PMID:25715848

  2. MicroRNA-23a regulates 3T3-L1 adipocyte differentiation.

    PubMed

    Shen, Linyuan; Zhang, Yi; Du, Jingjing; Chen, Li; Luo, Jia; Li, Xuewei; Li, Mingzhou; Tang, Guoqing; Zhang, Shunhua; Zhu, Li

    2016-01-10

    MicroRNAs (miRNAs) are small, non-coding RNAs, which are involved in regulation of a variety of biological processes. Since previous studies regarding the role of miRNAs in the regulation of adipogenic differentiation have shown that miRNA-27a, one member of miRNA-23a∼27a∼24 cluster, could suppress adipogenesis. We now investigated whether miRNA-23a regulates adipogenic differentiation. In the present study, we showed that the expression of miRNA-23a is decreased during the process of adipogenic differentiation. Over-expression of miRNA-23a decreased lipid accumulation and triglyceride content in 3T3-L1 adipocytes. Our results also demonstrated that miRNA-23a decreases mRNA levels of adipocyte-specific genes involved in lipogenic transcription, fatty acid synthesis and fatty acid transport. These findings suggested miRNA-23a to be a new type of adipogenic depressor and to play an important role in regulating adipocyte differentiation. PMID:26415879

  3. Epstein-Barr Virus MicroRNA Expression Increases Aggressiveness of Solid Malignancies

    PubMed Central

    Pandya, Deep; Mariani, Marisa; He, Shiquan; Andreoli, Mirko; Spennato, Manuela; Dowell-Martino, Candice; Fiedler, Paul; Ferlini, Cristiano

    2015-01-01

    The Cancer Genome Atlas (TCGA) microRNA (miRNA) initiative has revealed a pivotal role for miRNAs in cancer. Utilizing the TCGA raw data, we performed the first mapping of viral miRNA sequences within cancer and adjacent normal tissues. Results were integrated with TCGA RNA-seq to link the expression of viral miRNAs to the phenotype. Using clinical data and viral miRNA mapping results we also performed outcome analysis. Three lines of evidence lend credence to an active role of viral miRNAs in solid malignancies. First, expression of viral miRNA is consistently higher in cancerous compared to adjacent noncancerous tissues. Second, viral miRNA expression is associated with significantly worse clinical outcome among patients with early stage malignancy. These patients are also featured by increased expression of PD1/PD-L1, a pathway implicated in tumors escaping immune destruction. Finally, a particular cluster of EBV-miRNA (miR-BART2, miR-BART4, miR-BART5, miR-BART18, and miR-BART22) is associated with expression of cytokines known to inhibit host response to cancer. Quantification of specific viral miRNAs may help identify patients who are at risk of poor outcome. These patients may be candidates for novel therapeutic strategies incorporating antiviral agents and/or inhibitors of the PD-1/PD-L1 pathway. PMID:26375401

  4. MicroRNA processing pathway regulates olfactory neuron morphogenesis.

    PubMed

    Berdnik, Daniela; Fan, Audrey P; Potter, Christopher J; Luo, Liqun

    2008-11-25

    The microRNA (miRNA) processing pathway produces miRNAs as posttranscriptional regulators of gene expression. The nuclear RNase III Drosha catalyzes the first processing step together with the dsRNA binding protein DGCR8/Pasha generating pre-miRNAs [1, 2]. The next cleavage employs the cytoplasmic RNase III Dicer producing miRNA duplexes [3, 4]. Finally, Argonautes are recruited with miRNAs into an RNA-induced silencing complex for mRNA recognition (Figure 1A). Here, we identify two members of the miRNA pathway, Pasha and Dicer-1, in a forward genetic screen for mutations that disrupt wiring specificity of Drosophila olfactory projection neurons (PNs). The olfactory system is built as discrete map of highly stereotyped neuronal connections [5, 6]. Each PN targets dendrites to a specific glomerulus in the antennal lobe and projects axons stereotypically into higher brain centers [7-9]. In selected PN classes, pasha and Dicer-1 mutants cause specific PN dendrite mistargeting in the antennal lobe and altered axonal terminations in higher brain centers. Furthermore, Pasha and Dicer-1 act cell autonomously in postmitotic neurons to regulate dendrite and axon targeting during development. However, Argonaute-1 and Argonaute-2 are dispensable for PN morphogenesis. Our findings suggest a role for the miRNA processing pathway in establishing wiring specificity in the nervous system. PMID:19013069

  5. Birth and expression evolution of mammalian microRNA genes.

    PubMed

    Meunier, Julien; Lemoine, Frédéric; Soumillon, Magali; Liechti, Angélica; Weier, Manuela; Guschanski, Katerina; Hu, Haiyang; Khaitovich, Philipp; Kaessmann, Henrik

    2013-01-01

    MicroRNAs (miRNAs) are major post-transcriptional regulators of gene expression, yet their origins and functional evolution in mammals remain little understood due to the lack of appropriate comparative data. Using RNA sequencing, we have generated extensive and comparable miRNA data for five organs in six species that represent all main mammalian lineages and birds (the evolutionary outgroup) with the aim to unravel the evolution of mammalian miRNAs. Our analyses reveal an overall expansion of miRNA repertoires in mammals, with threefold accelerated birth rates of miRNA families in placentals and marsupials, facilitated by the de novo emergence of miRNAs in host gene introns. Generally, our analyses suggest a high rate of miRNA family turnover in mammals with many newly emerged miRNA families being lost soon after their formation. Selectively preserved mammalian miRNA families gradually evolved higher expression levels, as well as altered mature sequences and target gene repertoires, and were apparently mainly recruited to exert regulatory functions in nervous tissues. However, miRNAs that originated on the X chromosome evolved high expression levels and potentially diverse functions during spermatogenesis, including meiosis, through selectively driven duplication-divergence processes. Overall, our study thus provides detailed insights into the birth and evolution of mammalian miRNA genes and the associated selective forces. PMID:23034410

  6. MicroRNA-27b Regulates Mitochondria Biogenesis in Myocytes

    PubMed Central

    Zhang, Shunhua; Du, Jingjing; Bai, Lin; Zhang, Yi; Jiang, Yanzhi; Li, Xuewei; Wang, Jinyong; Zhu, Li

    2016-01-01

    MicroRNAs (miRNAs) are small, non-coding RNAs that affect the post-transcriptional regulation of various biological pathways. To date, it is not fully understood how miRNAs regulate mitochondrial biogenesis. This study aimed at the identification of the role of miRNA-27b in mitochondria biogenesis. The mitochondria content in C2C12 cells was significantly increased during myogenic differentiation and accompanied by a marked decrease of miRNA-27b expression. Furthermore, the expression of the predicted target gene of miRNA-27b, forkhead box j3 (Foxj3), was also increased during myogenic differentiation. Luciferase activity assays confirmed that miRNA-27b directly targets the 3’-untranslated region (3’-UTR) of Foxj3. Overexpression of miRNA-27b provoked a decrease of mitochondria content and diminished expression of related mitochondrial genes and Foxj3 both at mRNA and protein levels. The expression levels of downstream genes of Foxj3, such as Mef2c, PGC1α, NRF1 and mtTFA, were also decreased in C2C12 cells upon overexpression of miRNA-27b. These results suggested that miRNA-27b may affect mitochondria biogenesis by down-regulation of Foxj3 during myocyte differentiation. PMID:26849429

  7. Profiling of microRNA expression by mRAP.

    PubMed

    Takada, Shuji; Mano, Hiroyuki

    2007-01-01

    MicroRNA (miRNA) amplification profiling (mRAP) is a sensitive method for the determination of miRNA expression profiles. The method relies on a long, optimized 5' adaptor and the SMART (switching mechanism at the 5' end of RNA templates of reverse transcriptase) reaction to yield miRNA-derived cDNAs flanked by synthesized oligomers at each end. The cDNAs are PCR-amplified with primers corresponding to the oligomers, and the products are concatamerized for nucleotide sequencing. The expression level of each miRNA can be estimated from the frequency of the occurrence of its sequence in the data set, provided that sufficient clones of the cDNAs are sequenced. This method potentially yields millions of miRNA-derived clones from as few as 1 x 10(4) cells, thus allowing the characterization of miRNA expression profiles with small quantities of starting material such as those available for fresh clinical specimens or organs of developing embryos. This protocol can be completed in 10 d. PMID:18079713

  8. MicroRNA control of lymphocyte differentiation and function

    PubMed Central

    Belver, Laura; Papavasiliou, Nina F; Ramiro, Almudena R

    2011-01-01

    Summary MicroRNAs (miRNAs) are a class of endogenous, non-coding regulatory RNAs that control gene regulation by guiding silencing protein complexes to mRNA in a sequence-dependent manner. In this way miRNAs are able to repress gene expression post-transcriptionally by affecting mRNA stability or translation. These ubiquitous molecules play central roles in a wide range of biological processes, including cell proliferation, differentiation and apoptosis. Within the context of the immune system, genetic studies have identified distinct roles for specific miRNAs in gene regulation during development, activation and maturation. Conversely, dysregulation of miRNA expression has been specifically correlated with cancer. This review outlines our current understanding of miRNA function in lymphocytes as it impacts expression of protein-coding genes in the context of proper development, as well as oncogenesis. PMID:21353514

  9. Evolutionary Transitions of MicroRNA-Target Pairs

    PubMed Central

    Nozawa, Masafumi; Fujimi, Mai; Iwamoto, Chie; Onizuka, Kanako; Fukuda, Nana; Ikeo, Kazuho; Gojobori, Takashi

    2016-01-01

    How newly generated microRNA (miRNA) genes are integrated into gene regulatory networks during evolution is fundamental in understanding the molecular and evolutionary bases of robustness and plasticity in gene regulation. A recent model proposed that after the birth of a miRNA, the miRNA is generally integrated into the network by decreasing the number of target genes during evolution. However, this decreasing model remains to be carefully examined by considering in vivo conditions. In this study, we therefore compared the number of target genes among miRNAs with different ages, combining experiments with bioinformatics predictions. First, we focused on three Drosophila miRNAs with different ages. As a result, we found that an older miRNA has a greater number of target genes than a younger miRNA, suggesting the increasing number of targets for each miRNA during evolution (increasing model). To further confirm our results, we also predicted all target genes for all miRNAs in D. melanogaster, considering co-expression of miRNAs and mRNAs in vivo. The results obtained also do not support the decreasing model but are reasonably consistent with the increasing model of miRNA-target pairs. Furthermore, our large-scale analyses of currently available experimental data of miRNA-target pairs also showed a weak but the same trend in humans. These results indicate that the current decreasing model of miRNA-target pairs should be reconsidered and the increasing model may be more appropriate to explain the evolutionary transitions of miRNA-target pairs in many organisms. PMID:27189995

  10. Evolutionary Transitions of MicroRNA-Target Pairs.

    PubMed

    Nozawa, Masafumi; Fujimi, Mai; Iwamoto, Chie; Onizuka, Kanako; Fukuda, Nana; Ikeo, Kazuho; Gojobori, Takashi

    2016-01-01

    How newly generated microRNA (miRNA) genes are integrated into gene regulatory networks during evolution is fundamental in understanding the molecular and evolutionary bases of robustness and plasticity in gene regulation. A recent model proposed that after the birth of a miRNA, the miRNA is generally integrated into the network by decreasing the number of target genes during evolution. However, this decreasing model remains to be carefully examined by considering in vivo conditions. In this study, we therefore compared the number of target genes among miRNAs with different ages, combining experiments with bioinformatics predictions. First, we focused on three Drosophila miRNAs with different ages. As a result, we found that an older miRNA has a greater number of target genes than a younger miRNA, suggesting the increasing number of targets for each miRNA during evolution (increasing model). To further confirm our results, we also predicted all target genes for all miRNAs in D. melanogaster, considering co-expression of miRNAs and mRNAs in vivo The results obtained also do not support the decreasing model but are reasonably consistent with the increasing model of miRNA-target pairs. Furthermore, our large-scale analyses of currently available experimental data of miRNA-target pairs also showed a weak but the same trend in humans. These results indicate that the current decreasing model of miRNA-target pairs should be reconsidered and the increasing model may be more appropriate to explain the evolutionary transitions of miRNA-target pairs in many organisms. PMID:27189995

  11. MicroRNA expression profiles of porcine skeletal muscle.

    PubMed

    Zhou, B; Liu, H L; Shi, F X; Wang, J Y

    2010-10-01

    MicroRNAs (miRNAs) are endogenous non-coding RNAs of ∼22 nucleotides in length that play important roles in multiple biological processes by degrading targeted mRNAs or repressing mRNA translation. To evaluate the roles of miRNA in porcine skeletal muscle, miRNA expression profiles were investigated using longissimus muscle tissue from pigs at embryonic day 90 (E90) and postpartum day 120 (PD120). First, we used previously known miRNA sequences from humans and mice to perform blast searches against the porcine expressed sequence tag (EST) database; 98 new miRNA candidates were identified according to a range of filtering criteria. These miRNA candidates and 73 known miRNAs (miRBase 13.0) from pigs were chosen for porcine miRNA microarray analysis. A total of 16 newly identified miRNAs and 31 previously known miRNAs were detected in porcine skeletal muscle tissues. During later foetal development at E90, miR-1826, miR-26a, miR-199b and let-7 were highly expressed, whilst miR-1a, miR-133a, miR-26a and miR-1826 showed highest abundance during the fast growing stage at PD120. Using the 47 miRNAs detected by the microarray assay, we performed further investigations using the publicly available porcine mRNA database from NCBI and computed potential target hits using the software rnahybrid. This study identified 16 new miRNA candidates, computed potential target hits for 18 miRNA families and determined the miRNA expression profiles in porcine skeletal muscle tissues at different developmental stages. These results provide a valuable resource for investigators interested in post-transcriptional gene regulation in pigs and related animals. PMID:20331612

  12. Multicolor microRNA FISH effectively differentiates tumor types

    PubMed Central

    Renwick, Neil; Cekan, Pavol; Masry, Paul A.; McGeary, Sean E.; Miller, Jason B.; Hafner, Markus; Li, Zhen; Mihailovic, Aleksandra; Morozov, Pavel; Brown, Miguel; Gogakos, Tasos; Mobin, Mehrpouya B.; Snorrason, Einar L.; Feilotter, Harriet E.; Zhang, Xiao; Perlis, Clifford S.; Wu, Hong; Suárez-Fariñas, Mayte; Feng, Huichen; Shuda, Masahiro; Moore, Patrick S.; Tron, Victor A.; Chang, Yuan; Tuschl, Thomas

    2013-01-01

    MicroRNAs (miRNAs) are excellent tumor biomarkers because of their cell-type specificity and abundance. However, many miRNA detection methods, such as real-time PCR, obliterate valuable visuospatial information in tissue samples. To enable miRNA visualization in formalin-fixed paraffin-embedded (FFPE) tissues, we developed multicolor miRNA FISH. As a proof of concept, we used this method to differentiate two skin tumors, basal cell carcinoma (BCC) and Merkel cell carcinoma (MCC), with overlapping histologic features but distinct cellular origins. Using sequencing-based miRNA profiling and discriminant analysis, we identified the tumor-specific miRNAs miR-205 and miR-375 in BCC and MCC, respectively. We addressed three major shortcomings in miRNA FISH, identifying optimal conditions for miRNA fixation and ribosomal RNA (rRNA) retention using model compounds and high-pressure liquid chromatography (HPLC) analyses, enhancing signal amplification and detection by increasing probe-hapten linker lengths, and improving probe specificity using shortened probes with minimal rRNA sequence complementarity. We validated our method on 4 BCC and 12 MCC tumors. Amplified miR-205 and miR-375 signals were normalized against directly detectable reference rRNA signals. Tumors were classified using predefined cutoff values, and all were correctly identified in blinded analysis. Our study establishes a reliable miRNA FISH technique for parallel visualization of differentially expressed miRNAs in FFPE tumor tissues. PMID:23728175

  13. Sequence-non-specific effects of RNA interference triggers and microRNA regulators

    PubMed Central

    Olejniczak, Marta; Galka, Paulina; Krzyzosiak, Wlodzimierz J.

    2010-01-01

    RNA reagents of diverse lengths and structures, unmodified or containing various chemical modifications are powerful tools of RNA interference and microRNA technologies. These reagents which are either delivered to cells using appropriate carriers or are expressed in cells from suitable vectors often cause unintended sequence-non-specific immune responses besides triggering intended sequence-specific silencing effects. This article reviews the present state of knowledge regarding the cellular sensors of foreign RNA, the signaling pathways these sensors mobilize and shows which specific features of the RNA reagents set the responsive systems on alert. The representative examples of toxic effects caused in the investigated cell lines and tissues by the RNAs of specific types and structures are collected and may be instructive for further studies of sequence-non-specific responses to foreign RNA in human cells. PMID:19843612

  14. Human Argonaute 2 Is Tethered to Ribosomal RNA through MicroRNA Interactions.

    PubMed

    Atwood, Blake L; Woolnough, Jessica L; Lefevre, Gaelle M; Saint Just Ribeiro, Mariana; Felsenfeld, Gary; Giles, Keith E

    2016-08-19

    The primary role of the RNAi machinery is to promote mRNA degradation within the cytoplasm in a microRNA-dependent manner. However, both Dicer and the Argonaute protein family have expanded roles in gene regulation within the nucleus. To further our understanding of this role, we have identified chromatin binding sites for AGO2 throughout the 45S region of the human rRNA gene. The location of these sites was mirrored by the positions of AGO2 cross-linking sites identified via PAR-CLIP-seq. AGO2 binding to the rRNA within the nucleus was confirmed by RNA immunoprecipitation and quantitative-PCR. To explore a possible mechanism by which AGO2 could be recruited to the rRNA, we identified 1174 regions within the 45S rRNA transcript that have the ability to form a perfect duplex with position 2-6 (seed sequence) of each microRNA expressed in HEK293T cells. Of these potential AGO2 binding sites, 479 occurred within experimentally verified AGO2-rRNA cross-linking sites. The ability of AGO2 to cross-link to rRNA was almost completely lost in a DICER knock-out cell line. The transfection of miR-92a-2-3p into the noDICE cell line facilitated AGO2 cross-linking at a region of the rRNA that has a perfect seed match at positions 3-8, including a single G-U base pair. Knockdown of AGO2 within HEK293T cells causes a slight, but statistically significant increase in the overall rRNA synthesis rate but did not impact the ratio of processing intermediates or the recruitment of the Pol I transcription factor UBTF. PMID:27288410

  15. Current Status and Strategy of microRNA Research for Cartilage Development and Osteoarthritis Pathogenesis

    PubMed Central

    2016-01-01

    MicroRNAs (miRNAs), which are small (~21 nucleotides) non-coding RNAs, are important players in endochondral ossification, articular cartilage homeostasis, and arthritis pathogenesis. Comprehensive and genetic analyses of cartilage-specific or cartilage-related miRNAs have provided new information on cartilage development, homeostasis, and related diseases. State-of-the-art combinatorial approaches, including transcription-activator like effector nuclease (TALEN)/clustered regularly interspaced short palindromic repeats (CRISPR) technique for targeting miRNAs and high-throughput sequencing of RNA isolated by cross-linking immunoprecipitation for identifying target messenger RNAs, should be used to determine complex miRNA networks and miRNA-dependent cartilage regulation. Use of advanced drug delivery systems involving cartilage-specific miRNAs will accelerate the application of these new findings in arthritis therapy. PMID:27622175

  16. Current Status and Strategy of microRNA Research for Cartilage Development and Osteoarthritis Pathogenesis.

    PubMed

    Asahara, Hiroshi

    2016-08-01

    MicroRNAs (miRNAs), which are small (~21 nucleotides) non-coding RNAs, are important players in endochondral ossification, articular cartilage homeostasis, and arthritis pathogenesis. Comprehensive and genetic analyses of cartilage-specific or cartilage-related miRNAs have provided new information on cartilage development, homeostasis, and related diseases. State-of-the-art combinatorial approaches, including transcription-activator like effector nuclease (TALEN)/clustered regularly interspaced short palindromic repeats (CRISPR) technique for targeting miRNAs and high-throughput sequencing of RNA isolated by cross-linking immunoprecipitation for identifying target messenger RNAs, should be used to determine complex miRNA networks and miRNA-dependent cartilage regulation. Use of advanced drug delivery systems involving cartilage-specific miRNAs will accelerate the application of these new findings in arthritis therapy. PMID:27622175

  17. Vulnerability of microRNA biogenesis in FTD-ALS.

    PubMed

    Eitan, Chen; Hornstein, Eran

    2016-09-15

    The genetics of the neurodegenerative diseases amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) turn our attention to RNA metabolism, primarily because many of the identified diseases-associated genes encode for RNA-binding proteins. microRNAs (miRNAs) are endogenous noncoding RNAs that play critical roles in maintaining brain integrity. The current review sheds light on miRNA dysregulation in neurodegenerative diseases, focusing on FTD-ALS. We propose that miRNAs are susceptible to fail when protein factors that are critical for miRNA biogenesis malfunction. Accordingly, potential insufficiencies of the 'microprocessor' complex, the nucleo-cytoplasmic export of miRNA precursors or their processing by Dicer were recently reported. Furthermore, specific miRNAs are involved in the regulation of pathways that are essential for neuronal survival or function. Any change in the expression of these specific miRNAs or in their ability to recognize their target sequences will have negative consequences. Taken together, recent reports strengthens the hypothesis that dysregulation of miRNAs might play an important role in the pathogenesis of neurodegenerative diseases, and highlights the miRNA biogenesis machinery as an interesting target for therapeutic interventions for ALS as well as FTD. This article is part of a Special Issue entitled SI:RNA Metabolism in Disease. PMID:26778173

  18. microRNA Processing Pathway Regulates Olfactory Neuron Morphogenesis

    PubMed Central

    Berdnik, Daniela; Fan, Audrey P.; Potter, Christopher J.; Luo, Liqun

    2008-01-01

    Summary The micro(mi)RNA processing pathway produces miRNAs as posttranscriptional regulators of gene expression. The nuclear RNase III Drosha catalyzes the first processing step together with the dsRNA binding protein DGCR8/Pasha generating pre-miRNAs [1, 2]. The next cleavage employs the cytoplasmic RNase III Dicer producing miRNA duplexes [3, 4]. Finally, Argonautes are recruited with miRNAs into an RNA-induced silencing complex for mRNA recognition (Figure 1A). Here, we identify two members of the miRNA pathway, Pasha and Dicer-1, in a forward genetic screen for mutations that disrupt wiring specificity of Drosophila olfactory projection neurons (PNs). The olfactory system is built as discrete map of highly stereotyped neuronal connections [5, 6]. Each PN targets dendrites to a specific glomerulus in the antennal lobe and projects axons stereotypically into higher brain centers [7–9]. In selected PN classes, pasha and Dicer-1 mutants cause specific PN dendrite mistargeting in the antennal lobe and altered axonal terminations in higher brain centers. Furthermore, Pasha and Dicer-1 act cell-autonomously in postmitotic neurons to regulate dendrite and axon targeting during development. However, Argonaute-1 and Argonaute-2 are dispensable for PN morphogenesis. Our findings suggest a role for the miRNA processing pathway in establishing wiring specificity in the nervous system. PMID:19013069

  19. miEAA: microRNA enrichment analysis and annotation.

    PubMed

    Backes, Christina; Khaleeq, Qurratulain T; Meese, Eckart; Keller, Andreas

    2016-07-01

    Similar to the development of gene set enrichment and gene regulatory network analysis tools over a decade ago, microRNA enrichment tools are currently gaining importance. Building on our experience with the gene set analysis toolkit GeneTrail, we implemented the miRNA Enrichment Analysis and Annotation tool (miEAA). MiEAA is a web-based application that offers a variety of commonly applied statistical tests such as over-representation analysis and miRNA set enrichment analysis, which is similar to Gene Set Enrichment Analysis. Besides the different statistical tests, miEAA also provides rich functionality in terms of miRNA categories. Altogether, over 14 000 miRNA sets have been added, including pathways, diseases, organs and target genes. Importantly, our tool can be applied for miRNA precursors as well as mature miRNAs. To make the tool as useful as possible we additionally implemented supporting tools such as converters between different miRBase versions and converters from miRNA names to precursor names. We evaluated the performance of miEAA on two sets of miRNAs that are affected in lung adenocarcinomas and have been detected by array analysis. The web-based application is freely accessible at: http://www.ccb.uni-saarland.de/mieaa_tool/. PMID:27131362

  20. miEAA: microRNA enrichment analysis and annotation

    PubMed Central

    Backes, Christina; Khaleeq, Qurratulain T.; Meese, Eckart; Keller, Andreas

    2016-01-01

    Similar to the development of gene set enrichment and gene regulatory network analysis tools over a decade ago, microRNA enrichment tools are currently gaining importance. Building on our experience with the gene set analysis toolkit GeneTrail, we implemented the miRNA Enrichment Analysis and Annotation tool (miEAA). MiEAA is a web-based application that offers a variety of commonly applied statistical tests such as over-representation analysis and miRNA set enrichment analysis, which is similar to Gene Set Enrichment Analysis. Besides the different statistical tests, miEAA also provides rich functionality in terms of miRNA categories. Altogether, over 14 000 miRNA sets have been added, including pathways, diseases, organs and target genes. Importantly, our tool can be applied for miRNA precursors as well as mature miRNAs. To make the tool as useful as possible we additionally implemented supporting tools such as converters between different miRBase versions and converters from miRNA names to precursor names. We evaluated the performance of miEAA on two sets of miRNAs that are affected in lung adenocarcinomas and have been detected by array analysis. The web-based application is freely accessible at: http://www.ccb.uni-saarland.de/mieaa_tool/. PMID:27131362

  1. MicroRNA Detection: Current Technology and Research Strategies

    NASA Astrophysics Data System (ADS)

    Hunt, Eric A.; Broyles, David; Head, Trajen; Deo, Sapna K.

    2015-07-01

    The relatively new field of microRNA (miR) has experienced rapid growth in methodology associated with its detection and bioanalysis as well as with its role in -omics research, clinical diagnostics, and new therapeutic strategies. The breadth of this area of research and the seemingly exponential increase in number of publications on the subject can present scientists new to the field with a daunting amount of information to evaluate. This review aims to provide a collective overview of miR detection methods by relating conventional, established techniques [such as quantitative reverse transcription polymerase chain reaction (RT-qPCR), microarray, and Northern blotting (NB)] and relatively recent advancements [such as next-generation sequencing (NGS), highly sensitive biosensors, and computational prediction of microRNA/targets] to common miR research strategies. This should guide interested readers toward a more focused study of miR research and the surrounding technology.

  2. MicroRNA Profiles Discriminate among Colon Cancer Metastasis

    PubMed Central

    Drusco, Alessandra; Nuovo, Gerard J.; Zanesi, Nicola; Di Leva, Gianpiero; Pichiorri, Flavia; Volinia, Stefano; Fernandez, Cecilia; Antenucci, Anna; Costinean, Stefan; Bottoni, Arianna; Rosito, Immacolata A.; Liu, Chang-Gong; Burch, Aaron; Acunzo, Mario; Pekarsky, Yuri; Alder, Hansjuerg; Ciardi, Antonio; Croce, Carlo M.

    2014-01-01

    MicroRNAs are being exploited for diagnosis, prognosis and monitoring of cancer and other diseases. Their high tissue specificity and critical role in oncogenesis provide new biomarkers for the diagnosis and classification of cancer as well as predicting patients' outcomes. MicroRNAs signatures have been identified for many human tumors, including colorectal cancer (CRC). In most cases, metastatic disease is difficult to predict and to prevent with adequate therapies. The aim of our study was to identify a microRNA signature for metastatic CRC that could predict and differentiate metastatic target organ localization. Normal and cancer tissues of three different groups of CRC patients were analyzed. RNA microarray and TaqMan Array analysis were performed on 66 Italian patients with or without lymph nodes and/or liver recurrences. Data obtained with the two assays were analyzed separately and then intersected to identify a primary CRC metastatic signature. Five differentially expressed microRNAs (hsa-miR-21, -103, -93, -31 and -566) were validated by qRT-PCR on a second group of 16 American metastatic patients. In situ hybridization was performed on the 16 American patients as well as on three distinct commercial tissues microarray (TMA) containing normal adjacent colon, the primary adenocarcinoma, normal and metastatic lymph nodes and liver. Hsa-miRNA-21, -93, and -103 upregulation together with hsa-miR-566 downregulation defined the CRC metastatic signature, while in situ hybridization data identified a lymphonodal invasion profile. We provided the first microRNAs signature that could discriminate between colorectal recurrences to lymph nodes and liver and between colorectal liver metastasis and primary hepatic tumor. PMID:24921248

  3. Widespread context dependency of microRNA-mediated regulation

    PubMed Central

    Erhard, Florian; Haas, Jürgen; Lieber, Diana; Malterer, Georg; Jaskiewicz, Lukasz; Zavolan, Mihaela; Dölken, Lars; Zimmer, Ralf

    2014-01-01

    Gene expression is regulated in a context-dependent, cell-type-specific manner. Condition-specific transcription is dependent on the presence of transcription factors (TFs) that can activate or inhibit its target genes (global context). Additional factors, such as chromatin structure, histone, or DNA modifications, also influence the activity of individual target genes (individual context). The role of the global and individual context for post-transcriptional regulation has not systematically been investigated on a large scale and is poorly understood. Here we show that global and individual context dependency is a pervasive feature of microRNA-mediated regulation. Our comprehensive and highly consistent data set from several high-throughput technologies (PAR-CLIP, RIP-chip, 4sU-tagging, and SILAC) provides strong evidence that context-dependent microRNA target sites (CDTS) are as frequent and functionally relevant as constitutive target sites (CTS). Furthermore, we found the global context to be insufficient to explain the CDTS, and that flanking sequence motifs provide individual context that is an equally important factor. Our results demonstrate that, similar to TF-mediated regulation, global and individual context dependency are prevalent in microRNA-mediated gene regulation, implying a much more complex post-transcriptional regulatory network than is currently known. The necessary tools to unravel post-transcriptional regulations and mechanisms need to be much more involved, and much more data will be needed for particular cell types and cellular conditions in order to understand microRNA-mediated regulation and the context-dependent post-transcriptional regulatory network. PMID:24668909

  4. Widespread context dependency of microRNA-mediated regulation.

    PubMed

    Erhard, Florian; Haas, Jürgen; Lieber, Diana; Malterer, Georg; Jaskiewicz, Lukasz; Zavolan, Mihaela; Dölken, Lars; Zimmer, Ralf

    2014-06-01

    Gene expression is regulated in a context-dependent, cell-type-specific manner. Condition-specific transcription is dependent on the presence of transcription factors (TFs) that can activate or inhibit its target genes (global context). Additional factors, such as chromatin structure, histone, or DNA modifications, also influence the activity of individual target genes (individual context). The role of the global and individual context for post-transcriptional regulation has not systematically been investigated on a large scale and is poorly understood. Here we show that global and individual context dependency is a pervasive feature of microRNA-mediated regulation. Our comprehensive and highly consistent data set from several high-throughput technologies (PAR-CLIP, RIP-chip, 4sU-tagging, and SILAC) provides strong evidence that context-dependent microRNA target sites (CDTS) are as frequent and functionally relevant as constitutive target sites (CTS). Furthermore, we found the global context to be insufficient to explain the CDTS, and that flanking sequence motifs provide individual context that is an equally important factor. Our results demonstrate that, similar to TF-mediated regulation, global and individual context dependency are prevalent in microRNA-mediated gene regulation, implying a much more complex post-transcriptional regulatory network than is currently known. The necessary tools to unravel post-transcriptional regulations and mechanisms need to be much more involved, and much more data will be needed for particular cell types and cellular conditions in order to understand microRNA-mediated regulation and the context-dependent post-transcriptional regulatory network. PMID:24668909

  5. RNA Secondary Structure Modulates FMRP’s Bi-Functional Role in the MicroRNA Pathway

    PubMed Central

    Kenny, Phillip; Ceman, Stephanie

    2016-01-01

    MicroRNAs act by post-transcriptionally regulating the gene expression of 30%–60% of mammalian genomes. MicroRNAs are key regulators in all cellular processes, though the mechanism by which the cell activates or represses microRNA-mediated translational regulation is poorly understood. In this review, we discuss the RNA binding protein Fragile X Mental Retardation Protein (FMRP) and its role in microRNA-mediated translational regulation. Historically, FMRP is known to function as a translational suppressor. However, emerging data suggests that FMRP has both an agonistic and antagonistic role in regulating microRNA-mediated translational suppression. This bi-functional role is dependent on FMRP’s interaction with the RNA helicase Moloney leukemia virus 10 (MOV10), which modifies the structural landscape of bound mRNA, therefore facilitating or inhibiting its association with the RNA-Induced Silencing Complex. PMID:27338369

  6. Evolution of the let-7 microRNA Family

    PubMed Central

    Hertel, Jana; Bartschat, Sebastian; Wintsche, Axel; Otto, Christian; of the Bioinformatics Computer Lab, The Students; Stadler, Peter F.

    2012-01-01

    The increase of bodyplan complexity in early bilaterian evolution is correlates with the advent and diversification of microRNAs. These small RNAs guide animal development by regulating temporal transitions in gene expression involved in cell fate choices and transitions between pluripotency and differentiation. One of the two known microRNAs whose origins date back before the bilaterian ancestor is mir-100. In Bilateria, it appears stably associated in polycistronic transcripts with let-7 and mir-125, two key regulators of development. In vertebrates, these three microRNA families have expanded to form a complex system of developmental regulators. In this contribution, we disentangle the evolutionary history of the let-7 locus, which was restructured independently in nematodes, platyhelminths, and deuterostomes. The foundation of a second let-7 locus in the common ancestor of vertebrates and urochordates predates the vertebrate-specific genome duplications, which then caused a rapid expansion of the let-7 family. PMID:22617875

  7. MicroRNA-429 Modulates Hepatocellular Carcinoma Prognosis and Tumorigenesis

    PubMed Central

    Huang, Xiao-Ying; Yao, Jin-Guang; Wang, Chao; Ma, Yun; Xia, Qiang

    2013-01-01

    MicroRNA-429 (miR-429) may modify the development and progression of cancers; however, the role of this microRNA in the hepatocellular carcinoma (HCC) has not been well elaborated. Here, we tested miR-429 expression in 138 pathology-diagnosed HCC cases and SMMC-7721 cells. We found that miR-429 was upregulated in HCC tumor tissues and that the high expression of miR-429 was significantly correlated with larger tumor size (odd ratio (OR), 2.70; 95% confidence interval (CI), 1.28–5.56) and higher aflatoxin B1-DNA adducts (OR = 3.13, 95% CI = 1.47–6.67). Furthermore, this microRNA overexpression modified the recurrence-free survival and overall survival of HCC patients. Functionally, miR-429 overexpression progressed tumor cells proliferation and inhibited cell apoptosis. These results indicate for the first time that miR-429 may modify HCC prognosis and tumorigenesis and may be a potential tumor therapeutic target. PMID:24204382

  8. MicroRNA-21 Increases Proliferation and Cisplatin Sensitivity of Osteosarcoma-Derived Cells.

    PubMed

    Vanas, Vanita; Haigl, Barbara; Stockhammer, Verena; Sutterlüty-Fall, Hedwig

    2016-01-01

    Osteosarcoma is the most common primary bone tumor and poor prognosis for osteosarcoma patients is mainly due to chemotherapy resistance. MicroRNAs are important to maintain pathophysiological mechanisms of cancer and influence cell sensitivity to chemotherapy. In this study, we tested the functions of microRNA-21 for malignant features as well as for drug resistance of osteosarcoma. We used Northern blot to measure microRNA-21 levels in osteosarcoma-derived cell lines. MicroRNA-21 activity was modulated by either expressing a sponge to decrease its activity in an osteosarcoma-derived cell line expressing high levels of microRNA-21 or by introducing pri-microRNA-21 in a cell line with low endogenous levels. Cell migration was determined in a scratch assay and cell proliferation was measured by performing growth curve analysis. Sensitivity of the cells towards chemotherapeutics was investigated by performing cell viability assays and calculating the IC50 values. While cell migration was unaffected by modulated microRNA-21 levels, microRNA-21 inhibition slowed proliferation and exogenously expressed microRNA-21 promoted this process. Modulated microRNA-21 activity failed to effect sensitivity of osteosarcoma-derived cell lines to doxorubicin or methotrexate. Contrarily, reduction of microRNA-21 activity resulted in enhanced resistance towards cisplatin while ectopic expression of microRNA-21 showed the opposite effect. Increased microRNA-21 levels repressed the expression of Sprouty2 and ectopic expression of Sprouty2 was able to largely rescue the observed effects of microRNA-21 in osteosarcoma. In summary, our data indicate that in osteosarcoma microRNA-21 expression is an important component for regulation of cell proliferation and for determining sensitivity to cisplatin. PMID:27513462

  9. MicroRNA-21 Increases Proliferation and Cisplatin Sensitivity of Osteosarcoma-Derived Cells

    PubMed Central

    Vanas, Vanita; Haigl, Barbara; Stockhammer, Verena; Sutterlüty-Fall, Hedwig

    2016-01-01

    Osteosarcoma is the most common primary bone tumor and poor prognosis for osteosarcoma patients is mainly due to chemotherapy resistance. MicroRNAs are important to maintain pathophysiological mechanisms of cancer and influence cell sensitivity to chemotherapy. In this study, we tested the functions of microRNA-21 for malignant features as well as for drug resistance of osteosarcoma. We used Northern blot to measure microRNA-21 levels in osteosarcoma-derived cell lines. MicroRNA-21 activity was modulated by either expressing a sponge to decrease its activity in an osteosarcoma-derived cell line expressing high levels of microRNA-21 or by introducing pri-microRNA-21 in a cell line with low endogenous levels. Cell migration was determined in a scratch assay and cell proliferation was measured by performing growth curve analysis. Sensitivity of the cells towards chemotherapeutics was investigated by performing cell viability assays and calculating the IC50 values. While cell migration was unaffected by modulated microRNA-21 levels, microRNA-21 inhibition slowed proliferation and exogenously expressed microRNA-21 promoted this process. Modulated microRNA-21 activity failed to effect sensitivity of osteosarcoma-derived cell lines to doxorubicin or methotrexate. Contrarily, reduction of microRNA-21 activity resulted in enhanced resistance towards cisplatin while ectopic expression of microRNA-21 showed the opposite effect. Increased microRNA-21 levels repressed the expression of Sprouty2 and ectopic expression of Sprouty2 was able to largely rescue the observed effects of microRNA-21 in osteosarcoma. In summary, our data indicate that in osteosarcoma microRNA-21 expression is an important component for regulation of cell proliferation and for determining sensitivity to cisplatin. PMID:27513462

  10. MicroRNA Regulation in Systemic Lupus Erythematosus Pathogenesis

    PubMed Central

    Yan, Sheng; Yim, Lok Yan; Lu, Liwei; Lau, Chak Sing

    2014-01-01

    MicroRNAs (miRNAs) are endogenous small RNA molecules best known for their function in post-transcriptional gene regulation. Immunologically, miRNA regulates the differentiation and function of immune cells and its malfunction contributes to the development of various autoimmune diseases including systemic lupus erythematosus (SLE). Over the last decade, accumulating researches provide evidence for the connection between dysregulated miRNA network and autoimmunity. Interruption of miRNA biogenesis machinery contributes to the abnormal T and B cell development and particularly a reduced suppressive function of regulatory T cells, leading to systemic autoimmune diseases. Additionally, multiple factors under autoimmune conditions interfere with miRNA generation via key miRNA processing enzymes, thus further skewing the miRNA expression profile. Indeed, several independent miRNA profiling studies reported significant differences between SLE patients and healthy controls. Despite the lack of a consistent expression pattern on individual dysregulated miRNAs in SLE among these studies, the aberrant expression of distinct groups of miRNAs causes overlapping functional outcomes including perturbed type I interferon signalling cascade, DNA hypomethylation and hyperactivation of T and B cells. The impact of specific miRNA-mediated regulation on function of major immune cells in lupus is also discussed. Although research on the clinical application of miRNAs is still immature, through an integrated approach with advances in next generation sequencing, novel tools in bioinformatics database analysis and new in vitro and in vivo models for functional evaluation, the diagnostic and therapeutic potentials of miRNAs may bring to fruition in the future. PMID:24999310

  11. Capturing microRNA targets using an RNA-induced silencing complex (RISC)-trap approach

    PubMed Central

    Cambronne, Xiaolu A.; Shen, Rongkun; Auer, Paul L.; Goodman, Richard H.

    2012-01-01

    Identifying targets is critical for understanding the biological effects of microRNA (miRNA) expression. The challenge lies in characterizing the cohort of targets for a specific miRNA, especially when targets are being actively down-regulated in miRNA– RNA-induced silencing complex (RISC)–messengerRNA (mRNA) complexes. We have developed a robust and versatile strategy called RISCtrap to stabilize and purify targets from this transient interaction. Its utility was demonstrated by determining specific high-confidence target datasets for miR-124, miR-132, and miR-181 that contained known and previously unknown transcripts. Two previously unknown miR-132 targets identified with RISCtrap, adaptor protein CT10 regulator of kinase 1 (CRK1) and tight junction-associated protein 1 (TJAP1), were shown to be endogenously regulated by miR-132 in adult mouse forebrain. The datasets, moreover, differed in the number of targets and in the types and frequency of microRNA recognition element (MRE) motifs, thus revealing a previously underappreciated level of specificity in the target sets regulated by individual miRNAs. PMID:23184980

  12. Genomic architecture and inheritance of human ribosomal RNA gene clusters

    PubMed Central

    Stults, Dawn M.; Killen, Michael W.; Pierce, Heather H.; Pierce, Andrew J.

    2008-01-01

    The finishing of the Human Genome Project largely completed the detailing of human euchromatic sequences; however, the most highly repetitive regions of the genome still could not be assembled. The 12 gene clusters producing the structural RNA components of the ribosome are critically important for cellular viability, yet fall into this unassembled region of the Human Genome Project. To determine the extent of human variation in ribosomal RNA gene content (rDNA) and patterns of rDNA cluster inheritance, we have determined the physical lengths of the rDNA clusters in peripheral blood white cells of healthy human volunteers. The cluster lengths exhibit striking variability between and within human individuals, ranging from 50 kb to >6 Mb, manifest essentially complete heterozygosity, and provide each person with their own unique rDNA electrophoretic karyotype. Analysis of these rDNA fingerprints in multigenerational human families demonstrates that the rDNA clusters are subject to meiotic rearrangement at a frequency >10% per cluster, per meiosis. With this high intrinsic recombinational instability, the rDNA clusters may serve as a unique paradigm of potential human genomic plasticity. PMID:18025267

  13. Breast cancer targeting novel microRNA-nanoparticles for imaging

    NASA Astrophysics Data System (ADS)

    Natarajan, Arutselvan; Venugopal, Senthil K.; DeNardo, Sally J.; Zern, Mark A.

    2009-02-01

    MicroRNAs (miRNAs) are one of the most prevalent small (~22 nucleotide) regulatory RNA classes in animals. These miRNAs constitute nearly one percent of genes in the human genome, making miRNA genes one of the more abundant types of regulatory molecules. MiRNAs have been shown to play important roles in cell development, apoptosis, and other fundamental biological processes. MiRNAs exert their influence through complementary base-pairing with specific target mRNAs, leading to degradation or translational repression of the targeted mRNA. We have identified and tested a novel microRNA (miR-491) and demonstrated increased apoptosis in hepatocellular carcinoma cells (HepG2) and in human breast cancer cells (HBT3477) in vitro. We prepared a novel cancer targeting assembly of gold nanoparticles (GNP) with Quantum dots, miR-491, and MAb-ChL6 coupled through streptavidin/biotin for effective transfection, and to induce apoptosis in specific cancer cells for imaging and targeted therapy. The targeting and apoptosis inducing ability was tested by confocal and electron microscopy. The MAb-GNP-miR491-Qdot construct effectively transfected into the HBT3477 cells and induced apoptosis the confirmation of these results would suggest a new class of molecules for the imaging and therapy of breast cancer.

  14. Regulation of Senescence by microRNA Biogenesis Factors

    PubMed Central

    Abdelmohsen, Kotb; Srikantan, Subramanya; Kang, Min-Ju; Gorospe, Myriam

    2012-01-01

    Senescence represents a state of indefinite growth arrest in cells that have reached their replicative life span, have become damaged, or express aberrant levels of cancer-related proteins. While senescence is widely considered to represent tumor-suppressive mechanism, the accumulation of senescent cells in tissues of older organisms is believed to underlie age-associated losses in physiologic function and age-related diseases. With the emergence of microRNAs (miRNAs) as a major class of molecular regulators of senescence, we review the transcriptional and post-transcriptional factors that control senescence-associated microRNA biosynthesis. Focusing on their enhancement or repression of senescence, we describe the transcription factors that govern the synthesis of primary (pri-)miRNAs, the proteins that control the nuclear processing of pri-miRNAs into precursor (pre-)miRNAs, including RNA editing enzymes, RNases, and RNA helicases, and the cytoplasmic proteins that affect the final processing of pre-miRNAs into mature miRNAs. We discuss how miRNA biogenesis proteins enhance or repress senescence, and thus influence the senescent phenotype that affects normal tissue function and pathology. PMID:22306790

  15. Efficient synthesis of stably adenylated DNA and RNA adapters for microRNA capture using T4 RNA ligase 1.

    PubMed

    Song, Yunke; Liu, Kelvin J; Wang, Tza-Huei

    2015-01-01

    MicroRNA profiling methods have become increasingly important due to the rapid rise of microRNA in both basic and translational sciences. A critical step in many microRNA profiling assays is adapter ligation using pre-adenylated adapters. While pre-adenylated adapters can be chemically or enzymatically prepared, enzymatic adenylation is preferred due to its ease and high yield. However, previously reported enzymatic methods either require tedious purification steps or use thermostable ligases that can generate side products during the subsequent ligation step. We have developed a highly efficient, template- and purification-free, adapter adenylation method using T4 RNA ligase 1. This method is capable of adenylating large amounts of adapter at ~100% efficiency and can efficiently adenylate both DNA and RNA bases. We find that the adenylation reaction speed can differ between DNA and RNA and between terminal nucleotides, leading to bias if reactions are not allowed to run to completion. We further find that the addition of high PEG levels can effectively suppress these differences. PMID:26500066

  16. MicroRNA Biomarkers for Coronary Artery Disease?

    PubMed

    Kaudewitz, Dorothee; Zampetaki, Anna; Mayr, Manuel

    2015-12-01

    MicroRNA (miRNA, miR) measurements in patients with coronary heart disease are hampered by the confounding effects of medication commonly used in cardiovascular patients such as statins, antiplatelet drugs, and heparin administration. Statins reduce the circulating levels of liver-derived miR-122. Antiplatelet medication attenuates the release of platelet-derived miRNAs. Heparin inhibits the polymerase chain reactions, in particular the amplification of the exogenous Caenorhabditis elegans spike-in control, thereby resulting in an artefactual rise of endogenous miRNAs. As these limitations have not been previously recognised, a reevaluation of the current miRNA literature, in particular of case-control studies in patients with cardiovascular disease or coronary interventions, is required. PMID:26490079

  17. A Review of Computational Tools in microRNA Discovery

    PubMed Central

    Gomes, Clarissa P. C.; Cho, Ji-Hoon; Hood, Leroy; Franco, Octávio L.; Pereira, Rinaldo W.; Wang, Kai

    2013-01-01

    Since microRNAs (miRNAs) were discovered, their impact on regulating various biological activities has been a surprising and exciting field. Knowing the entire repertoire of these small molecules is the first step to gain a better understanding of their function. High throughput discovery tools such as next-generation sequencing significantly increased the number of known miRNAs in different organisms in recent years. However, the process of being able to accurately identify miRNAs is still a complex and difficult task, requiring the integration of experimental approaches with computational methods. A number of prediction algorithms based on characteristics of miRNA molecules have been developed to identify new miRNA species. Different approaches have certain strengths and weaknesses and in this review, we aim to summarize several commonly used tools in metazoan miRNA discovery. PMID:23720668

  18. MicroRNA-mediated repression of nonsense mRNAs

    PubMed Central

    Zhao, Ya; Lin, Jimin; Xu, Beiying; Hu, Sida; Zhang, Xue; Wu, Ligang

    2014-01-01

    Numerous studies have established important roles for microRNAs (miRNAs) in regulating gene expression. Here, we report that miRNAs also serve as a surveillance system to repress the expression of nonsense mRNAs that may produce harmful truncated proteins. Upon recognition of the premature termination codon by the translating ribosome, the downstream portion of the coding region of an mRNA is redefined as part of the 3′ untranslated region; as a result, the miRNA-responsive elements embedded in this region can be detected by miRNAs, triggering accelerated mRNA deadenylation and translational inhibition. We demonstrate that naturally occurring cancer-causing APC (adenomatous polyposis coli) nonsense mutants which escape nonsense-mediated mRNA decay (NMD) are repressed by miRNA-mediated surveillance. In addition, we show that miRNA-mediated surveillance and exon–exon junction complex-mediated NMD are not mutually exclusive and act additively to enhance the repressive activity. Therefore, we have uncovered a new role for miRNAs in repressing nonsense mutant mRNAs. DOI: http://dx.doi.org/10.7554/eLife.03032.001 PMID:25107276

  19. MicroRNA in autoimmunity and autoimmune diseases

    PubMed Central

    Pauley, Kaleb M.; Cha, Seunghee; Chan, Edward K.L.

    2009-01-01

    MicroRNAs (miRNAs) are small conserved non-coding RNA molecules that post-transcriptionally regulate gene expression by targeting the 3′ untranslated region (UTR) of specific messenger RNAs (mRNAs) for degradation or translational repression. miRNA-mediated gene regulation is critical for normal cellular functions such as the cell cycle, differentiation, and apoptosis, and as much as one-third of human mRNAs may be miRNA targets. Emerging evidence has demonstrated that miRNAs play a vital role in the regulation of immunological functions and the prevention of autoimmunity. Here we review the many newly discovered roles of miRNA regulation in immune functions and in the development of autoimmunity and autoimmune disease. Specifically, we discuss the involvement of miRNA regulation in innate and adaptive immune responses, immune cell development, T regulatory cell stability and function, and differential miRNA expression in rheumatoid arthritis and systemic lupus erythematosus. PMID:19303254

  20. A Dynamic Search Process Underlies MicroRNA Targeting

    PubMed Central

    Chandradoss, Stanley D.; MacRae, Ian J.; Joo, Chirlmin

    2016-01-01

    Summary Argonaute proteins play a central role in mediating post-transcriptional gene regulation by microRNAs (miRNAs). Argonautes use the nucleotide sequences in miRNAs as guides for identifying target messenger RNAs for repression. Here we used single-molecule FRET to directly visualize how human Argonaute-2 (Ago2) searches for and identifies target sites in RNAs complementary to its miRNA guide. Our results suggest that Ago2 initially scans for target sites with complementarity to nucleotides 2–4 of the miRNA. This initial transient interaction propagates into a stable association when target complementarity extends to nucleotides 2–8. This stepwise recognition process is coupled to lateral diffusion of Ago2 along the target RNA, which promotes target search by enhancing the retention of Ago2 on the RNA. The combined results reveal the mechanisms that Argonaute likely uses to efficiently identify miRNA target sites within the vast and dynamic agglomeration of RNA molecules in the living cell. PMID:26140593

  1. A Dynamic Search Process Underlies MicroRNA Targeting.

    PubMed

    Chandradoss, Stanley D; Schirle, Nicole T; Szczepaniak, Malwina; MacRae, Ian J; Joo, Chirlmin

    2015-07-01

    Argonaute proteins play a central role in mediating post-transcriptional gene regulation by microRNAs (miRNAs). Argonautes use the nucleotide sequences in miRNAs as guides for identifying target messenger RNAs for repression. Here, we used single-molecule FRET to directly visualize how human Argonaute-2 (Ago2) searches for and identifies target sites in RNAs complementary to its miRNA guide. Our results suggest that Ago2 initially scans for target sites with complementarity to nucleotides 2-4 of the miRNA. This initial transient interaction propagates into a stable association when target complementarity extends to nucleotides 2-8. This stepwise recognition process is coupled to lateral diffusion of Ago2 along the target RNA, which promotes the target search by enhancing the retention of Ago2 on the RNA. The combined results reveal the mechanisms that Argonaute likely uses to efficiently identify miRNA target sites within the vast and dynamic agglomeration of RNA molecules in the living cell. PMID:26140593

  2. Experimental strategies for microRNA target identification

    PubMed Central

    Thomson, Daniel W.; Bracken, Cameron P.; Goodall, Gregory J.

    2011-01-01

    MicroRNAs (miRNAs) are important regulators of eukaryotic gene expression in most biological processes. They act by guiding the RNAi-induced silencing complex (RISC) to partially complementary sequences in target mRNAs to suppress gene expression by a combination of translation inhibition and mRNA decay. The commonly accepted mechanism of miRNA targeting in animals involves an interaction between the 5′-end of the miRNA called the ‘seed region’ and the 3′ untranslated region (3′-UTR) of the mRNA. Many target prediction algorithms are based around such a model, though increasing evidence demonstrates that targeting can also be mediated through sites other than the 3′-UTR and that seed region base pairing is not always required. The power and validity of such in silico data can be therefore hindered by the simplified rules used to represent targeting interactions. Experimentation is essential to identify genuine miRNA targets, however many experimental modalities exist and their limitations need to be understood. This review summarizes and critiques the existing experimental techniques for miRNA target identification. PMID:21652644

  3. Using artificial microRNA sponges to achieve microRNA loss-of-function in cancer cells.

    PubMed

    Tay, Felix Chang; Lim, Jia Kai; Zhu, Haibao; Hin, Lau Cia; Wang, Shu

    2015-01-01

    Widely observed dysregulation of microRNAs (miRNAs) in human cancer has led to substantial speculation regarding possible functions of these short, non-coding RNAs in cancer development and manipulation of miRNA expression to treat cancer. To achieve miRNA loss-of-function, miRNA sponge technology has been developed to use plasmid or viral vectors for intracellular expression of tandemly arrayed, bulged miRNA binding sites complementary to a miRNA target to saturate its ability to regulate natural mRNAs. A strong viral promoter can be used in miRNA sponge vectors to generate high-level expression of the competitive inhibitor transcripts for either transient or long-term inhibition of miRNA function. Taking the advantage of sharing a common seed sequence by members of a miRNA family, this technology is especially useful in knocking down the expression of a family of miRNAs, providing a powerful means for simultaneous inhibition of multiple miRNAs of interest with a single inhibitor. Knockdown of overexpressed oncogenic miRNAs with the technology can be a rational therapeutic strategy for cancer, whereas inhibition of tumor-suppressive miRNAs by the sponges will be useful in deciphering functions of miRNAs in oncogenesis. Herein, we discuss the design of miRNA sponge expression vectors and the use of the vectors to gain better understanding of miRNA's roles in cancer biology and as an alternative tool for anticancer gene therapy. PMID:24859534

  4. Targeted Regression of Hepatocellular Carcinoma by Cancer-Specific RNA Replacement through MicroRNA Regulation.

    PubMed

    Kim, Juhyun; Won, Ranhui; Ban, Guyee; Ju, Mi Ha; Cho, Kyung Sook; Young Han, Sang; Jeong, Jin-Sook; Lee, Seong-Wook

    2015-01-01

    Hepatocellular carcinoma (HCC) has a high fatality rate and limited therapeutic options with side effects and low efficacy. Here, we proposed a new anti-HCC approach based on cancer-specific post-transcriptional targeting. To this end, trans-splicing ribozymes from Tetrahymena group I intron were developed, which can specifically induce therapeutic gene activity through HCC-specific replacement of telomerase reverse transcriptase (TERT) RNA. To circumvent side effects due to TERT expression in regenerating liver tissue, liver-specific microRNA-regulated ribozymes were constructed by incorporating complementary binding sites for the hepatocyte-selective microRNA-122a (miR-122a), which is down-regulated in HCC. The ribozyme activity in vivo was assessed in mouse models orthotopically implanted with HCC. Systemic administration of adenovirus encoding the developed ribozymes caused efficient anti-cancer effect and the least hepatotoxicity with regulation of ribozyme expression by miR-122a in both xenografted and syngeneic orthotopic murine model of multifocal HCC. Of note, the ribozyme induced local and systemic antitumor immunity, thereby completely suppressing secondary tumor challenge in the syngeneic mouse. The cancer specific trans-splicing ribozyme system, which mediates tissue-specific microRNA-regulated RNA replacement, provides a clinically relevant, safe, and efficient strategy for HCC treatment. PMID:26189916

  5. Targeted Regression of Hepatocellular Carcinoma by Cancer-Specific RNA Replacement through MicroRNA Regulation

    PubMed Central

    Kim, Juhyun; Won, Ranhui; Ban, Guyee; Ha Ju, Mi; Sook Cho, Kyung; Young Han, Sang; Jeong, Jin-Sook; Lee, Seong-Wook

    2015-01-01

    Hepatocellular carcinoma (HCC) has a high fatality rate and limited therapeutic options with side effects and low efficacy. Here, we proposed a new anti-HCC approach based on cancer-specific post-transcriptional targeting. To this end, trans-splicing ribozymes from Tetrahymena group I intron were developed, which can specifically induce therapeutic gene activity through HCC-specific replacement of telomerase reverse transcriptase (TERT) RNA. To circumvent side effects due to TERT expression in regenerating liver tissue, liver-specific microRNA-regulated ribozymes were constructed by incorporating complementary binding sites for the hepatocyte-selective microRNA-122a (miR-122a), which is down-regulated in HCC. The ribozyme activity in vivo was assessed in mouse models orthotopically implanted with HCC. Systemic administration of adenovirus encoding the developed ribozymes caused efficient anti-cancer effect and the least hepatotoxicity with regulation of ribozyme expression by miR-122a in both xenografted and syngeneic orthotopic murine model of multifocal HCC. Of note, the ribozyme induced local and systemic antitumor immunity, thereby completely suppressing secondary tumor challenge in the syngeneic mouse. The cancer specific trans-splicing ribozyme system, which mediates tissue-specific microRNA-regulated RNA replacement, provides a clinically relevant, safe, and efficient strategy for HCC treatment. PMID:26189916

  6. Differential MicroRNA Expression Profile in Myxomatous Mitral Valve Prolapse and Fibroelastic Deficiency Valves.

    PubMed

    Chen, Yei-Tsung; Wang, Juan; Wee, Abby S Y; Yong, Quek-Wei; Tay, Edgar Lik-Wui; Woo, Chin Cheng; Sorokin, Vitaly; Richards, Arthur Mark; Ling, Lieng-Hsi

    2016-01-01

    Myxomatous mitral valve prolapse (MMVP) and fibroelastic deficiency (FED) are two common variants of degenerative mitral valve disease (DMVD), which is a leading cause of mitral regurgitation worldwide. While pathohistological studies have revealed differences in extracellular matrix content in MMVP and FED, the molecular mechanisms underlying these two disease entities remain to be elucidated. By using surgically removed valvular specimens from MMVP and FED patients that were categorized on the basis of echocardiographic, clinical and operative findings, a cluster of microRNAs that expressed differentially were identified. The expressions of has-miR-500, -3174, -17, -1193, -646, -1273e, -4298, -203, -505, and -939 showed significant differences between MMVP and FED after applying Bonferroni correction (p < 0.002174). The possible involvement of microRNAs in the pathogenesis of DMVD were further suggested by the presences of in silico predicted target sites on a number of genes reported to be involved in extracellular matrix homeostasis and marker genes for cellular composition of mitral valves, including decorin (DCN), aggrecan (ACAN), fibromodulin (FMOD), α actin 2 (ACTA2), extracellular matrix protein 2 (ECM2), desmin (DES), endothelial cell specific molecule 1 (ESM1), and platelet/ endothelial cell adhesion molecule 1 (PECAM1), as well as inverse correlations of selected microRNA and mRNA expression in MMVP and FED groups. Our results provide evidence that distinct molecular mechanisms underlie MMVP and FED. Moreover, the microRNAs identified may be targets for the future development of diagnostic biomarkers and therapeutics. PMID:27213335

  7. Differential MicroRNA Expression Profile in Myxomatous Mitral Valve Prolapse and Fibroelastic Deficiency Valves

    PubMed Central

    Chen, Yei-Tsung; Wang, Juan; Wee, Abby S. Y.; Yong, Quek-Wei; Tay, Edgar Lik-Wui; Woo, Chin Cheng; Sorokin, Vitaly; Richards, Arthur Mark; Ling, Lieng-Hsi

    2016-01-01

    Myxomatous mitral valve prolapse (MMVP) and fibroelastic deficiency (FED) are two common variants of degenerative mitral valve disease (DMVD), which is a leading cause of mitral regurgitation worldwide. While pathohistological studies have revealed differences in extracellular matrix content in MMVP and FED, the molecular mechanisms underlying these two disease entities remain to be elucidated. By using surgically removed valvular specimens from MMVP and FED patients that were categorized on the basis of echocardiographic, clinical and operative findings, a cluster of microRNAs that expressed differentially were identified. The expressions of has-miR-500, -3174, -17, -1193, -646, -1273e, -4298, -203, -505, and -939 showed significant differences between MMVP and FED after applying Bonferroni correction (p < 0.002174). The possible involvement of microRNAs in the pathogenesis of DMVD were further suggested by the presences of in silico predicted target sites on a number of genes reported to be involved in extracellular matrix homeostasis and marker genes for cellular composition of mitral valves, including decorin (DCN), aggrecan (ACAN), fibromodulin (FMOD), α actin 2 (ACTA2), extracellular matrix protein 2 (ECM2), desmin (DES), endothelial cell specific molecule 1 (ESM1), and platelet/ endothelial cell adhesion molecule 1 (PECAM1), as well as inverse correlations of selected microRNA and mRNA expression in MMVP and FED groups. Our results provide evidence that distinct molecular mechanisms underlie MMVP and FED. Moreover, the microRNAs identified may be targets for the future development of diagnostic biomarkers and therapeutics. PMID:27213335

  8. Radiation-Induced Micro-RNA Expression Changes in Peripheral Blood Cells of Radiotherapy Patients

    SciTech Connect

    Templin, Thomas; Paul, Sunirmal; Amundson, Sally A.; Young, Erik F.; Barker, Christopher A.; Wolden, Suzanne L.; Smilenov, Lubomir B.

    2011-06-01

    Purpose: MicroRNAs (miRNAs), a class of noncoding small RNAs that regulate gene expression, are involved in numerous physiologic processes in normal and malignant cells. Our in vivo study measured miRNA and gene expression changes in human blood cells in response to ionizing radiation, to develop miRNA signatures that can be used as biomarkers for radiation exposure. Methods and Materials: Blood from 8 radiotherapy patients in complete remission 1 or 2 was collected immediately before and 4 hours after total body irradiation with 1.25 Gy x-rays. Both miRNA and gene expression changes were measured by means of quantitative polymerase chain reaction and microarray hybridization, respectively. Hierarchic clustering, multidimensional scaling, class prediction, and gene ontology analysis were performed to investigate the potential of miRNAs to serve as radiation biomarkers and to elucidate their likely physiologic roles in the radiation response. Results: The expression levels of 45 miRNAs were statistically significantly upregulated 4 hours after irradiation with 1.25 Gy x-rays, 27 of them in every patient. Nonirradiated and irradiated samples form separate clusters in hierarchic clustering and multidimensional scaling. Out of 223 differentially expressed genes, 37 were both downregulated and predicted targets of the upregulated miRNAs. Paired and unpaired miRNA-based classifiers that we developed can predict the class membership of a sample with unknown irradiation status, with accuracies of 100% when all 45 upregulated miRNAs are included. Both miRNA control of and gene involvement in biologic processes such as hemopoiesis and the immune response are increased after irradiation, whereas metabolic processes are underrepresented among all differentially expressed genes and the genes controlled by miRNAs. Conclusions: Exposure to ionizing radiation leads to the upregulation of the expression of a considerable proportion of the human miRNAome of peripheral blood cells

  9. Identification of factors involved in target RNA-directed microRNA degradation.

    PubMed

    Haas, Gabrielle; Cetin, Semih; Messmer, Mélanie; Chane-Woon-Ming, Béatrice; Terenzi, Olivier; Chicher, Johana; Kuhn, Lauriane; Hammann, Philippe; Pfeffer, Sébastien

    2016-04-01

    The mechanism by which micro (mi)RNAs control their target gene expression is now well understood. It is however less clear how the level of miRNAs themselves is regulated. Under specific conditions, abundant and highly complementary target RNA can trigger miRNA degradation by a mechanism involving nucleotide addition and exonucleolytic degradation. One such mechanism has been previously observed to occur naturally during viral infection. To date, the molecular details of this phenomenon are not known. We report here that both the degree of complementarity and the ratio of miRNA/target abundance are crucial for the efficient decay of the small RNA. Using a proteomic approach based on the transfection of biotinylated antimiRNA oligonucleotides, we set to identify the factors involved in target-mediated miRNA degradation. Among the retrieved proteins, we identified members of the RNA-induced silencing complex, but also RNA modifying and degradation enzymes. We further validate and characterize the importance of one of these, the Perlman Syndrome 3'-5' exonuclease DIS3L2. We show that this protein interacts with Argonaute 2 and functionally validate its role in target-directed miRNA degradation both by artificial targets and in the context of mouse cytomegalovirus infection. PMID:26809675

  10. Identification of factors involved in target RNA-directed microRNA degradation

    PubMed Central

    Haas, Gabrielle; Cetin, Semih; Messmer, Mélanie; Chane-Woon-Ming, Béatrice; Terenzi, Olivier; Chicher, Johana; Kuhn, Lauriane; Hammann, Philippe; Pfeffer, Sébastien

    2016-01-01

    The mechanism by which micro (mi)RNAs control their target gene expression is now well understood. It is however less clear how the level of miRNAs themselves is regulated. Under specific conditions, abundant and highly complementary target RNA can trigger miRNA degradation by a mechanism involving nucleotide addition and exonucleolytic degradation. One such mechanism has been previously observed to occur naturally during viral infection. To date, the molecular details of this phenomenon are not known. We report here that both the degree of complementarity and the ratio of miRNA/target abundance are crucial for the efficient decay of the small RNA. Using a proteomic approach based on the transfection of biotinylated antimiRNA oligonucleotides, we set to identify the factors involved in target-mediated miRNA degradation. Among the retrieved proteins, we identified members of the RNA-induced silencing complex, but also RNA modifying and degradation enzymes. We further validate and characterize the importance of one of these, the Perlman Syndrome 3′-5′ exonuclease DIS3L2. We show that this protein interacts with Argonaute 2 and functionally validate its role in target-directed miRNA degradation both by artificial targets and in the context of mouse cytomegalovirus infection. PMID:26809675

  11. MicroRNA Expression during Bovine Oocyte Maturation and Fertilization

    PubMed Central

    Gilchrist, Graham C.; Tscherner, Allison; Nalpathamkalam, Thomas; Merico, Daniele; LaMarre, Jonathan

    2016-01-01

    Successful fertilization and subsequent embryo development rely on complex molecular processes starting with the development of oocyte competence through maturation. MicroRNAs (miRNAs) are small non-coding RNA molecules that function as gene regulators in many biological systems, including the oocyte and embryo. In order to further explore the roles of miRNAs in oocyte maturation, we employed small RNA sequencing as a screening tool to identify and characterize miRNA populations present in pools of bovine germinal vesicle (GV) oocytes, metaphase II (MII) oocytes, and presumptive zygotes (PZ). Each stage contained a defined miRNA population, some of which showed stable expression while others showed progressive changes between stages that were subsequently confirmed by quantitative reverse transcription polymerase chain reaction (RT-PCR). Bta-miR-155, bta-miR-222, bta-miR-21, bta-let-7d, bta-let-7i, and bta-miR-190a were among the statistically significant differentially expressed miRNAs (p < 0.05). To determine whether changes in specific primary miRNA (pri-miRNA) transcripts were responsible for the observed miRNA changes, we evaluated pri-miR-155, -222 and let-7d expression. Pri-miR-155 and -222 were not detected in GV oocytes but pri-miR-155 was present in MII oocytes, indicating transcription during maturation. In contrast, levels of pri-let-7d decreased during maturation, suggesting that the observed increase in let-7d expression was likely due to processing of the primary transcript. This study demonstrates that both dynamic and stable populations of miRNAs are present in bovine oocytes and zygotes and extend previous studies supporting the importance of the small RNA landscape in the maturing bovine oocyte and early embryo. PMID:26999121

  12. MicroRNA Expression during Bovine Oocyte Maturation and Fertilization.

    PubMed

    Gilchrist, Graham C; Tscherner, Allison; Nalpathamkalam, Thomas; Merico, Daniele; LaMarre, Jonathan

    2016-01-01

    Successful fertilization and subsequent embryo development rely on complex molecular processes starting with the development of oocyte competence through maturation. MicroRNAs (miRNAs) are small non-coding RNA molecules that function as gene regulators in many biological systems, including the oocyte and embryo. In order to further explore the roles of miRNAs in oocyte maturation, we employed small RNA sequencing as a screening tool to identify and characterize miRNA populations present in pools of bovine germinal vesicle (GV) oocytes, metaphase II (MII) oocytes, and presumptive zygotes (PZ). Each stage contained a defined miRNA population, some of which showed stable expression while others showed progressive changes between stages that were subsequently confirmed by quantitative reverse transcription polymerase chain reaction (RT-PCR). Bta-miR-155, bta-miR-222, bta-miR-21, bta-let-7d, bta-let-7i, and bta-miR-190a were among the statistically significant differentially expressed miRNAs (p < 0.05). To determine whether changes in specific primary miRNA (pri-miRNA) transcripts were responsible for the observed miRNA changes, we evaluated pri-miR-155, -222 and let-7d expression. Pri-miR-155 and -222 were not detected in GV oocytes but pri-miR-155 was present in MII oocytes, indicating transcription during maturation. In contrast, levels of pri-let-7d decreased during maturation, suggesting that the observed increase in let-7d expression was likely due to processing of the primary transcript. This study demonstrates that both dynamic and stable populations of miRNAs are present in bovine oocytes and zygotes and extend previous studies supporting the importance of the small RNA landscape in the maturing bovine oocyte and early embryo. PMID:26999121

  13. MicroRNA polymorphisms and risk of colorectal cancer

    PubMed Central

    Schmit, Stephanie L.; Gollub, Jeremy; Shapero, Michael H.; Huang, Shu-Chen; Rennert, Hedy S.; Finn, Andrea; Rennert, Gad; Gruber, Stephen B.

    2016-01-01

    Background MicroRNAs (miRNAs) act as post-transcriptional regulators of gene expression. Genetic variation in miRNA-encoding sequences or their corresponding binding sites may affect the fidelity of the miRNA-messenger RNA interaction and subsequently alter risk of cancer development. Methods This study expanded the search for miRNA-related polymorphisms contributing to the etiology of colorectal cancer (CRC) across the genome using a novel platform, the Axiom® miRNA Target Site Genotyping Array (237,858 markers). After quality control, the study included 596 cases and 429 controls from the Molecular Epidemiology of Colorectal Cancer study, a population-based case-control study of CRC in northern Israel. The association between each marker and CRC status was examined assuming a log-additive genetic model using logistic regression adjusted for sex, age, and two principal components. Results Twenty-three markers had p-values less than 5.0E-04, and the most statistically significant association involved rs2985 (chr6:34845648; intronic of UHRF1BP1; OR=0.66; p-value=3.7E-05). Further, this study replicated a previously published locus, rs1051690 in the 3’-untranslated region of the insulin receptor gene INSR (OR = 1.38; p = 0.03), with strong evidence of differences in INSR gene expression by genotype. Conclusions This study is the first to examine associations between genetic variation in miRNA target sites and CRC using a genome-wide approach. Functional studies to identify allele-specific effects on miRNA binding are needed to confirm the regulatory capacity of genetic variation to influence risk of CRC. Impact This study demonstrates the potential for a miRNA-targeted genome-wide association study to identify candidate susceptibility loci and prioritize them for functional characterization. PMID:25342389

  14. MicroRNA Related Polymorphisms and Breast Cancer Risk

    PubMed Central

    Khan, Sofia; Greco, Dario; Michailidou, Kyriaki; Milne, Roger L.; Muranen, Taru A.; Heikkinen, Tuomas; Aaltonen, Kirsimari; Dennis, Joe; Bolla, Manjeet K.; Liu, Jianjun; Hall, Per; Irwanto, Astrid; Humphreys, Keith; Li, Jingmei; Czene, Kamila; Chang-Claude, Jenny; Hein, Rebecca; Rudolph, Anja; Seibold, Petra; Flesch-Janys, Dieter; Fletcher, Olivia; Peto, Julian; dos Santos Silva, Isabel; Johnson, Nichola; Gibson, Lorna; Aitken, Zoe; Hopper, John L.; Tsimiklis, Helen; Bui, Minh; Makalic, Enes; Schmidt, Daniel F.; Southey, Melissa C.; Apicella, Carmel; Stone, Jennifer; Waisfisz, Quinten; Meijers-Heijboer, Hanne; Adank, Muriel A.; van der Luijt, Rob B.; Meindl, Alfons; Schmutzler, Rita K.; Müller-Myhsok, Bertram; Lichtner, Peter; Turnbull, Clare; Rahman, Nazneen; Chanock, Stephen J.; Hunter, David J.; Cox, Angela; Cross, Simon S.; Reed, Malcolm W. R.; Schmidt, Marjanka K.; Broeks, Annegien; Veer, Laura J. V. a. n't.; Hogervorst, Frans B.; Fasching, Peter A.; Schrauder, Michael G.; Ekici, Arif B.; Beckmann, Matthias W.; Bojesen, Stig E.; Nordestgaard, Børge G.; Nielsen, Sune F.; Flyger, Henrik; Benitez, Javier; Zamora, Pilar M.; Perez, Jose I. A.; Haiman, Christopher A.; Henderson, Brian E.; Schumacher, Fredrick; Le Marchand, Loic; Pharoah, Paul D. P.; Dunning, Alison M.; Shah, Mitul; Luben, Robert; Brown, Judith; Couch, Fergus J.; Wang, Xianshu; Vachon, Celine; Olson, Janet E.; Lambrechts, Diether; Moisse, Matthieu; Paridaens, Robert; Christiaens, Marie-Rose; Guénel, Pascal; Truong, Thérèse; Laurent-Puig, Pierre; Mulot, Claire; Marme, Frederick; Burwinkel, Barbara; Schneeweiss, Andreas; Sohn, Christof; Sawyer, Elinor J.; Tomlinson, Ian; Kerin, Michael J.; Miller, Nicola; Andrulis, Irene L.; Knight, Julia A.; Tchatchou, Sandrine; Mulligan, Anna Marie; Dörk, Thilo; Bogdanova, Natalia V.; Antonenkova, Natalia N.; Anton-Culver, Hoda; Darabi, Hatef; Eriksson, Mikael; Garcia-Closas, Montserrat; Figueroa, Jonine; Lissowska, Jolanta; Brinton, Louise; Devilee, Peter; Tollenaar, Robert A. E. M.; Seynaeve, Caroline; van Asperen, Christi J.; Kristensen, Vessela N.; Slager, Susan; Toland, Amanda E.; Ambrosone, Christine B.; Yannoukakos, Drakoulis; Lindblom, Annika; Margolin, Sara; Radice, Paolo; Peterlongo, Paolo; Barile, Monica; Mariani, Paolo; Hooning, Maartje J.; Martens, John W. M.; Collée, J. Margriet; Jager, Agnes; Jakubowska, Anna; Lubinski, Jan; Jaworska-Bieniek, Katarzyna; Durda, Katarzyna; Giles, Graham G.; McLean, Catriona; Brauch, Hiltrud; Brüning, Thomas; Ko, Yon-Dschun; Brenner, Hermann; Dieffenbach, Aida Karina; Arndt, Volker; Stegmaier, Christa; Swerdlow, Anthony; Ashworth, Alan; Orr, Nick; Jones, Michael; Simard, Jacques; Goldberg, Mark S.; Labrèche, France; Dumont, Martine; Winqvist, Robert; Pylkäs, Katri; Jukkola-Vuorinen, Arja; Grip, Mervi; Kataja, Vesa; Kosma, Veli-Matti; Hartikainen, Jaana M.; Mannermaa, Arto; Hamann, Ute; Chenevix-Trench, Georgia; Blomqvist, Carl; Aittomäki, Kristiina; Easton, Douglas F.; Nevanlinna, Heli

    2014-01-01

    Genetic variations, such as single nucleotide polymorphisms (SNPs) in microRNAs (miRNA) or in the miRNA binding sites may affect the miRNA dependent gene expression regulation, which has been implicated in various cancers, including breast cancer, and may alter individual susceptibility to cancer. We investigated associations between miRNA related SNPs and breast cancer risk. First we evaluated 2,196 SNPs in a case-control study combining nine genome wide association studies (GWAS). Second, we further investigated 42 SNPs with suggestive evidence for association using 41,785 cases and 41,880 controls from 41 studies included in the Breast Cancer Association Consortium (BCAC). Combining the GWAS and BCAC data within a meta-analysis, we estimated main effects on breast cancer risk as well as risks for estrogen receptor (ER) and age defined subgroups. Five miRNA binding site SNPs associated significantly with breast cancer risk: rs1045494 (odds ratio (OR) 0.92; 95% confidence interval (CI): 0.88–0.96), rs1052532 (OR 0.97; 95% CI: 0.95–0.99), rs10719 (OR 0.97; 95% CI: 0.94–0.99), rs4687554 (OR 0.97; 95% CI: 0.95–0.99, and rs3134615 (OR 1.03; 95% CI: 1.01–1.05) located in the 3′ UTR of CASP8, HDDC3, DROSHA, MUSTN1, and MYCL1, respectively. DROSHA belongs to miRNA machinery genes and has a central role in initial miRNA processing. The remaining genes are involved in different molecular functions, including apoptosis and gene expression regulation. Further studies are warranted to elucidate whether the miRNA binding site SNPs are the causative variants for the observed risk effects. PMID:25390939

  15. MicroRNA related polymorphisms and breast cancer risk.

    PubMed

    Khan, Sofia; Greco, Dario; Michailidou, Kyriaki; Milne, Roger L; Muranen, Taru A; Heikkinen, Tuomas; Aaltonen, Kirsimari; Dennis, Joe; Bolla, Manjeet K; Liu, Jianjun; Hall, Per; Irwanto, Astrid; Humphreys, Keith; Li, Jingmei; Czene, Kamila; Chang-Claude, Jenny; Hein, Rebecca; Rudolph, Anja; Seibold, Petra; Flesch-Janys, Dieter; Fletcher, Olivia; Peto, Julian; dos Santos Silva, Isabel; Johnson, Nichola; Gibson, Lorna; Aitken, Zoe; Hopper, John L; Tsimiklis, Helen; Bui, Minh; Makalic, Enes; Schmidt, Daniel F; Southey, Melissa C; Apicella, Carmel; Stone, Jennifer; Waisfisz, Quinten; Meijers-Heijboer, Hanne; Adank, Muriel A; van der Luijt, Rob B; Meindl, Alfons; Schmutzler, Rita K; Müller-Myhsok, Bertram; Lichtner, Peter; Turnbull, Clare; Rahman, Nazneen; Chanock, Stephen J; Hunter, David J; Cox, Angela; Cross, Simon S; Reed, Malcolm W R; Schmidt, Marjanka K; Broeks, Annegien; Van't Veer, Laura J; Hogervorst, Frans B; Fasching, Peter A; Schrauder, Michael G; Ekici, Arif B; Beckmann, Matthias W; Bojesen, Stig E; Nordestgaard, Børge G; Nielsen, Sune F; Flyger, Henrik; Benitez, Javier; Zamora, Pilar M; Perez, Jose I A; Haiman, Christopher A; Henderson, Brian E; Schumacher, Fredrick; Le Marchand, Loic; Pharoah, Paul D P; Dunning, Alison M; Shah, Mitul; Luben, Robert; Brown, Judith; Couch, Fergus J; Wang, Xianshu; Vachon, Celine; Olson, Janet E; Lambrechts, Diether; Moisse, Matthieu; Paridaens, Robert; Christiaens, Marie-Rose; Guénel, Pascal; Truong, Thérèse; Laurent-Puig, Pierre; Mulot, Claire; Marme, Frederick; Burwinkel, Barbara; Schneeweiss, Andreas; Sohn, Christof; Sawyer, Elinor J; Tomlinson, Ian; Kerin, Michael J; Miller, Nicola; Andrulis, Irene L; Knight, Julia A; Tchatchou, Sandrine; Mulligan, Anna Marie; Dörk, Thilo; Bogdanova, Natalia V; Antonenkova, Natalia N; Anton-Culver, Hoda; Darabi, Hatef; Eriksson, Mikael; Garcia-Closas, Montserrat; Figueroa, Jonine; Lissowska, Jolanta; Brinton, Louise; Devilee, Peter; Tollenaar, Robert A E M; Seynaeve, Caroline; van Asperen, Christi J; Kristensen, Vessela N; Slager, Susan; Toland, Amanda E; Ambrosone, Christine B; Yannoukakos, Drakoulis; Lindblom, Annika; Margolin, Sara; Radice, Paolo; Peterlongo, Paolo; Barile, Monica; Mariani, Paolo; Hooning, Maartje J; Martens, John W M; Collée, J Margriet; Jager, Agnes; Jakubowska, Anna; Lubinski, Jan; Jaworska-Bieniek, Katarzyna; Durda, Katarzyna; Giles, Graham G; McLean, Catriona; Brauch, Hiltrud; Brüning, Thomas; Ko, Yon-Dschun; Brenner, Hermann; Dieffenbach, Aida Karina; Arndt, Volker; Stegmaier, Christa; Swerdlow, Anthony; Ashworth, Alan; Orr, Nick; Jones, Michael; Simard, Jacques; Goldberg, Mark S; Labrèche, France; Dumont, Martine; Winqvist, Robert; Pylkäs, Katri; Jukkola-Vuorinen, Arja; Grip, Mervi; Kataja, Vesa; Kosma, Veli-Matti; Hartikainen, Jaana M; Mannermaa, Arto; Hamann, Ute; Chenevix-Trench, Georgia; Blomqvist, Carl; Aittomäki, Kristiina; Easton, Douglas F; Nevanlinna, Heli

    2014-01-01

    Genetic variations, such as single nucleotide polymorphisms (SNPs) in microRNAs (miRNA) or in the miRNA binding sites may affect the miRNA dependent gene expression regulation, which has been implicated in various cancers, including breast cancer, and may alter individual susceptibility to cancer. We investigated associations between miRNA related SNPs and breast cancer risk. First we evaluated 2,196 SNPs in a case-control study combining nine genome wide association studies (GWAS). Second, we further investigated 42 SNPs with suggestive evidence for association using 41,785 cases and 41,880 controls from 41 studies included in the Breast Cancer Association Consortium (BCAC). Combining the GWAS and BCAC data within a meta-analysis, we estimated main effects on breast cancer risk as well as risks for estrogen receptor (ER) and age defined subgroups. Five miRNA binding site SNPs associated significantly with breast cancer risk: rs1045494 (odds ratio (OR) 0.92; 95% confidence interval (CI): 0.88-0.96), rs1052532 (OR 0.97; 95% CI: 0.95-0.99), rs10719 (OR 0.97; 95% CI: 0.94-0.99), rs4687554 (OR 0.97; 95% CI: 0.95-0.99, and rs3134615 (OR 1.03; 95% CI: 1.01-1.05) located in the 3' UTR of CASP8, HDDC3, DROSHA, MUSTN1, and MYCL1, respectively. DROSHA belongs to miRNA machinery genes and has a central role in initial miRNA processing. The remaining genes are involved in different molecular functions, including apoptosis and gene expression regulation. Further studies are warranted to elucidate whether the miRNA binding site SNPs are the causative variants for the observed risk effects. PMID:25390939

  16. MicroRNA-binding viral protein interferes with Arabidopsis development.

    PubMed

    Chellappan, Padmanabhan; Vanitharani, Ramachandran; Fauquet, Claude M

    2005-07-19

    MicroRNAs (miRNAs) are small (approximately 21 nt), noncoding RNAs that negatively regulate target mRNAs at the posttranscriptional level that are involved in development. In plants, virus-induced disease symptoms often result in developmental abnormalities resembling perturbation of miRNA-mediated function. Here, we report that expression in transgenic plants of a geminivirus-encoded AC4 protein from African cassava mosaic virus Cameroon Strain (ACMV), a suppressor of posttranscriptional gene silencing, was correlated with decreased accumulation of host miRNAs and increased development abnormalities in Arabidopsis. Down-regulation of miRNA correlated with an up-regulation of target mRNA level. In vitro binding assays revealed the ability of AC4 of ACMV (A-AC4) but not East African cassava mosaic Cameroon virus AC2 to bind single-stranded forms of miRNAs and short interfering RNAs but not double-stranded RNA forms. Normally, a labile intermediate during the miRNA biogenesis/RNA-induced silencing complex assembly, miRNA*, was below the level of detection, indicating that AC4 might interfere at a point downstream of the miRNA duplex unwinding process. The association of AC4 with miRNA was demonstrated by the association of A-AC4-GFP fusion protein, extracted from Arabidopsis protoplasts, with 2'-O-methyloligonucleotide complementary to miR159 (miR159*) and by the presence of miRNA with the A-AC4-GFP fusion protein after immunoprecipitation with antibody against GFP. In both assays, A-AC4 protein and miRNA complexes were copurified. These results provide direct evidence that AC4 is a unique virus-encoded posttranscriptional gene-silencing suppressor protein that binds to and presumably inactivates mature miRNAs and thus blocks the normal miRNA-mediated regulation of target mRNAs, resulting in developmental defects in Arabidopsis. PMID:16006510

  17. Multifaceted enrichment analysis of RNA–RNA crosstalk reveals cooperating micro-societies in human colorectal cancer

    PubMed Central

    Mazza, Tommaso; Mazzoccoli, Gianluigi; Fusilli, Caterina; Capocefalo, Daniele; Panza, Anna; Biagini, Tommaso; Castellana, Stefano; Gentile, Annamaria; De Cata, Angelo; Palumbo, Orazio; Stallone, Raffaella; Rubino, Rosa; Carella, Massimo; Piepoli, Ada

    2016-01-01

    Alterations in the balance of mRNA and microRNA (miRNA) expression profiles contribute to the onset and development of colorectal cancer. The regulatory functions of individual miRNA-gene pairs are widely acknowledged, but group effects are largely unexplored. We performed an integrative analysis of mRNA–miRNA and miRNA–miRNA interactions using high-throughput mRNA and miRNA expression profiles obtained from matched specimens of human colorectal cancer tissue and adjacent non-tumorous mucosa. This investigation resulted in a hypernetwork-based model, whose functional backbone was fulfilled by tight micro-societies of miRNAs. These proved to modulate several genes that are known to control a set of significantly enriched cancer-enhancer and cancer-protection biological processes, and that an array of upstream regulatory analyses demonstrated to be dependent on miR-145, a cell cycle and MAPK signaling cascade master regulator. In conclusion, we reveal miRNA-gene clusters and gene families with close functional relationships and highlight the role of miR-145 as potent upstream regulator of a complex RNA–RNA crosstalk, which mechanistically modulates several signaling pathways and regulatory circuits that when deranged are relevant to the changes occurring in colorectal carcinogenesis. PMID:27067546

  18. Systematic study of Drosophila microRNA functions using a collection of targeted knockout mutations.

    PubMed

    Chen, Ya-Wen; Song, Shilin; Weng, Ruifen; Verma, Pushpa; Kugler, Jan-Michael; Buescher, Marita; Rouam, Sigrid; Cohen, Stephen M

    2014-12-22

    MicroRNAs are abundant in animal genomes, yet little is known about their functions in vivo. Here, we report the production of 80 new Drosophila miRNA mutants by targeted homologous recombination. These mutants remove 104 miRNAs. Together with 15 previously reported mutants, this collection includes 95 mutants deleting 130 miRNAs. Collectively, these genes produce over 99% of all Drosophila miRNAs, measured by miRNA sequence reads. We present a survey of developmental and adult miRNA phenotypes. Over 80% of the mutants showed at least one phenotype using a p < 0.01 significance threshold. We observed a significant correlation between miRNA abundance and phenotypes related to survival and lifespan, but not to most other phenotypes. miRNA cluster mutants were no more likely than single miRNA mutants to produce significant phenotypes. This mutant collection will provide a resource for future analysis of the biological roles of Drosophila miRNAs. PMID:25535920

  19. Retrieving relevant experiments: The case of microRNA microarrays.

    PubMed

    Açıcı, Koray; Terzi, Yunus Kasım; Oğul, Hasan

    2015-08-01

    Content-based retrieval of biological experiments in large public repositories is a recent challenge in computational biology and bioinformatics. The task is, in general, to search in a database using a query-by-example without any experimental meta-data annotation. Here, we consider a more specific problem that seeks a solution for retrieving relevant microRNA experiments from microarray repositories. A computational framework is proposed with this objective. The framework adapts a normal-uniform mixture model for identifying differentially expressed microRNAs in microarray profiling experiments. A rank-based thresholding scheme is offered to binarize real-valued experiment fingerprints based on differential expression. An effective similarity metric is introduced to compare categorical fingerprints, which in turn infers the relevance between two experiments. Two different views of experimental relevance are evaluated, one for disease association and another for embryonic germ layer, to discern the retrieval ability of the proposed model. To the best of our knowledge, the experiment retrieval task is investigated for the first time in the context of microRNA microarrays. PMID:26116091

  20. MicroRNA-206: a Promising Theranostic Marker

    PubMed Central

    Novák, Jan; Kružliak, Peter; Bienertová-Vašků, Julie; Slabý, Ondřej; Novák, Miroslav

    2014-01-01

    MicroRNAs (miRs) are small non-coding RNAs that negatively regulate gene expression by binding to the 3` untranslated regions (3`UTR) of their target mRNAs. MiRs were shown to play pivotal roles in tissue development and function and are also involved in the pathogenesis of various diseases including cancer. MicroRNA-206, which belongs to the group of so-called “myomiRs”, is one of the most studied miRs thus far. In addition to being involved in skeletal muscle development and pathology, it has also been established that it is involved in the pathogenesis of numerous diseases including heart failure, chronic obstructive pulmonary disease, Alzheimer's disease and various types of cancers. The aim of this review is to provide a complex overview of microRNA-206, including regulating its expression, a brief description of its known functions in skeletal muscle and a complex overview of its roles in the biology and pathology of other tissues, emphasizing its significant diagnostic and therapeutic potential. PMID:24465270

  1. MicroRNA-183/182/96 cooperatively regulates the proliferation of colon cancer cells.

    PubMed

    Zhang, Qingquan; Ren, Wei; Huang, Bin; Yi, Liang; Zhu, Hongtao

    2015-07-01

    The microRNA (miR/miRNA)-182/183/96 cluster comprises miR-96, -182 and -183. The present study examined five previous microarray-based human colon cancer miR expression profiling studies and the expression of these three miRs was found to be upregulated in colon cancer tissues. Subsequently, in vitro assays were performed to determine the role of the miR-183/182/96 cluster in colon cancer cells. The results demonstrated that inhibiting miR-183, miR-182 or miR-96 with antisense oligonucleotide (ASO)-mimics inhibited the proliferation of colon cancer cells. Notably, further investigation revealed that inhibiting their expression simultaneously led to a more efficient reduction in cancer cell proliferation. These results suggested that miR-182/183/96, which resides in clusters in the genome, functioned synergistically in colon cancer and implied that co-expression of the miR cluster ASOs was efficient in reducing tumorigenesis, offering novel insight into the use of miRNAs in tumor therapy. PMID:25695717

  2. Quadratic isothermal amplification for the detection of microRNA.

    PubMed

    Duan, Ruixue; Zuo, Xiaolei; Wang, Shutao; Quan, Xiyun; Chen, Dongliang; Chen, Zhifei; Jiang, Lei; Fan, Chunhai; Xia, Fan

    2014-03-01

    This protocol describes an isothermal amplification approach for ultrasensitive detection of specific microRNAs (miRNAs). It achieves this level of sensitivity through quadratic amplification of the target oligonucleotide by using a Bst DNA polymerase-induced strand-displacement reaction and a lambda exonuclease-aided recycling reaction. First, the target miRNA binds to a specifically designed molecular beacon, causing it to become a fluorescence emitter. A primer then binds to the activated beacon, and Bst polymerase initiates the synthesis of a double-stranded DNA segment templated on the molecular beacon. This causes the concomitant release of the target miRNA from the beacon--the first round of 'recycling'. Second, the duplex beacon thus produced is a suitable substrate for a nicking enzyme present in solution. After the duplex beacon is nicked, the lambda exonuclease digests the beacon and releases the DNA single strand just synthesized, which is complementary to the molecular beacon, inducing the second round of recycling. The miRNA detection limit of this protocol is 10 fmol at 37 °C and 1 amol at 4 °C. This approach also affords high selectivity when applied to miRNA extracted from MCF-7 and PC3 cell lines and even from breast cancer tissue samples. Upon isolation of miRNA, the detection process can be completed in ∼2 h. PMID:24525753

  3. A new plasmid-based microRNA inhibitor system that inhibits microRNA families in transgenic mice and cells: a potential new therapeutic reagent.

    PubMed

    Cao, H; Yu, W; Li, X; Wang, J; Gao, S; Holton, N E; Eliason, S; Sharp, T; Amendt, B A

    2016-06-01

    Current tools for the inhibition of microRNA (miR) function are limited to modified antisense oligonucleotides, sponges and decoy RNA molecules and none have been used to understand miR function during development. CRISPR/Cas-mediated deletion of miR sequences within the genome requires multiple chromosomal deletions to remove all functional miR family members because of duplications. Here, we report a novel plasmid-based miR inhibitor system (PMIS) that expresses a new RNA molecule, which inhibits miR family members in cells and mice. The PMIS engineered RNA optimal secondary structure, flanking sequences and specific antisense miR oligonucleotide sequence bind the miR in a stable complex to inhibit miR activity. In cells, one PMIS can effectively inhibit miR family members that share the same seed sequence. The PMIS shows no off-target effects or toxicity and is highly specific for miRs sharing identical seed sequences. Transgenic mice expressing both PMIS-miR-17-18 and PMIS-miR-19-92 show similar phenotypes of miR-17-92-knockout mice. Interestingly, mice only expressing PMIS-miR-17-18 have developmental defects distinct from mice only expressing PMIS-miR-19-92 demonstrating usefulness of the PMIS system to dissect different functions of miRs within clusters. Different PMIS miR inhibitors can be linked together to knock down multiple miRs expressed from different chromosomes. Inhibition of the miR-17-92, miR-106a-363 and miR-106b-25 clusters reveals new mechanisms and developmental defects for these miRs. We report a new tool to dissect the role of miRs in development without genome editing, inhibit miR function in cells and as a potential new therapeutic reagent. PMID:26934100

  4. A new plasmid-based microRNA inhibitor system that inhibits microRNA families in transgenic mice and cells: a potential new therapeutic reagent

    PubMed Central

    Cao, H; Yu, W; Li, X; Wang, J; Gao, S; Holton, N E; Eliason, S; Sharp, T; Amendt, B A

    2016-01-01

    Current tools for the inhibition of microRNA (miR) function are limited to modified antisense oligonucleotides, sponges and decoy RNA molecules and none have been used to understand miR function during development. CRISPR/Cas-mediated deletion of miR sequences within the genome requires multiple chromosomal deletions to remove all functional miR family members because of duplications. Here, we report a novel plasmid-based miR inhibitor system (PMIS) that expresses a new RNA molecule, which inhibits miR family members in cells and mice. The PMIS engineered RNA optimal secondary structure, flanking sequences and specific antisense miR oligonucleotide sequence bind the miR in a stable complex to inhibit miR activity. In cells, one PMIS can effectively inhibit miR family members that share the same seed sequence. The PMIS shows no off-target effects or toxicity and is highly specific for miRs sharing identical seed sequences. Transgenic mice expressing both PMIS-miR-17-18 and PMIS-miR-19-92 show similar phenotypes of miR-17-92-knockout mice. Interestingly, mice only expressing PMIS-miR-17-18 have developmental defects distinct from mice only expressing PMIS-miR-19-92 demonstrating usefulness of the PMIS system to dissect different functions of miRs within clusters. Different PMIS miR inhibitors can be linked together to knock down multiple miRs expressed from different chromosomes. Inhibition of the miR-17-92, miR-106a-363 and miR-106b-25 clusters reveals new mechanisms and developmental defects for these miRs. We report a new tool to dissect the role of miRs in development without genome editing, inhibit miR function in cells and as a potential new therapeutic reagent. PMID:26934100

  5. A new unique form of microRNA from human heart, microRNA-499c, promotes myofibril formation and rescues cardiac development in mutant axolotl embryos

    PubMed Central

    2013-01-01

    Background A recessive mutation “c” in the Mexican axolotl, Ambystoma mexicanum, results in the failure of normal heart development. In homozygous recessive embryos, the hearts do not have organized myofibrils and fail to beat. In our previous studies, we identified a noncoding Myofibril-Inducing RNA (MIR) from axolotls which promotes myofibril formation and rescues heart development. Results We randomly cloned RNAs from fetal human heart. RNA from clone #291 promoted myofibril formation and induced heart development of mutant axolotls in organ culture. This RNA induced expression of cardiac markers in mutant hearts: tropomyosin, troponin and α-syntrophin. This cloned RNA matches in partial sequence alignment to human microRNA-499a and b, although it differs in length. We have concluded that this cloned RNA is unique in its length, but is still related to the microRNA-499 family. We have named this unique RNA, microRNA-499c. Thus, we will refer to this RNA derived from clone #291 as microRNA-499c throughout the rest of the paper. Conclusions This new form, microRNA-499c, plays an important role in cardiac development. PMID:23522091

  6. MicroRNA transcriptome in the newborn mouse ovaries determined by massive parallel sequencing.

    PubMed

    Ahn, Hyo Won; Morin, Ryan D; Zhao, Han; Harris, Ronald A; Coarfa, Cristian; Chen, Zi-Jiang; Milosavljevic, Aleksandar; Marra, Marco A; Rajkovic, Aleksandar

    2010-07-01

    Small non-coding RNAs, such as microRNAs (miRNAs), are involved in diverse biological processes including organ development and tissue differentiation. Global disruption of miRNA biogenesis in Dicer knockout mice disrupts early embryogenesis and primordial germ cell formation. However, the role of miRNAs in early folliculogenesis is poorly understood. In order to identify a full transcriptome set of small RNAs expressed in the newborn (NB) ovary, we extracted small RNA fraction from mouse NB ovary tissues and subjected it to massive parallel sequencing using the Genome Analyzer from Illumina. Massive sequencing produced 4 655 992 reads of 33 bp each representing a total of 154 Mbp of sequence data. The Pash alignment algorithm mapped 50.13% of the reads to the mouse genome. Sequence reads were clustered based on overlapping mapping coordinates and intersected with known miRNAs, small nucleolar RNAs (snoRNAs), piwi-interacting RNA (piRNA) clusters and repetitive genomic regions; 25.2% of the reads mapped to known miRNAs, 25.5% to genomic repeats, 3.5% to piRNAs and 0.18% to snoRNAs. Three hundred and ninety-eight known miRNA species were among the sequenced small RNAs, and 118 isomiR sequences that are not in the miRBase database. Let-7 family was the most abundantly expressed miRNA, and mmu-mir-672, mmu-mir-322, mmu-mir-503 and mmu-mir-465 families are the most abundant X-linked miRNA detected. X-linked mmu-mir-503, mmu-mir-672 and mmu-mir-465 family showed preferential expression in testes and ovaries. We also identified four novel miRNAs that are preferentially expressed in gonads. Gonadal selective miRNAs may play important roles in ovarian development, folliculogenesis and female fertility. PMID:20215419

  7. MicroRNA: a small molecule with a big biological impact.

    PubMed

    Zhou, Xiaofeng; Yang, Pan-Chyr

    2012-01-01

    One of the most significant achievements in biological science in the last decade is the discovery of RNA interference (RNAi), a process within living cells that regulates gene expression at post-transcriptional levels. Historically, this process was described by other more generic names, such as co-suppression and post transcriptional gene silencing. Only after the molecular mechanism underlying these apparently unrelated processes was fully understood did it become apparent that they all described the RNAi phenomenon. In 2006, Dr. Andrew Fire and Dr. Craig C. Mello were awarded the Nobel Prize in Physiology or Medicine for their work on RNAi interference. RNAi is an RNA-dependent gene silencing process that is controlled by the RNA-induced silencing complex (RISC) and is initiated by two types of small RNA molecules - microRNA (miRNA) and small interfering RNA (siRNA). However, the function of microRNA appears to be far beyond RNAi alone, including direct interaction with the gene promoter and epigenetic regulation of the DNA methylation and histone modification. By regulating gene expression, miRNAs are likely to be involved in diverse biological activities, such as tumorigenesis, immune response, insulin secretion, neurotransmitter synthesis, and circadian rhythm, to name a few. MicroRNAs are 21-23 nucleotide single stranded RNA molecules found in eukaryotic cells. The first miRNA, lin-4, was characterized in C. elegans in the early 1990s [1]. In the early years, the progress on microRNA research was slow and experienced substantial growing pains. The short length and uniqueness of each microRNA rendered many conventional hybridization based methods ineffective; very small RNAs are difficult to reliably amplify or label without introducing bias. In addition, hybridization-based methods for microRNA profiling relied on probes designed to detect known microRNAs or known microRNA species previously identified by sequencing or homology search. Recent evidence of

  8. Evolution of a research field—a micro (RNA) example

    PubMed Central

    Casey, Máire-Caitlín; Kerin, Michael J.

    2015-01-01

    Background. Every new scientific field can be traced back to a single, seminal publication. Therefore, a bibliometric analysis can yield significant insights into the history and potential future of a research field. This year marks 21 years since that first ground-breaking microRNA (miRNA) publication. Here, we make the case that the miRNA field is mature, utilising bibliometrics. Methods. Utilising the Web of Science™ (WoS) database publication and citation information, we charted the history of miRNA-related publications, describing and dissecting contributions by publication type (plus category, pay-per-view or open access), journal (highlighting dominant journals), by country, citations and languages. Results. We found that the United States of America (USA) publishes the most miRNA papers, followed by China and Germany. Significantly, publications attributed to the USA also receive the most citations per publication, followed by a close grouping of England, Germany and France. We also describe the relevance and acceptance of the miRNA field to different research areas, through its uptake in areas from oncology to plant sciences. Exploring the recent momentous change in publishing, we find that although pay-per view articles vastly out-number open-access articles, the citation rate of pay-per-view articles is currently less than double that of open-access. Conclusions. We believe the trends described here represent the typical evolution of a research field. By analysing publications, citations and distribution patterns, key moments in the evolution of this research area are recognised, indicating the maturation of the miRNA field and providing guidance for future research endeavours. PMID:25802804

  9. microRNA Profiles in Parkinson's Disease Prefrontal Cortex

    PubMed Central

    Hoss, Andrew G.; Labadorf, Adam; Beach, Thomas G.; Latourelle, Jeanne C.; Myers, Richard H.

    2016-01-01

    Objective: The goal of this study was to compare the microRNA (miRNA) profile of Parkinson's disease (PD) frontal cortex with normal control brain, allowing for the identification of PD specific signatures as well as study the disease-related phenotypes of onset age and dementia. Methods: Small RNA sequence analysis was performed from prefrontal cortex for 29 PD samples and 33 control samples. After sample QC, normalization and batch correction, linear regression was employed to identify miRNAs altered in PD, and a PD classifier was developed using weighted voting class prediction. The relationship of miRNA levels to onset age and PD with dementia (PDD) was also characterized in case-only analyses. Results: One twenty five miRNAs were differentially expressed in PD at a genome-wide level of significance (FDR q < 0.05). A set of 29 miRNAs classified PD from non-diseased brain (93.9% specificity, 96.6% sensitivity). The majority of differentially expressed miRNAs (105/125) showed an ordinal relationship from control, to PD without dementia (PDN), to PDD. Among PD brains, 36 miRNAs classified PDD from PDN (sensitivity = 81.2%, specificity = 88.9%). Among differentially expressed miRNAs, miR-10b-5p had a positive association with onset age (q = 4.7e-2). Conclusions: Based on cortical miRNA levels, PD brains were accurately classified from non-diseased brains. Additionally, the PDD miRNA profile exhibited a more severe pattern of alteration among those differentially expressed in PD. To evaluate the clinical utility of miRNAs as potential clinical biomarkers, further characterization and testing of brain-related miRNA alterations in peripheral biofluids is warranted. PMID:26973511

  10. City block distance and rough-fuzzy clustering for identification of co-expressed microRNAs.

    PubMed

    Paul, Sushmita; Maji, Pradipta

    2014-06-01

    The microRNAs or miRNAs are short, endogenous RNAs having ability to regulate mRNA expression at the post-transcriptional level. Various studies have revealed that miRNAs tend to cluster on chromosomes. The members of a cluster that are in close proximity on chromosomes are highly likely to be processed as co-transcribed units. Therefore, a large proportion of miRNAs are co-expressed. Expression profiling of miRNAs generates a huge volume of data. Complicated networks of miRNA-mRNA interaction increase the challenges of comprehending and interpreting the resulting mass of data. In this regard, this paper presents a clustering algorithm in order to extract meaningful information from miRNA expression data. It judiciously integrates the merits of rough sets, fuzzy sets, the c-means algorithm, and the normalized range-normalized city block distance to discover co-expressed miRNA clusters. While the membership functions of fuzzy sets enable efficient handling of overlapping partitions in a noisy environment, the concept of lower and upper approximations of rough sets deals with uncertainty, vagueness, and incompleteness in cluster definition. The city block distance is used to compute the membership functions of fuzzy sets and to find initial partition of a data set, and therefore helps to handle minute differences between two miRNA expression profiles. The effectiveness of the proposed approach, along with a comparison with other related methods, is demonstrated for several miRNA expression data sets using different cluster validity indices. Moreover, the gene ontology is used to analyze the functional consistency and biological significance of generated miRNA clusters. PMID:24682049

  11. A small molecule enhances RNA interference and promotes microRNA processing

    PubMed Central

    Shan, Ge; Li, Yujing; Zhang, Junliang; Li, Wendi; Szulwach, Keith E; Duan, Ranhui; Faghihi, Mohammad A; Khalil, Ahmad M; Lu, Lianghua; Paroo, Zain; Chan, Anthony W S; Shi, Zhangjie; Liu, Qinghua; Wahlestedt, Claes; He, Chuan; Jin, Peng

    2010-01-01

    Small interfering RNAs (siRNAs) and microRNAs (miRNAs) are sequence-specific post-transcriptional regulators of gene expression. Although major components of the RNA interference (RNAi) pathway have been identified, regulatory mechanisms for this pathway remain largely unknown. Here we demonstrate that the RNAi pathway can be modulated intracellularly by small molecules. We have developed a cell-based assay to monitor the activity of the RNAi pathway and find that the small-molecule enoxacin (Penetrex) enhances siRNA-mediated mRNA degradation and promotes the biogenesis of endogenous miRNAs. We show that this RNAi-enhancing activity depends on the trans-activation-responsive region RNA-binding protein. Our results provide a proof-of-principle demonstration that small molecules can be used to modulate the activity of the RNAi pathway. RNAi enhancers may be useful in the development of research tools and therapeutics. PMID:18641635

  12. MicroRNA response to environmental mutagens in liver.

    PubMed

    Elamin, Bahaeldin K; Callegari, Elisa; Gramantieri, Laura; Sabbioni, Silvia; Negrini, Massimo

    2011-12-01

    During the recent few years, microRNAs emerged as key molecules in the regulation of mammalian cell functions. It was also shown that their altered expression can promote pathologic conditions, such as cancer and other common diseases. Because environmental exposure to biological, chemical or physical agents may be responsible for human diseases, including cancer, uncovering relationships between exposure to environmental carcinogens and expression of microRNAs may help to disclose early mechanisms of disease and it may potentially lead to the development of useful indicators of toxic exposure or novel biomarkers for carcinogenicity testing. The unique expression profile of microRNAs in different types and at different stages of cancer coupled to their remarkable stability in tissues and in serum/plasma suggests that these little molecules may find application as sensitive biomarkers. This review will concentrate on the alterations in microRNA expression in response to environmental factors in relation to the risk of developing liver cancer. PMID:21514310

  13. Missing link between microRNA and prostate cancer.

    PubMed

    Gill, Balraj Singh; Alex, Jimi Marin; Navgeet; Kumar, Sanjeev

    2016-05-01

    MicroRNAs are the non-coding RNAs which regulate endogenous gene expression in animal and plant cells. Alterations in the level of micro-ribonucleic acids (miRNAs) involving the deletions, overexpression, mutations, epigenetic silencing, or dysregulation of transcription factors that target specific miRNAs may culminate in various diseases including cancer. Recent findings demonstrate the role of miRNAs in prostate cancer. Numerous discoveries of miRNAs have marked the research and development surrounding prostate cancer management, diagnosis, and therapy which has made prediction easy, but the effective treatment strategy remains a mystery. This review seeks to draw a link between miRNA and prostate cancer through an understanding of the numerous signaling pathways that these miRNAs control, which may prove to be helpful in identifying therapeutically interesting molecular targets. PMID:26822307

  14. A microRNA Signature Associated with Early Recurrence in Breast Cancer

    PubMed Central

    Carmona, Rosario; de Luque, Vanessa; Vicioso, Luis; Claros, M. Gonzalo; Viguera, Enrique; Pajares, Bella; Sánchez, Alfonso; Ribelles, Nuria; Alba, Emilio; Lozano, José

    2014-01-01

    Recurrent breast cancer occurring after the initial treatment is associated with poor outcome. A bimodal relapse pattern after surgery for primary tumor has been described with peaks of early and late recurrence occurring at about 2 and 5 years, respectively. Although several clinical and pathological features have been used to discriminate between low- and high-risk patients, the identification of molecular biomarkers with prognostic value remains an unmet need in the current management of breast cancer. Using microarray-based technology, we have performed a microRNA expression analysis in 71 primary breast tumors from patients that either remained disease-free at 5 years post-surgery (group A) or developed early (group B) or late (group C) recurrence. Unsupervised hierarchical clustering of microRNA expression data segregated tumors in two groups, mainly corresponding to patients with early recurrence and those with no recurrence. Microarray data analysis and RT-qPCR validation led to the identification of a set of 5 microRNAs (the 5-miRNA signature) differentially expressed between these two groups: miR-149, miR-10a, miR-20b, miR-30a-3p and miR-342-5p. All five microRNAs were down-regulated in tumors from patients with early recurrence. We show here that the 5-miRNA signature defines a high-risk group of patients with shorter relapse-free survival and has predictive value to discriminate non-relapsing versus early-relapsing patients (AUC = 0.993, p-value<0.05). Network analysis based on miRNA-target interactions curated by public databases suggests that down-regulation of the 5-miRNA signature in the subset of early-relapsing tumors would result in an overall increased proliferative and angiogenic capacity. In summary, we have identified a set of recurrence-related microRNAs with potential prognostic value to identify patients who will likely develop metastasis early after primary breast surgery. PMID:24632820

  15. MicroRNA and HER2-overexpressing Cancer

    PubMed Central

    Wang, Shizhen Emily; Lin, Ren-Jang

    2013-01-01

    The discovery of microRNAs (miRNAs) has opened up new avenues for studying cancer at the molecular level, featuring a post-genomic era of biomedical research. These non-coding regulatory RNA molecules of ~22 nucleotides have emerged as important cancer biomarkers, effectors, and targets. In this review, we focus on the dysregulated biogenesis and function of miRNAs in cancers with an overexpression of the proto-oncogene HER2. Many of the studies reviewed here were carried out in breast cancer, where HER2 overexpression has been extensively studied and HER2-targeted therapy practiced for more than a decade. MiRNA signatures that can be used to classify tumors with different HER2 status have been reported but little consensus can be established among various studies, emphasizing the needs for additional well-controlled profiling approaches and meta-analyses in large and well-balanced patient cohorts. We further discuss three aspects of microRNA dysregulation in or contribution to HER2-associated malignancies or therapies: (a) miRNAs that are up- or down-regulated by HER2 and mediate the downstream signaling of HER2; (b) miRNAs that suppress the expression of HER2 or a factor in HER2 receptor complexes, such as HER3; and (c) miRNAs that affect responses to anti-HER2 therapies. The regulatory mechanisms are elaborated using mainly examples of miR-205, miR-125, and miR-21. Understanding the regulation and function of miRNAs in HER2-overexpressing tumors shall shed new light on the pathogenic mechanisms of microRNAs and the HER2 proto-oncogene in cancer, as well as on individualized or combinatorial anti-HER2 therapies. PMID:25070783

  16. Transgenic Mouse Expressing Optical MicroRNA Reporter for Monitoring MicroRNA-124 Action during Development.

    PubMed

    Choi, Yoori; Hwang, Do Won; Kim, Mee Young; Kim, Joo Yeon; Sun, Woong; Lee, Dong Soo

    2016-01-01

    MicroRNAs (miRNAs) fine-tune target protein synthesis by suppressing gene expression, temporally changing along development and possibly in pathological conditions. A method to monitor the action of miRNAs in vivo shall help understand their dynamic behavior during development. In this study, we established a transgenic mouse harboring miR-124 responsive element in their luciferase-eGFP reporter transgenes which enabled monitoring the action of miR-124 in the brain and other organs in vivo by the bioluminescence imaging. The mouse model was produced and verified by imaging ex vivo so that luminescence by luciferase shone and then reduced during development with miR-124 expression. Bioluminescence dramatically decreased in the brain between embryonic day 13 and 16 as endogenous miR-124 expression increased, which sustained into adulthood. The inverse relationship of miR-124 expression was observed with luciferase bioluminescence and activity ex vivo as well as in vivo. Taken together, one can use this microRNA-transgenic mouse to investigate the temporal changes of microRNA action in vivo in the brain as well as in other organs. PMID:27462205

  17. Transgenic Mouse Expressing Optical MicroRNA Reporter for Monitoring MicroRNA-124 Action during Development

    PubMed Central

    Choi, Yoori; Hwang, Do won; Kim, Mee Young; Kim, Joo Yeon; Sun, Woong; Lee, Dong Soo

    2016-01-01

    MicroRNAs (miRNAs) fine-tune target protein synthesis by suppressing gene expression, temporally changing along development and possibly in pathological conditions. A method to monitor the action of miRNAs in vivo shall help understand their dynamic behavior during development. In this study, we established a transgenic mouse harboring miR-124 responsive element in their luciferase-eGFP reporter transgenes which enabled monitoring the action of miR-124 in the brain and other organs in vivo by the bioluminescence imaging. The mouse model was produced and verified by imaging ex vivo so that luminescence by luciferase shone and then reduced during development with miR-124 expression. Bioluminescence dramatically decreased in the brain between embryonic day 13 and 16 as endogenous miR-124 expression increased, which sustained into adulthood. The inverse relationship of miR-124 expression was observed with luciferase bioluminescence and activity ex vivo as well as in vivo. Taken together, one can use this microRNA-transgenic mouse to investigate the temporal changes of microRNA action in vivo in the brain as well as in other organs. PMID:27462205

  18. MicroRNA-155 and Its Role in Malignant Hematopoiesis

    PubMed Central

    Ranganath, Prajnya

    2015-01-01

    MicroRNA-155 (miR-155) is a multifunctional molecule involved in both normal and malignant hematopoiesis. It has been found to be involved in the pathogenesis of many different hematological malignancies with either an oncogenic or a tumor-repressor effect, depending on the nature of the cell and the type of malignancy. In particular, it has been strongly implicated in the causation of diffuse large B-cell lymphomas. This review focuses on the molecular interactions of miR-155, its oncogenic mechanisms, and its potential as an effective therapeutic target for the associated malignancies. PMID:26523117

  19. Regulating the Regulators: microRNA and Asthma

    PubMed Central

    2011-01-01

    One obstacle to developing an effective therapeutic strategy to treat or prevent asthma is that the fundamental causes of asthma are not totally understood. Asthma is thought to be a chronic TH2 immune-mediated inflammatory disease. Epigenetic changes are recognized to play a role in the initiation and maintenance of a TH2 response. MicroRNAs (miRNAs) are key epigenetic regulators of gene expression, and their expression is highly regulated, therefore, deregulation of miRNAs may play an important role in the pathogenesis of asthma. Profiling circulating miRNA might provide the highest specificity and sensitivity to diagnose asthma; similarly, correcting potential defects in the miRNA regulation network may lead to new therapeutic modalities to treat this disease. PMID:23282474

  20. MicroRNA Regulators of Anxiety and Metabolic Disorders.

    PubMed

    Meydan, Chanan; Shenhar-Tsarfaty, Shani; Soreq, Hermona

    2016-09-01

    Anxiety-related and metabolic disorders are under intense research focus. Anxiety-induced microRNAs (miRNAs) are emerging as regulators that are not only capable of suppressing inflammation but can also induce metabolic syndrome-related processes. We summarize here evidence linking miRNA pathways which share regulatory networks in metabolic and anxiety-related conditions. In particular, miRNAs involved in these disorders include regulators of acetylcholine signaling in the nervous system and their accompanying molecular machinery. These have been associated with anxiety-prone states in individuals, while also acting as inflammatory suppressors. In peripheral tissues, altered miRNA pathways can lead to dysregulated metabolism. Common pathways in metabolic and anxiety-related phenomena might offer an opportunity to reclassify 'healthy' and 'unhealthy', as well as metabolic and anxiety-prone biological states, and inform putative strategies to treat these disorders. PMID:27496210

  1. Effects of microRNA-21 and microRNA-24 inhibitors on neuronal apoptosis in ischemic stroke

    PubMed Central

    Liu, Wansheng; Chen, Xiaosheng; Zhang, Yu

    2016-01-01

    Objectives: The purpose of our study was aimed to investigate the effects of microRNA-21 (miR-21) and microRNA-24 (miR-24) inhibitors on ischemic stroke. Methods: MiR-21 inhibitor or miR-24 inhibitor was delivered to Sprague Dawley (SD) rats by continuous intracerebroventricular infusion. Two days later, middle cerebral artery occlusion (MCAO) was performed to induce ischemic stroke. Quantitative real-time PCR was performed to confirm transfection efficiency. The number of apoptotic neurons was detected using TUNEL method. Besides, primary hippocampal or cortical neuronal cultures were prepared from embryonic day 16-18 C57BL/6 mice. These cells were transfected with miR-21 inhibitor, miR-24 inhibitor, or negative scramble RNA. Then the cell viability was detected after transfection, as well as the protein levels of Caspase-3, B-cell lymphoma (Bcl)-xL, and heat shock protein (HSP) 70. Results: Both the levels of miR-21 and miR-24 were significantly reduced by transfection with inhibitors compared to control group or scramble RNA group (both P < 0.05). The apoptosis was significantly reduced in both hippocampal neuron and cortical neuron by miR-24 inhibitor rather than miR-21 inhibitor (P < 0.05), while the cell viability was significantly increased compared to the control group or the scramble group (P < 0.05). In addition, the levels of Bcl-xL and HSP70 were significantly increased, and the levels of Caspase-3 were statistically decreased by transfection with miR-24 inhibitor. Conclusion: MiRNA-24 but not miR-21 inhibitor prevents apoptosis in ischemic stroke by regulation of Bcl-xL, Caspase-3 and HSP70. PMID:27508039

  2. A population-based statistical approach identifies parameters characteristic of human microRNA-mRNA interactions

    PubMed Central

    Smalheiser, Neil R; Torvik, Vetle I

    2004-01-01

    Background MicroRNAs are ~17–24 nt. noncoding RNAs found in all eukaryotes that degrade messenger RNAs via RNA interference (if they bind in a perfect or near-perfect complementarity to the target mRNA), or arrest translation (if the binding is imperfect). Several microRNA targets have been identified in lower organisms, but only one mammalian microRNA target has yet been validated experimentally. Results We carried out a population-wide statistical analysis of how human microRNAs interact complementarily with human mRNAs, looking for characteristics that differ significantly as compared with scrambled control sequences. These characteristics were used to identify a set of 71 outlier mRNAs unlikely to have been hit by chance. Unlike the case in C. elegans and Drosophila, many human microRNAs exhibited long exact matches (10 or more bases in a row), up to and including perfect target complementarity. Human microRNAs hit outlier mRNAs within the protein coding region about 2/3 of the time. And, the stretches of perfect complementarity within microRNA hits onto outlier mRNAs were not biased near the 5'-end of the microRNA. In several cases, an individual microRNA hit multiple mRNAs that belonged to the same functional class. Conclusions The analysis supports the notion that sequence complementarity is the basis by which microRNAs recognize their biological targets, but raises the possibility that human microRNA-mRNA target interactions follow different rules than have been previously characterized in Drosophila and C. elegans. PMID:15453917

  3. Crucial microRNAs and genes of human primary breast cancer explored by microRNA-mRNA integrated analysis.

    PubMed

    Yang, Yang; Xing, Yiqiao; Liang, Chaoqun; Hu, Liya; Xu, Fei; Chen, Yuan

    2015-07-01

    This study aimed to screen potential microRNAs (miRNAs) and genes related to human primary breast cancer. The gene and miRNA expression profile data of GSE19783 was obtained from Gene Expression Omnibus. The matched messenger RNA (mRNA) and miRNA expression profiles of 100 human primary breast cancer samples were chosen for further analysis. The miRNA-gene regulatory modules were screened via iterative multiplicative updating algorithm. The potential functions of genes in modules were predicted by functional and pathway enrichment analysis; meanwhile, the potential functions of miRNAs were predicted by functional enrichment analysis. Furthermore, miRNA-miRNA functional synergistic network and miRNA-miRNA co-regulatory network were constructed. Totally, 16 miRNA-gene modules were screened, containing 222 miRNA-gene interactions. The genes in these modules were mainly related to breast cancer. Genes in module 6 (e.g., SFRP1) were enriched in cell junction assembly; genes in module 8 and 12 (e.g., ESR1 and ERBB4) were significantly implicated in mammary gland alveolus and lobule development. Meanwhile, genes in module 12 (e.g., ERBB4) were enriched in the pathway of endocytosis. Besides, several miRNAs (e.g., miR-375) were enriched in inflammatory cell apoptotic process; some other miRNAs (e.g., miR-139-5p and miR-9) were enriched in response to vitamin D. Additionally, miR-139-5p with several other miRNAs (e.g., miR-9) co-regulated SFRP1; miR-375, miR-592, and miR-135a co-regulated ESR1 and ERBB4. Some miRNAs (e.g., miR-139-5p and miR-9) and their target gene SFRP1, as well as several other miRNAs (e.g., miR-375, miR-592, and miR-135a) and their target genes (e.g., ESR1 and ERBB4), might be crucial in the pathogenesis of primary breast cancer. PMID:25680412

  4. MicroRNA profiles classify papillary renal cell carcinoma subtypes

    PubMed Central

    Wach, S; Nolte, E; Theil, A; Stöhr, C; T Rau, T; Hartmann, A; Ekici, A; Keck, B; Taubert, H; Wullich, B

    2013-01-01

    Background: Besides the conventional clear-cell renal cell carcinoma (ccRCC), papillary RCC (pRCC) is the second most common renal malignancy. Papillary RCCs can further be subdivided into two distinct subtypes. Although a clinical relevance of pRCC subtyping has been shown, little is known about the molecular characteristics of both pRCC subtypes. Methods: We performed microarray-based microRNA (miRNA) expression profiling of primary ccRCC and pRCC cases. A subset of miRNAs was identified and used to establish a classification model for ccRCC, pRCC types 1 and 2 and normal tissue. Furthermore, we performed gene set enrichment analysis with the predicted miRNA target genes. Results: Only five miRNAs (miR-145, -200c, -210, -502-3p and let-7c) were sufficient to identify the samples with high accuracy. In a collection of 111 tissue samples, 73.9% were classified correctly. An enrichment of miRNA target genes in the family of multidrug-resistance proteins was noted in all tumours. Several components of the Jak-STAT signalling pathway might be targets for miRNAs that define pRCC tumour subtypes. Conclusion: MicroRNAs are able to accurately classify RCC samples. Deregulated miRNAs might contribute to the high chemotherapy resistance of RCC. Furthermore, our results indicate that pRCC type 2 tumours could be dependent on oncogenic MYC signalling. PMID:23799849

  5. MicroRNA-Detargeted Mengovirus for Oncolytic Virotherapy

    PubMed Central

    Ruiz, Autumn J.; Hadac, Elizabeth M.; Nace, Rebecca A.

    2016-01-01

    ABSTRACT Mengovirus, a member of the Picornaviridae family, has a broad cell tropism and can cause encephalitis and myocarditis in multiple mammalian species. Attenuation has been achieved by shortening the polycytidine tract in the 5′ noncoding region (NCR). A poly(C)-truncated strain of mengovirus, vMC24, resulted in significant tumor regression in immunocompetent BALB/c mice bearing syngeneic MPC-11 plasmacytomas, but the associated toxicities were unacceptable. To enhance its safety profile, microRNA target sequences complementary to miR-124 or miR-125 (enriched in nervous tissue), miR-133 and miR-208 (enriched in cardiac tissue), or miR-142 (control; enriched in hematopoietic tissues) were inserted into the vMC24 NCRs. The microRNA-detargeted viruses showed reduced replication and cell killing specifically in cells expressing the cognate microRNAs, but certain insertions additionally were associated with nonspecific suppression of viral fitness in vivo. In vivo toxicity testing confirmed that miR-124 targets within the 5′ NCR suppressed virus replication in the central nervous system while miR-133 and miR-208 targets in the 3′ NCR suppressed viral replication in cardiac tissue. A dual-detargeted virus named vMC24-NC, with miR-124 targets in the 5′ NCR and miR-133 plus miR-208 targets in the 3′ NCR, showed the suppression of replication in both nervous and cardiac tissues but retained full oncolytic potency when administered by intratumoral (106 50% tissue culture infectious doses [TCID50]) or intravenous (107 to 108 TCID50) injection into BALB/c mice bearing MPC-11 plasmacytomas. Overall survival of vMC24-NC-treated tumor-bearing mice was significantly improved compared to that of nontreated mice. MicroRNA-detargeted mengoviruses offer a promising oncolytic virotherapy platform that merits further development for clinical translation. IMPORTANCE The clinical potential of oncolytic virotherapy for cancer treatment has been well demonstrated

  6. Functional Analysis of microRNA in Multiple Myeloma.

    PubMed

    Di Martino, Maria Teresa; Amodio, Nicola; Tassone, Pierfrancesco; Tagliaferri, Pierosandro

    2016-01-01

    MicroRNAs (miRNAs) are short non coding RNAs that regulate the gene expression and play a relevant role in physiopathological mechanisms such as development, proliferation, death, and differentiation of normal and cancer cells. Recently, abnormal expression of miRNAs has been reported in most of solid or hematopoietic malignancies, including multiple myeloma (MM), where miRNAs have been found deeply dysregulated and act as oncogenes or tumor suppressors. Presently, the most recognized approach for definition of miRNA portraits is based on microarray profiling analysis. We here describe a workflow based on the identification of dysregulated miRNAs in plasma cells from MM patients based on Affymetrix technology. We describe how it is possible to search miRNA putative targets performing whole gene expression profile on MM cell lines transfected with miRNA mimics or inhibitors followed by luciferase reporter assay to analyze the specific targeting of the 3'untranslated region (UTR) sequence of a mRNA by selected miRNAs. These technological approaches are suitable strategies for the identification of relevant druggable targets in MM. PMID:25971914

  7. Identification and Expression Profiles of microRNA in Dolphin.

    PubMed

    Segawa, Takao; Kobayashi, Yuki; Inamoto, Satoko; Suzuki, Miwa; Endoh, Tomoko; Itou, Takuya

    2016-02-01

    Recently, microRNAs (miRNAs) are focused on the role of biomarker because they are stable in serum and plasma, and some of them express in the specific organs and increase with the organ injury. Thus miRNAs may be very useful as biomarkers for monitoring the health and condition of dolphins and for detecting disorders in aquariums. Here, a small RNA library was made from dolphin lung, liver and spleen, and miRNA expression patterns were then determined for 15 different tissues. We identified 62 conserved miRNA homologs in the dolphin small RNA library and found high expression miRNAs in specific tissues: miR-125b and miR-221 were highly expressed in brain, miR-23b in heart, miR-199a and miR-223 in lung, and miR-122-5p in liver. Some of these tissue-enriched miRNAs may be useful as specific and sensitive diagnostic blood biomarkers for organ injury in dolphins. PMID:26853874

  8. MicroRNA in carcinogenesis & cancer diagnostics: A new paradigm

    PubMed Central

    Ahmad, Javed; Hasnain, Seyed E.; Siddiqui, Maqsood A.; Ahamed, Maqusood; Musarrat, Javed; Al-Khedhairy, Abdulaziz A.

    2013-01-01

    MicroRNAs (miRNAs) are small 22-25 nucleotides long non-coding RNAs, that are conserved during evolution, and control gene expression in metazoan animals, plants, viruses, and bacteria primarily at post-transcriptional and transcriptional levels. MiRNAs ultimately regulate target gene expression by degrading the corresponding mRNA and/or inhibiting their translation. Currently, the critical functions of miRNAs have been established in regulating immune system, cell proliferation, differentiation and development, cancer and cell cycle by as yet unknown control mechanism. MiRNAs play an essential role in malignancy, and as tumour suppressors and oncogenes. Thus, discovery of new miRNAs will probably change the landscape of cancer genetics. Significantly different miRNA profiles can be assigned to various types of tumours, which could serve as phenotypic signatures for different cancers for their exploitation in cancer diagnostics, prognostics and therapeutics. If miRNA profiles can accurately predict malignancies, this technology could be exploited as a tool to surmount the diagnostic challenges. This review provides comprehensive and systematic information on miRNA biogenesis and their implications in human health. PMID:23703335

  9. MicroRNA Targeting to Modulate Tumor Microenvironment

    PubMed Central

    Kuninty, Praneeth R.; Schnittert, Jonas; Storm, Gert; Prakash, Jai

    2016-01-01

    Communication between stromal cells and tumor cells initiates tumor growth, angiogenesis, invasion, and metastasis. Stromal cells include cancer-associated fibroblasts, tumor-associated macrophages, pericytes, endothelial cells, and infiltrating immune cells. MicroRNAs (miRNAs) in the tumor microenvironment have emerged as key players involved in the development of cancer and its progression. miRNAs are small endogenous non-protein-coding RNAs that negatively regulate the expression of multiple target genes at post-transcriptional level and thereby control many cellular processes. In this review, we provide a comprehensive overview of miRNAs dysregulated in different stromal cells and their impact on the regulation of intercellular crosstalk in the tumor microenvironment. We also discuss the therapeutic significance potential of miRNAs to modulate the tumor microenvironment. Since miRNA delivery is quite challenging and the biggest hurdle for clinical translation of miRNA therapeutics, we review various non-viral miRNA delivery systems that can potentially be used for targeting miRNA to stromal cells within the tumor microenvironment. PMID:26835418

  10. MicroRNA and messenger RNA profiling reveals new biomarkers and mechanisms for RDX induced neurotoxicity

    PubMed Central

    2014-01-01

    Background RDX is a well-known pollutant to induce neurotoxicity. MicroRNAs (miRNA) and messenger RNA (mRNA) profiles are useful tools for toxicogenomics studies. It is worthy to integrate MiRNA and mRNA expression data to understand RDX-induced neurotoxicity. Results Rats were treated with or without RDX for 48 h. Both miRNA and mRNA profiles were conducted using brain tissues. Nine miRNAs were significantly regulated by RDX. Of these, 6 and 3 miRNAs were up- and down-regulated respectively. The putative target genes of RDX-regulated miRNAs were highly nervous system function genes and pathways enriched. Fifteen differentially genes altered by RDX from mRNA profiles were the putative targets of regulated miRNAs. The induction of miR-71, miR-27ab, miR-98, and miR-135a expression by RDX, could reduce the expression of the genes POLE4, C5ORF13, SULF1 and ROCK2, and eventually induce neurotoxicity. Over-expression of miR-27ab, or reduction of the expression of unknown miRNAs by RDX, could up-regulate HMGCR expression and contribute to neurotoxicity. RDX regulated immune and inflammation response miRNAs and genes could contribute to RDX- induced neurotoxicity and other toxicities as well as animal defending reaction response to RDX exposure. Conclusions Our results demonstrate that integrating miRNA and mRNA profiles is valuable to indentify novel biomarkers and molecular mechanisms for RDX-induced neurological disorder and neurotoxicity. PMID:25559034

  11. MicroRNA Predictors of Longevity in Caenorhabditis elegans

    PubMed Central

    Pincus, Zachary; Smith-Vikos, Thalyana; Slack, Frank J.

    2011-01-01

    Neither genetic nor environmental factors fully account for variability in individual longevity: genetically identical invertebrates in homogenous environments often experience no less variability in lifespan than outbred human populations. Such variability is often assumed to result from stochasticity in damage accumulation over time; however, the identification of early-life gene expression states that predict future longevity would suggest that lifespan is least in part epigenetically determined. Such “biomarkers of aging,” genetic or otherwise, nevertheless remain rare. In this work, we sought early-life differences in organismal robustness in unperturbed individuals and examined the utility of microRNAs, known regulators of lifespan, development, and robustness, as aging biomarkers. We quantitatively examined Caenorhabditis elegans reared individually in a novel apparatus and observed throughout their lives. Early-to-mid–adulthood measures of homeostatic ability jointly predict 62% of longevity variability. Though correlated, markers of growth/muscle maintenance and of metabolic by-products (“age pigments”) report independently on lifespan, suggesting that graceful aging is not a single process. We further identified three microRNAs in which early-adulthood expression patterns individually predict up to 47% of lifespan differences. Though expression of each increases throughout this time, mir-71 and mir-246 correlate with lifespan, while mir-239 anti-correlates. Two of these three microRNA “biomarkers of aging” act upstream in insulin/IGF-1–like signaling (IIS) and other known longevity pathways, thus we infer that these microRNAs not only report on but also likely determine longevity. Thus, fluctuations in early-life IIS, due to variation in these microRNAs and from other causes, may determine individual lifespan. PMID:21980307

  12. Cross Talk Between MicroRNA and Coding Cancer Genes

    PubMed Central

    Kunej, T; Godnic, I; Horvat, S; Zorc, M; Calin, GA

    2012-01-01

    MicroRNAs (miRNAs) are a class of non-coding RNAs (ncRNAs) and post-transcriptional gene regulators shown to be involved in pathogenesis of all types of human cancers. Their aberrant expression as tumor suppressors can lead to cancerogenesis by inhibiting malignant potential, or when acting as oncogenes, by activating malignant potential. Differential expression of miRNA genes in tumorous tissues can occur due to several factors including positional effects when mapping to cancer-associated genomic regions, epigenetic mechanisms and malfunctioning of the miRNA processing machinery, all of which can contribute to a complex miRNA-mediated gene network misregulation. They may increase or decrease expression of protein-coding genes, can target 3’-UTR or other genic regions (5'-UTR, promoter, coding sequences), and can function in various subcellular compartments, developmental and metabolic processes. Because expanding research on miRNA-cancer associations has already produced large amounts of data, our main objective here was to summarize main findings and critically examine the intricate network connecting the miRNAs and coding genes in regulatory mechanisms, their function and phenotypic consequences for cancer. By examining such interactions we aimed to gain insights for development of new diagnostic markers as well as identify potential venues for more selective tumor therapy. To enable efficient examination of the main past and current miRNA discoveries, we developed a web based miRNA timeline tool that will be regularly updated (http://www.integratomics-time.com/miRNA_timeline). Further development of this tool will be directed at providing additional analyses to clarify complex network interactions between miRNAs, other classes of ncRNAs and protein coding genes and their involvement in development of diseases including cancer. This tool therefore provides curated relevant information about the miRNA basic research and therapeutic application all at hand on

  13. MicroRNA-Offset RNA Alters Gene Expression and Cell Proliferation

    PubMed Central

    Zhao, Jin; Schnitzler, Gavin R.; Iyer, Lakshmanan K.; Aronovitz, Mark J.; Baur, Wendy E.; Karas, Richard H.

    2016-01-01

    MicroRNA-offset RNAs (moRs) were first identified in simple chordates and subsequently in mouse and human cells by deep sequencing of short RNAs. MoRs are derived from sequences located immediately adjacent to microRNAs (miRs) in the primary miR (pri-miR). Currently moRs are considered to be simply a by-product of miR biosynthesis that lack biological activity. Here we show for the first time that a moR is biologically active. We demonstrate that endogenous or over-expressed moR-21 significantly alters gene expression and inhibits the proliferation of vascular smooth muscle cells (VSMC). In addition, we find that miR-21 and moR-21 may regulate different genes in a given pathway and can oppose each other in regulating certain genes. We report that there is a “seed region” of moR-21 as well as a “seed match region” in the target gene 3’UTR that are indispensable for moR-21-mediated gene down-regulation. We further demonstrate that moR-21-mediated gene repression is Argonaute 2 (Ago2) dependent. Taken together, these findings provide the first evidence that microRNA offset RNA alters gene expression and is biologically active. PMID:27276022

  14. Hierarchical Generative Biclustering for MicroRNA Expression Analysis

    NASA Astrophysics Data System (ADS)

    Caldas, José; Kaski, Samuel

    Clustering methods are a useful and common first step in gene expression studies, but the results may be hard to interpret. We bring in explicitly an indicator of which genes tie each cluster, changing the setup to biclustering. Furthermore, we make the indicators hierarchical, resulting in a hierarchy of progressively more specific biclusters. A non-parametric Bayesian formulation makes the model rigorous and yet flexible, and computations feasible. The formulation additionally offers a natural information retrieval relevance measure that allows relating samples in a principled manner. We show that the model outperforms other four biclustering procedures in a large miRNA data set. We also demonstrate the model's added interpretability and information retrieval capability in a case study that highlights the potential and novel role of miR-224 in the association between melanoma and non-Hodgkin lymphoma. Software is publicly available.

  15. Hierarchical generative biclustering for microRNA expression analysis.

    PubMed

    Caldas, José; Kaski, Samuel

    2011-03-01

    Clustering methods are a useful and common first step in gene expression studies, but the results may be hard to interpret. We bring in explicitly an indicator of which genes tie each cluster, changing the setup to biclustering. Furthermore, we make the indicators hierarchical, resulting in a hierarchy of progressively more specific biclusters. A non-parametric Bayesian formulation makes the model rigorous yet flexible and computations feasible. The model can additionally be used in information retrieval for relating relevant samples. We show that the model outperforms four other biclustering procedures on a large miRNA data set. We also demonstrate the model's added interpretability and information retrieval capability in a case study. Software is publicly available at http://research.ics.tkk.fi/mi/software/treebic/. PMID:21385032

  16. A comparative analysis of high-throughput platforms for validation of a circulating microRNA signature in diabetic retinopathy

    PubMed Central

    Farr, Ryan J.; Januszewski, Andrzej S.; Joglekar, Mugdha V.; Liang, Helena; McAulley, Annie K.; Hewitt, Alex W.; Thomas, Helen E.; Loudovaris, Tom; Kay, Thomas W. H.; Jenkins, Alicia; Hardikar, Anandwardhan A.

    2015-01-01

    MicroRNAs are now increasingly recognized as biomarkers of disease progression. Several quantitative real-time PCR (qPCR) platforms have been developed to determine the relative levels of microRNAs in biological fluids. We systematically compared the detection of cellular and circulating microRNA using a standard 96-well platform, a high-content microfluidics platform and two ultra-high content platforms. We used extensive analytical tools to compute inter- and intra-run variability and concordance measured using fidelity scoring, coefficient of variation and cluster analysis. We carried out unprejudiced next generation sequencing to identify a microRNA signature for Diabetic Retinopathy (DR) and systematically assessed the validation of this signature on clinical samples using each of the above four qPCR platforms. The results indicate that sensitivity to measure low copy number microRNAs is inversely related to qPCR reaction volume and that the choice of platform for microRNA biomarker validation should be made based on the abundance of miRNAs of interest. PMID:26035063

  17. MicroRNA Expression Signature in Degenerative Aortic Stenosis

    PubMed Central

    2016-01-01

    Degenerative aortic stenosis, characterized by narrowing of the exit of the left ventricle of the heart, has become the most common valvular heart disease in the elderly. The aim of this study was to investigate the microRNA (miRNA) signature in degenerative AS. Through microarray analysis, we identified the miRNA expression signature in the tissue samples from healthy individuals (n = 4) and patients with degenerative AS (n = 4). Six miRNAs (hsa-miR-193a-3p, hsa-miR-29b-1-5p, hsa-miR-505-5p, hsa-miR-194-5p, hsa-miR-99b-3p, and hsa-miR-200b-3p) were overexpressed and 14 (hsa-miR-3663-3p, hsa-miR-513a-5p, hsa-miR-146b-5p, hsa-miR-1972, hsa-miR-718, hsa-miR-3138, hsa-miR-21-5p, hsa-miR-630, hsa-miR-575, hsa-miR-301a-3p, hsa-miR-636, hsa-miR-34a-3p, hsa-miR-21-3p, and hsa-miR-516a-5p) were downregulated in aortic tissue from AS patients. GeneSpring 13.1 was used to identify potential human miRNA target genes by comparing a 3-way comparison of predictions from TargetScan, PITA, and microRNAorg databases. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were performed to identify potential pathways and functional annotations associated with AS. Twenty miRNAs were significantly differentially expressed between patients with AS samples and normal controls and identified potential miRNA targets and molecular pathways associated with this morbidity. This study describes the miRNA expression signature in degenerative AS and provides an improved understanding of the molecular pathobiology of this disease. PMID:27579316

  18. MicroRNA Expression Signature in Degenerative Aortic Stenosis.

    PubMed

    Shi, Jing; Liu, Hui; Wang, Hui; Kong, Xiangqing

    2016-01-01

    Degenerative aortic stenosis, characterized by narrowing of the exit of the left ventricle of the heart, has become the most common valvular heart disease in the elderly. The aim of this study was to investigate the microRNA (miRNA) signature in degenerative AS. Through microarray analysis, we identified the miRNA expression signature in the tissue samples from healthy individuals (n = 4) and patients with degenerative AS (n = 4). Six miRNAs (hsa-miR-193a-3p, hsa-miR-29b-1-5p, hsa-miR-505-5p, hsa-miR-194-5p, hsa-miR-99b-3p, and hsa-miR-200b-3p) were overexpressed and 14 (hsa-miR-3663-3p, hsa-miR-513a-5p, hsa-miR-146b-5p, hsa-miR-1972, hsa-miR-718, hsa-miR-3138, hsa-miR-21-5p, hsa-miR-630, hsa-miR-575, hsa-miR-301a-3p, hsa-miR-636, hsa-miR-34a-3p, hsa-miR-21-3p, and hsa-miR-516a-5p) were downregulated in aortic tissue from AS patients. GeneSpring 13.1 was used to identify potential human miRNA target genes by comparing a 3-way comparison of predictions from TargetScan, PITA, and microRNAorg databases. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were performed to identify potential pathways and functional annotations associated with AS. Twenty miRNAs were significantly differentially expressed between patients with AS samples and normal controls and identified potential miRNA targets and molecular pathways associated with this morbidity. This study describes the miRNA expression signature in degenerative AS and provides an improved understanding of the molecular pathobiology of this disease. PMID:27579316

  19. A conformation-induced fluorescence method for microRNA detection.

    PubMed

    Aw, Sherry S; Tang, Melissa Xm; Teo, Yin Nah; Cohen, Stephen M

    2016-06-01

    MicroRNAs play important roles in a large variety of biological systems and processes through their regulation of target mRNA expression, and show promise as clinical biomarkers. However, their small size presents challenges for tagging or direct detection. Innovation in techniques to sense and quantify microRNAs may aid research into novel aspects of microRNA biology and contribute to the development of diagnostics. By introducing an additional stem loop into the fluorescent RNA Spinach and altering its 3' and 5' ends, we have generated a new RNA, Pandan, that functions as the basis for a microRNA sensor. Pandan contains two sequence-variable stem loops that encode complementary sequence for a target microRNA of interest. In its sensor form, it requires the binding of a target microRNA in order to reconstitute the RNA scaffold for fluorophore binding and fluorescence. Binding of the target microRNA resulted in large changes in fluorescence intensity. The median fold change in fluorescence observed for the sensors tested was ∼50-fold. Pandan RNA sensors exhibit good signal-to-noise ratios, and can detect their target microRNAs within complex RNA mixtures. PMID:26951376

  20. A conformation-induced fluorescence method for microRNA detection

    PubMed Central

    Aw, Sherry S.; Tang, Melissa XM; Teo, Yin Nah; Cohen, Stephen M.

    2016-01-01

    MicroRNAs play important roles in a large variety of biological systems and processes through their regulation of target mRNA expression, and show promise as clinical biomarkers. However, their small size presents challenges for tagging or direct detection. Innovation in techniques to sense and quantify microRNAs may aid research into novel aspects of microRNA biology and contribute to the development of diagnostics. By introducing an additional stem loop into the fluorescent RNA Spinach and altering its 3′ and 5′ ends, we have generated a new RNA, Pandan, that functions as the basis for a microRNA sensor. Pandan contains two sequence-variable stem loops that encode complementary sequence for a target microRNA of interest. In its sensor form, it requires the binding of a target microRNA in order to reconstitute the RNA scaffold for fluorophore binding and fluorescence. Binding of the target microRNA resulted in large changes in fluorescence intensity. The median fold change in fluorescence observed for the sensors tested was ∼50-fold. Pandan RNA sensors exhibit good signal-to-noise ratios, and can detect their target microRNAs within complex RNA mixtures. PMID:26951376

  1. MicroRNA inhibition fine-tunes and provides robustness to the restriction point switch of the cell cycle.

    PubMed

    Del Rosario, Ricardo C H; Damasco, Joseph Ray Clarence G; Aguda, Baltazar D

    2016-01-01

    The restriction point marks a switch in G1 from growth factor-dependent to growth factor-independent progression of the cell cycle. The proper regulation of this switch is important for normal cell processes; aberrations could result in a number of diseases such as cancer, neurodegenerative disorders, stroke and myocardial infarction. To further understand the regulation of the restriction point, we extended a mathematical model of the Rb-E2F pathway to include members of the microRNA cluster miR-17-92. Our mathematical analysis shows that microRNAs play an essential role in fine-tuning and providing robustness to the switch. We also demonstrate how microRNA regulation can steer cells in or out of cancer states. PMID:27610602

  2. MicroRNA inhibition fine-tunes and provides robustness to the restriction point switch of the cell cycle

    PubMed Central

    del Rosario, Ricardo C. H.; Damasco, Joseph Ray Clarence G.; Aguda, Baltazar D.

    2016-01-01

    The restriction point marks a switch in G1 from growth factor-dependent to growth factor-independent progression of the cell cycle. The proper regulation of this switch is important for normal cell processes; aberrations could result in a number of diseases such as cancer, neurodegenerative disorders, stroke and myocardial infarction. To further understand the regulation of the restriction point, we extended a mathematical model of the Rb-E2F pathway to include members of the microRNA cluster miR-17-92. Our mathematical analysis shows that microRNAs play an essential role in fine-tuning and providing robustness to the switch. We also demonstrate how microRNA regulation can steer cells in or out of cancer states. PMID:27610602

  3. MicroRNA (miRNA) expression is regulated by butyrate induced epigenetic modulation of gene expression in bovine cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We present evidence that butyrate induced histone acetylation regulates miRNA expression. MicroRNA expression microarray profiling revealed that 35 miRNA transcripts are significantly (p <0.05) differentially expressed after cells were treated with 10 mM butyrate. Among them, 11 transcripts are dif...

  4. MicroRNA in TLR signaling and endotoxin tolerance

    PubMed Central

    Nahid, Md A; Satoh, Minoru; Chan, Edward KL

    2011-01-01

    Toll-like receptors (TLRs) in innate immune cells are the prime cellular sensors for microbial components. TLR activation leads to the production of proinflammatory mediators and thus TLR signaling must be properly regulated by various mechanisms to maintain homeostasis. TLR4-ligand lipopolysaccharide (LPS)-induced tolerance or cross-tolerance is one such mechanism, and it plays an important role in innate immunity. Tolerance is established and sustained by the activity of the microRNA miR-146a, which is known to target key elements of the myeloid differentiation factor 88 (MyD88) signaling pathway, including IL-1 receptor-associated kinase (IRAK1), IRAK2 and tumor-necrosis factor (TNF) receptor-associated factor 6 (TRAF6). In this review, we comprehensively examine the TLR signaling involved in innate immunity, with special focus on LPS-induced tolerance. The function of TLR ligand-induced microRNAs, including miR-146a, miR-155 and miR-132, in regulating inflammatory mediators, and their impact on the immune system and human diseases, are discussed. Modulation of these microRNAs may affect TLR pathway activation and help to develop therapeutics against inflammatory diseases. PMID:21822296

  5. Silencing microRNA-155 ameliorates experimental autoimmune encephalomyelitis.

    PubMed

    Murugaiyan, Gopal; Beynon, Vanessa; Mittal, Akanksha; Joller, Nicole; Weiner, Howard L

    2011-09-01

    IFN-γ-producing Th1 and IL-17-producing Th17 cells are the key participants in various autoimmune diseases, including multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE). Although both of these T cell subsets are known to be regulated by specific transcription factors and cytokines, the role of microRNAs that control these two inflammatory T cell subsets and whether targeting microRNAs can have therapeutic effects are not known. In this study, we show that microRNA-155 (Mir-155) expression is elevated in CD4(+) T cells during EAE, and Mir-155(-/-) mice had a delayed course and reduced severity of disease and less inflammation in the CNS. The attenuation of EAE in Mir-155(-/-) mice was associated with a decrease in Th1 and Th17 responses in the CNS and peripheral lymphoid organs. The T cell-intrinsic function of Mir-155(-/-) was demonstrated by the resistance of Mir-155(-/-) CD4(+) T cell-repleted Rag-1(-/-) mice to EAE. Finally, we found that anti-Mir-155 treatment reduced clinical severity of EAE when given before and after the appearance of clinical symptoms. These findings demonstrate that Mir-155 confers susceptibility to EAE by affecting inflammatory T cell responses and identify Mir-155 as a new target for therapeutic intervention in multiple sclerosis. PMID:21788439

  6. Osteoblast-Targeting-Peptide Modified Nanoparticle for siRNA/microRNA Delivery.

    PubMed

    Sun, Yao; Ye, Xiongzhen; Cai, Mingxiang; Liu, Xiangning; Xiao, Jia; Zhang, Chenyang; Wang, Yayu; Yang, Li; Liu, Jiafan; Li, Shannai; Kang, Chen; Zhang, Bin; Zhang, Qi; Wang, Zuolin; Hong, An; Wang, Xiaogang

    2016-06-28

    Antiosteoporosis gene-based drug development strategies are presently focused on targeting osteoblasts to either suppress bone loss or increase bone mass. Although siRNA/microRNA-based gene therapy has enormous potential, it is severely limited by the lack of specific cell-targeting delivery systems. We report an osteoblast-targeting peptide (SDSSD) that selectively binds to osteoblasts via periostin. We developed SDSSD-modified polyurethane (PU) nanomicelles encapsulating siRNA/microRNA that delivers drugs to osteoblasts; the data showed that SDSSD-PU could selectively target not only bone-formation surfaces but also osteoblasts without overt toxicity or eliciting an immune response in vivo. We used the SDSSD-PU delivery system to deliver anti-miR-214 to osteoblasts and our results showed increased bone formation, improved bone microarchitecture, and increased bone mass in an ovariectomized osteoporosis mouse model. SDSSD-PU may be a useful osteoblast-targeting small nucleic acid delivery system that could be used as an anabolic strategy to treat osteoblast-induced bone diseases. PMID:27176123

  7. MicroRNA-10 modulates Hox genes expression during Nile tilapia embryonic development.

    PubMed

    Giusti, Juliana; Pinhal, Danillo; Moxon, Simon; Campos, Camila Lovaglio; Münsterberg, Andrea; Martins, Cesar

    2016-05-01

    Hox gene clusters encode a family of transcription factors that govern anterior-posterior axis patterning during embryogenesis in all bilaterian animals. The time and place of Hox gene expression are largely determined by the relative position of each gene within its cluster. Furthermore, Hox genes were shown to have their expression fine-tuned by regulatory microRNAs (miRNAs). However, the mechanisms of miRNA-mediated regulation of these transcription factors during fish early development remain largely unknown. Here we have profiled three highly expressed miR-10 family members of Nile tilapia at early embryonic development, determined their genomic organization as well as performed functional experiments for validation of target genes. Quantitative analysis during developmental stages showed miR-10 family expression negatively correlates with the expression of HoxA3a, HoxB3a and HoxD10a genes, as expected for bona fide miRNA-mRNA interactions. Moreover, luciferase assays demonstrated that HoxB3a and HoxD10a are targeted by miR-10b-5p. Overall, our data indicate that the miR-10 family directly regulates members of the Hox gene family during Nile tilapia embryogenesis. PMID:26980108

  8. Identification of microRNAs by small RNA deep sequencing for synthetic microRNA mimics to control Spodoptera exigua.

    PubMed

    Zhang, Yu Liang; Huang, Qi Xing; Yin, Guo Hua; Lee, Samantha; Jia, Rui Zong; Liu, Zhi Xin; Yu, Nai Tong; Pennerman, Kayla K; Chen, Xin; Guo, An Ping

    2015-02-25

    Beet armyworm, Spodoptera exigua, is a major pest of cotton around the world. With the increase of resistance to Bacillus thuringiensis (Bt) toxin in transgenic cotton plants, there is a need to develop an alternative control approach that can be used in combination with Bt transgenic crops as part of resistance management strategies. MicroRNAs (miRNAs), a non-coding small RNA family (18-25 nt), play crucial roles in various biological processes and over-expression of miRNAs has been shown to interfere with the normal development of insects. In this study, we identified 127 conserved miRNAs in S. exigua by using small RNA deep sequencing technology. From this, we tested the effects of 11 miRNAs on larval development. We found three miRNAs, Sex-miR-10-1a, Sex-miR-4924, and Sex-miR-9, to be differentially expressed during larval stages of S. exigua. Oral feeding experiments using synthetic miRNA mimics of Sex-miR-10-1a, Sex-miR-4924, and Sex-miR-9 resulted in suppressed growth of S. exigua and mortality. Over-expression of Sex-miR-4924 caused a significant reduction in the expression level of chitinase 1 and caused abortive molting in the insects. Therefore, we demonstrated a novel approach of using miRNA mimics to control S. exigua development. PMID:25528266

  9. Genome-wide analysis of microRNA and mRNA expression signatures in cancer

    PubMed Central

    Li, Ming-hui; Fu, Sheng-bo; Xiao, Hua-sheng

    2015-01-01

    Cancer is an extremely diverse and complex disease that results from various genetic and epigenetic changes such as DNA copy-number variations, mutations, and aberrant mRNA and/or protein expression caused by abnormal transcriptional regulation. The expression profiles of certain microRNAs (miRNAs) and messenger RNAs (mRNAs) are closely related to cancer progression stages. In the past few decades, DNA microarray and next-generation sequencing techniques have been widely applied to identify miRNA and mRNA signatures for cancers on a genome-wide scale and have provided meaningful insights into cancer diagnosis, prognosis and personalized medicine. In this review, we summarize the progress in genome-wide analysis of miRNAs and mRNAs as cancer biomarkers, highlighting their diagnostic and prognostic roles. PMID:26299954

  10. microRNA expression profiling identifies molecular signatures associated with anaplastic large cell lymphoma

    PubMed Central

    Liu, Cuiling; Iqbal, Javeed; Teruya-Feldstein, Julie; Shen, Yulei; Dabrowska, Magdalena Julia; Dybkaer, Karen; Lim, Megan S.; Piva, Roberto; Barreca, Antonella; Pellegrino, Elisa; Spaccarotella, Elisa; Lachel, Cynthia M.; Kucuk, Can; Jiang, Chun-Sun; Hu, Xiaozhou; Bhagavathi, Sharathkumar; Greiner, Timothy C.; Weisenburger, Dennis D.; Aoun, Patricia; Perkins, Sherrie L.; McKeithan, Timothy W.; Inghirami, Giorgio

    2013-01-01

    Anaplastic large-cell lymphomas (ALCLs) encompass at least 2 systemic diseases distinguished by the presence or absence of anaplastic lymphoma kinase (ALK) expression. We performed genome-wide microRNA (miRNA) profiling on 33 ALK-positive (ALK[+]) ALCLs, 25 ALK-negative (ALK[−]) ALCLs, 9 angioimmunoblastic T-cell lymphomas, 11 peripheral T-cell lymphomas not otherwise specified (PTCLNOS), and normal T cells, and demonstrated that ALCLs express many of the miRNAs that are highly expressed in normal T cells with the prominent exception of miR-146a. Unsupervised hierarchical clustering demonstrated distinct clustering of ALCL, PTCL-NOS, and the AITL subtype of PTCL. Cases of ALK(+) ALCL and ALK(–) ALCL were interspersed in unsupervised analysis, suggesting a close relationship at the molecular level. We identified an miRNA signature of 7 miRNAs (5 upregulated: miR-512-3p, miR-886-5p, miR-886-3p, miR-708, miR-135b; 2 downregulated: miR-146a, miR-155) significantly associated with ALK(+) ALCL cases. In addition, we derived an 11-miRNA signature (4 upregulated: miR-210, miR-197, miR-191, miR-512-3p; 7 downregulated: miR-451, miR-146a, miR-22, miR-455-3p, miR-455-5p, miR-143, miR-494) that differentiates ALK(–) ALCL from other PTCLs. Our in vitro studies identified a set of 32 miRNAs associated with ALK expression. Of these, the miR-17∼92 cluster and its paralogues were also highly expressed in ALK(+) ALCL and may represent important downstream effectors of the ALK oncogenic pathway. PMID:23801630

  11. miRBase: integrating microRNA annotation and deep-sequencing data.

    PubMed

    Kozomara, Ana; Griffiths-Jones, Sam

    2011-01-01

    miRBase is the primary online repository for all microRNA sequences and annotation. The current release (miRBase 16) contains over 15,000 microRNA gene loci in over 140 species, and over 17,000 distinct mature microRNA sequences. Deep-sequencing technologies have delivered a sharp rise in the rate of novel microRNA discovery. We have mapped reads from short RNA deep-sequencing experiments to microRNAs in miRBase and developed web interfaces to view these mappings. The user can view all read data associated with a given microRNA annotation, filter reads by experiment and count, and search for microRNAs by tissue- and stage-specific expression. These data can be used as a proxy for relative expression levels of microRNA sequences, provide detailed evidence for microRNA annotations and alternative isoforms of mature microRNAs, and allow us to revisit previous annotations. miRBase is available online at: http://www.mirbase.org/. PMID:21037258

  12. mRNA and microRNA expression profiles of the NCI-60 integrated with drug activities

    PubMed Central

    Liu, Hongfang; D’Andrade, Petula; Fulmer-Smentek, Stephanie; Lorenzi, Philip; Kohn, Kurt W.; Weinstein, John N.; Pommier, Yves; Reinhold, William C.

    2010-01-01

    As part of the Spotlight on Molecular Profiling series, we present here new profiling studies of mRNA and microRNA expression for the 60 cell lines of the NCI DTP drug screen (NCI-60) using the 41,000-probe Agilent Whole Human Genome Oligo Microarray and the 15,000-feature Agilent Human microRNA Microarray V2. The expression levels of ~21,000 genes and 723 human microRNAs were measured. These profiling studies include quadruplicate technical replicates for six and eight cell lines for mRNA and microRNA, respectively, and duplicates for the remaining cell lines. The resulting data sets are freely available and searchable online in our CellMiner database. The result indicates high reproducibility for both platforms and an essential biological similarity across the various cell types. The mRNA and microRNA expression levels were integrated with our previously published 1,429-compound database of anticancer activity obtained from the NCI DTP drug screen. Large blocks of both mRNAs and microRNAs were identified with predominately unidirectional correlations to ~1,300 drugs including 121 drugs with known mechanisms of action. The data sets presented here will facilitate the identification of groups of mRNAs, microRNAs and drugs that potentially affect and interact with one another. PMID:20442302

  13. MicroRNA and Pathogenesis of Enterovirus Infection

    PubMed Central

    Ho, Bing-Ching; Yang, Pan-Chyr; Yu, Sung-Liang

    2016-01-01

    There are no currently available specific antiviral therapies for non-polio Enterovirus infections. Although several vaccines have entered clinical trials, the efficacy requires further evaluation, particularly for cross-strain protective activity. Curing patients with viral infections is a public health problem due to antigen alterations and drug resistance caused by the high genomic mutation rate. To conquer these limits in the development of anti-Enterovirus treatments, a comprehensive understanding of the interactions between Enterovirus and host cells is urgently needed. MicroRNA (miRNA) constitutes the biggest family of gene regulators in mammalian cells and regulates almost a half of all human genes. The roles of miRNAs in Enterovirus pathogenesis have recently begun to be noted. In this review, we shed light on recent advances in the understanding of Enterovirus infection-modulated miRNAs. The impacts of altered host miRNAs on cellular processes, including immune escape, apoptosis, signal transduction, shutdown of host protein synthesis and viral replication, are discussed. Finally, miRNA-based medication provides a promising strategy for the development of antiviral therapy. PMID:26751468

  14. Direct serum assay for microRNA in cancer patients.

    PubMed

    Asaga, Sota; Hoon, Dave S B

    2013-01-01

    MicroRNAs (miRs) are small noncoding RNAs which can be detected in body fluids as well as cells and tissues. miRs play important roles in various activities of cancer cells. The miRs in bloods called circulating miRs (cmiRs) are potential biomarkers for detecting and monitoring cancer progression. There are reports on the cmiR research which utilizes various primers, reagents, and instruments. Here, we introduce our protocols for RNA extraction and RT-qPCR for cmiRs as well as our novel RT-qPCR directly in serum assay (RT-qPCR-DS) where RT is directly performed in serum without the need for RNA extraction. Results from the two protocols are analyzed and compared. RT-qPCR-DS is logistically simpler and more sensitive in assessing cmiR in breast cancer patients than isolating RNA before RT-qPCR. This approach may be very useful as a diagnostic tool. PMID:23719948

  15. Circulating microRNA-based screening tool for breast cancer

    PubMed Central

    Boukerroucha, Meriem; Fasquelle, Corinne; Thiry, Jérôme; Bovy, Nicolas; Struman, Ingrid; Geurts, Pierre; Collignon, Joëlle; Schroeder, Hélène; Kridelka, Frédéric; Lifrange, Eric; Jossa, Véronique

    2016-01-01

    Circulating microRNAs (miRNAs) are increasingly recognized as powerful biomarkers in several pathologies, including breast cancer. Here, their plasmatic levels were measured to be used as an alternative screening procedure to mammography for breast cancer diagnosis. A plasma miRNA profile was determined by RT-qPCR in a cohort of 378 women. A diagnostic model was designed based on the expression of 8 miRNAs measured first in a profiling cohort composed of 41 primary breast cancers and 45 controls, and further validated in diverse cohorts composed of 108 primary breast cancers, 88 controls, 35 breast cancers in remission, 31 metastatic breast cancers and 30 gynecologic tumors. A receiver operating characteristic curve derived from the 8-miRNA random forest based diagnostic tool exhibited an area under the curve of 0.81. The accuracy of the diagnostic tool remained unchanged considering age and tumor stage. The miRNA signature correctly identified patients with metastatic breast cancer. The use of the classification model on cohorts of patients with breast cancers in remission and with gynecologic cancers yielded prediction distributions similar to that of the control group. Using a multivariate supervised learning method and a set of 8 circulating miRNAs, we designed an accurate, minimally invasive screening tool for breast cancer. PMID:26734993

  16. MicroRNA in prostate cancer: Practical aspects.

    PubMed

    Patil, Pallavi A; Magi-Galluzzi, Cristina

    2015-12-01

    In the last decade, microRNAs (miRNAs) have emerged as biomarkers for cancer diagnosis, prognosis, therapy and prediction of treatment response and have earned a promising role in prostate cancer (PCa) management. A plethora of studies has been conducted on miRNA expression in PCa compared to non-neoplastic prostatic tissue, in PCa of different histologic grades and pathologic stages, in castration resistance prostate cancer (CRPC), in metastatic disease and in response to therapy, with evidence pointing towards distinctive miRNAs differentially expressed in each of these phases. In addition to tissue, miRNA can be detected in blood, serum, and urine. The aim of this review is to survey studies conducted on human prostate tissue and biofluids and to consolidate trustworthy data on the role of miRNA in the occurrence and progression of PCa, with a delineation of differentially expressed miRNAs and an analysis of their association with PCa prognosis, progression to CRPC and metastatic disease, as well as their correlation with response to chemotherapy and hormonal therapy. Changes in circulating miRNAs may represent potentially useful non-invasive biomarkers for PCa diagnosis, staging and prediction of outcome. PMID:26186079

  17. A microRNA isolation method from clinical samples

    PubMed Central

    Zununi Vahed, Sepideh; Barzegari, Abolfazl; Rahbar Saadat, Yalda; Mohammadi, Somayeh; Samadi, Nasser

    2016-01-01

    Introduction: microRNAs (miRNAs) are considered to be novel molecular biomakers that could be exploited in the diagnosis and treatment of different diseases. The present study aimed to develop an efficient miRNA isolation method from different clinical specimens. Methods: Total RNAs were isolated by Trizol reagent followed by precipitation of the large RNAs with potassium acetate (KCH3COOH), polyethylene glycol (PEG) 4000 and 6000, and lithium chloride (LiCl). Then, small RNAs were enriched and recovered from the supernatants by applying a combination of LiCl and ethanol. The efficiency of the method was evaluated through the quality, quantity, and integrity of the recovered RNAs using the A260/280 absorbance ratio, reverse transcription PCR (RT-PCR), and quantitative real-time PCR (q-PCR). Results: Comparison of different RNA isolation methods based on the precipitation of DNA and large RNAs, high miRNA recovery and PCR efficiency revealed that applying potassium acetate with final precipitation of small RNAs using 2.5 M LiCl plus ethanol can provide high yield and quality small RNAs that can be exploited for clinical purposes. Conclusion: The current isolation method can be applied for most clinical samples including cells, formalin-fixed and paraffin-embedded (FFPE) tissues and even body fluids with a wide applicability in molecular biology investigations. PMID:27340621

  18. Circulating microRNA Signatures in Rodent Models of Pain.

    PubMed

    Qureshi, Rehman A; Tian, Yuzhen; McDonald, Marguerite K; Capasso, Kathryn E; Douglas, Sabrina R; Gao, Ruby; Orlova, Irina A; Barrett, James E; Ajit, Seena K; Sacan, Ahmet

    2016-07-01

    MicroRNAs (miRNAs) remain stable in circulation and have been identified as potential biomarkers for a variety of conditions. We report miRNA changes in blood from multiple rodent models of pain, including spinal nerve ligation and spared nerve injury models of neuropathic pain; a complete Freund's adjuvant (CFA) model of inflammatory pain; and a chemotherapy-induced model of pain using the histone deacetylase inhibitor JNJ-26481585. The effect of celecoxib, a cyclooxygenase-2-selective nonsteroidal anti-inflammatory drug, was investigated in the CFA model as proof of principle for assessing the utility of circulating miRNAs as biomarkers in determining treatment response. Each study resulted in a unique miRNA expression profile. Despite differences in miRNAs identified from various models, computational target prediction and functional enrichment have identified biological pathways common among different models. The Wnt signaling pathway was affected in all models, suggesting a crucial role for this pathway in the pathogenesis of pain. Our studies demonstrate the utility of circulating miRNAs as pain biomarkers and suggest the potential for rigorous forward and reverse translational approaches. Evaluating alterations in miRNA fingerprints under different pain conditions and after administering therapeutic agents may be beneficial in evaluating clinical trial outcomes, predicting treatment response, and developing correlational outcomes between preclinical and human studies. PMID:26081151

  19. Duplicate gene divergence by changes in microRNA binding sites in Arabidopsis and Brassica.

    PubMed

    Wang, Sishuo; Adams, Keith L

    2015-03-01

    Gene duplication provides large numbers of new genes that can lead to the evolution of new functions. Duplicated genes can diverge by changes in sequences, expression patterns, and functions. MicroRNAs play an important role in the regulation of gene expression in many eukaryotes. After duplication, two paralogs may diverge in their microRNA binding sites, which might impact their expression and function. Little is known about conservation and divergence of microRNA binding sites in duplicated genes in plants. We analyzed microRNA binding sites in duplicated genes in Arabidopsis thaliana and Brassica rapa. We found that duplicates are more often targeted by microRNAs than singletons. The vast majority of duplicated genes in A. thaliana with microRNA binding sites show divergence in those sites between paralogs. Analysis of microRNA binding sites in genes derived from the ancient whole-genome triplication in B. rapa also revealed extensive divergence. Paralog pairs with divergent microRNA binding sites show more divergence in expression patterns compared with paralog pairs with the same microRNA binding sites in Arabidopsis. Close to half of the cases of binding site divergence are caused by microRNAs that are specific to the Arabidopsis genus, indicating evolutionarily recent gain of binding sites after target gene duplication. We also show rapid evolution of microRNA binding sites in a jacalin gene family. Our analyses reveal a dynamic process of changes in microRNA binding sites after gene duplication in Arabidopsis and highlight the role of microRNA regulation in the divergence and contrasting evolutionary fates of duplicated genes. PMID:25644246

  20. MicroRNA and tasiRNA diversity in mature pollen of Arabidopsis thaliana

    PubMed Central

    2009-01-01

    Background New generation sequencing technology has allowed investigation of the small RNA populations of flowering plants at great depth. However, little is known about small RNAs in their reproductive cells, especially in post-meiotic cells of the gametophyte generation. Pollen - the male gametophyte - is the specialised haploid structure that generates and delivers the sperm cells to the female gametes at fertilisation. Whether development and differentiation of the male gametophyte depends on the action of microRNAs and trans-acting siRNAs guiding changes in gene expression is largely unknown. Here we have used 454 sequencing to survey the various small RNA populations present in mature pollen of Arabidopsis thaliana. Results In this study we detected the presence of 33 different microRNA families in mature pollen and validated the expression levels of 17 selected miRNAs by Q-RT-PCR. The majority of the selected miRNAs showed pollen-enriched expression compared with leaves. Furthermore, we report for the first time the presence of trans-acting siRNAs in pollen. In addition to describing new patterns of expression for known small RNAs in each of these classes, we identified 7 putative novel microRNAs. One of these, ath-MIR2939, targets a pollen-specific F-box transcript and we demonstrate cleavage of its target mRNA in mature pollen. Conclusions Despite the apparent simplicity of the male gametophyte, comprising just two different cell types, pollen not only utilises many miRNAs and trans-acting siRNAs expressed in the somatic tissues but also expresses novel miRNAs. PMID:20042113

  1. Human cerebrospinal fluid microRNA: temporal changes following subarachnoid hemorrhage.

    PubMed

    Powers, Ciarán J; Dickerson, Ryan; Zhang, Stacey W; Rink, Cameron; Roy, Sashwati; Sen, Chandan K

    2016-05-01

    Aneurysmal subarachnoid hemorrhage (aSAH) is a devastating form of hemorrhagic stroke with 30-day mortality between 33 and 45%. Delayed cerebral ischemia (DCI) is the chief cause of morbidity and mortality in patients who survive the initial aSAH. DCI accounts for almost 50% of deaths in patients surviving to treatment of the ruptured aneurysm. The mechanisms for brain injury after aSAH and the brain's response to this injury are not fully understood in humans. MicroRNAs (miRs) are 22- to 25-nucleotide single-stranded RNA molecules that inhibit the expression of specific messenger RNA targets. In this work, miR profiling of human cerebrospinal fluid from eight patients after aSAH was performed daily for 10 days with the goal of identifying changes in miR abundance. Using the nanoString nCounter Expression Assay, we identified two specific clusters of miR that were differentially regulated over time. Quantitative RT-PCR was performed on select miRs from each cluster. The first cluster contained miRs known to be present in blood and decreased in abundance over time. miRs in this group include miR-92a and let-7b. The second cluster contained several poorly characterized miRs that increased in abundance over time. miRs in this group included miR-491. This second cluster of miRs may be released into the CSF by the brain itself as a result of the initial SAH. Temporal changes in the abundance of specific miRs in human CSF after aSAH may provide novel insight into the role of miRs in brain injury and the brain's response. PMID:26945012

  2. Identifying significant microRNA-mRNA pairs associated with breast cancer subtypes.

    PubMed

    Bhattacharyya, Malay; Nath, Joyshree; Bandyopadhyay, Sanghamitra

    2016-07-01

    MicroRNAs (miRNAs) are small non-coding RNAs that help in post-transcriptional gene silencing. These endogenous RNAs develop a post-transcriptional gene-regulatory network by binding to complementary sequences of target mRNAs and essentially degrade them. Cancer is a class of diseases that is caused by the uncontrolled cell growth, thereby resulting into a gradual degradation of cell structure. Earlier researches have shown that miRNAs have significant biological involvement in cancer. Prolonged research in this genre has led to the identification of the functions of numerous miRNAs in cancer development. Studying the differential expression profiles of miRNAs and mRNAs together could help us in recognizing the significant miRNA-mRNA pairs from cancer samples. In this paper, we have analyzed the simultaneous over-expression of miRNAs and under-expression of mRNAs and vice versa to establish their association with cancer. This study focuses on breast tumor samples and the miRNA-mRNA target pairs that have a visible signature in such breast tumor samples. We have been able to identify the differentially expressed miRNAs and mRNAs, and further established relations between them to extract the miRNA-mRNA pairs that might be significant in the breast cancer types. This gives us the clue about the potential biomarkers for the breast cancer subtypes that can further help in understanding the progression of each of the subtypes separately. This might be helpful for the joint miRNA-mRNA biomarker identification. PMID:27245063

  3. GE-01MOLECULAR AND PATHOLOGIC SUBSETS OF LOW GRADE GLIOMAS AND GLIONEURONAL TUMORS IDENTIFIED BY microRNA PROFILING

    PubMed Central

    Ames, Heather; Vizcaino, M. Adelita; Rodriguez, Fausto

    2014-01-01

    Low-grade (WHO I-II) gliomas represent the most frequent primary tumors of the central nervous system in children. They often have a good prognosis following total resection, however they can create many neurological complications due to mass effect, and may be difficult to resect depending on anatomic location. MicroRNAs have been identified as molecular regulators of protein expression that can repress multiple mRNAs concurrently through base pairing. Specific microRNAs are often suppressed during early cell differentiation to promote the expression of mitogenic proteins that are associated with the maintenance of specific stem cell types, a mechanism for growth and survival that is frequently exploited in cancer cells. Identification of these microRNA signatures present in low grade glioma and glioneuronal tumor sub-types could therefore lead to a wealth of candidate biomarkers. We used NanoString technology to analyze the expression levels of 800 microRNAs in nine low-grade glial and glioneuronal tumor subtypes (n = 45) using formalin-fixed paraffin-embedded tissue. We then generated hierarchical clusters following evaluation via significant analysis of microarrays (SAMs). Hierarchical clustering separated tumors from non-neoplastic brain. When looking at individual tumors, subependymal giant cell astrocytomas (SEGA) clustered sharply together, consistent with a unique microRNA expression signature in this tuberous sclerosis associated tumor subtype, compared to other low grade glial and glioneuronal tumors. Candidate microRNAs were validated using qRT-PCR. In SEGAs, microRNAs miR-219-5p, miR-129-2-3p, miR-338-3p, miR-487b, miR-885-5p, and miR-323-3p were significantly down-regulated by more than 15 fold as compared to normal brain and were also significantly down-regulated as compared to other low grade gliomas. In summary, altered microRNA expression is a feature of low grade glial and glioneuronal tumors. MicroRNA profiling may therefore be useful in

  4. MicroRNA expression in ileal carcinoid tumors: downregulation of microRNA-133a with tumor progression.

    PubMed

    Ruebel, Katharina; Leontovich, Alexey A; Stilling, Gail A; Zhang, Shuya; Righi, Alberto; Jin, Long; Lloyd, Ricardo V

    2010-03-01

    MicroRNAs (miRNAs) are involved in cell proliferation, differentiation, and apoptosis and can function as tumor suppressor genes or oncogenes. The role of miRNAs in neuroendocrine tumors such as ileal carcinoids is largely unknown. We examined the differential expression of 95 miRNAs by RT-PCR using the QuantiMir System in eight matching primary and metastatic carcinoid tumors from the ileum. All miRNAs chosen for the QuantiMir System array were based on their potential functions related to cancer biology, cell development, and apoptosis. The expression of miRNAs for the samples was normalized to miRNA-197, and the matching primary and metastatic tumors were compared. There was downregulation of miRNA-133a, -145, -146, -222, and -10b in all samples between the primary and matching metastatic tumors and upregulation of miRNA-183, -488, and -19a+b in six of eight metastatic carcinoids compared to the primary tumors. miRNA-133a was further analyzed by TaqMan real-time RT-PCR and northern hybridization using six additional matching primary and metastatic samples, which supported the PCR array findings. There were significant differences in miRNA-133a expression with downregulation in the metastasis compared to the primary in the eight original cases (P<0.009) and in the six additional cases used for validation (P<0.014). Laser capture microdissection and real-time RT-PCR analysis using normal ileum found miRNA-133a expression in normal enterochromaffin cells. In situ hybridization in normal ileum showed that some of the mucosal endocrine cells expressed miRNA-133a. Both primary and metastatic ileal carcinoid tumors expressed miRNA-133a by in situ hybridization. These results provide information about novel marker miRNAs that may be used as biomarkers and/or therapeutic targets in intestinal carcinoid tumors. PMID:20037573

  5. Structural features of microRNA (miRNA) precursors and their relevance to miRNA biogenesis and small interfering RNA/short hairpin RNA design.

    PubMed

    Krol, Jacek; Sobczak, Krzysztof; Wilczynska, Urszula; Drath, Maria; Jasinska, Anna; Kaczynska, Danuta; Krzyzosiak, Wlodzimierz J

    2004-10-01

    We have established the structures of 10 human microRNA (miRNA) precursors using biochemical methods. Eight of these structures turned out to be different from those that were computer-predicted. The differences localized in the terminal loop region and at the opposite side of the precursor hairpin stem. We have analyzed the features of these structures from the perspectives of miRNA biogenesis and active strand selection. We demonstrated the different thermodynamic stability profiles for pre-miRNA hairpins harboring miRNAs at their 5'- and 3'-sides and discussed their functional implications. Our results showed that miRNA prediction based on predicted precursor structures may give ambiguous results, and the success rate is significantly higher for the experimentally determined structures. On the other hand, the differences between the predicted and experimentally determined structures did not affect the stability of termini produced through "conceptual dicing." This result confirms the value of thermodynamic analysis based on mfold as a predictor of strand section by RNAi-induced silencing complex (RISC). PMID:15292246

  6. Dietary MicroRNA Database (DMD): An Archive Database and Analytic Tool for Food-Borne microRNAs

    PubMed Central

    Chiang, Kevin; Shu, Jiang; Zempleni, Janos; Cui, Juan

    2015-01-01

    With the advent of high throughput technology, a huge amount of microRNA information has been added to the growing body of knowledge for non-coding RNAs. Here we present the Dietary MicroRNA Databases (DMD), the first repository for archiving and analyzing the published and novel microRNAs discovered in dietary resources. Currently there are fifteen types of dietary species, such as apple, grape, cow milk, and cow fat, included in the database originating from 9 plant and 5 animal species. Annotation for each entry, a mature microRNA indexed as DM0000*, covers information of the mature sequences, genome locations, hairpin structures of parental pre-microRNAs, cross-species sequence comparison, disease relevance, and the experimentally validated gene targets. Furthermore, a few functional analyses including target prediction, pathway enrichment and gene network construction have been integrated into the system, which enable users to generate functional insights through viewing the functional pathways and building protein-protein interaction networks associated with each microRNA. Another unique feature of DMD is that it provides a feature generator where a total of 411 descriptive attributes can be calculated for any given microRNAs based on their sequences and structures. DMD would be particularly useful for research groups studying microRNA regulation from a nutrition point of view. The database can be accessed at http://sbbi.unl.edu/dmd/. PMID:26030752

  7. Dietary MicroRNA Database (DMD): An Archive Database and Analytic Tool for Food-Borne microRNAs.

    PubMed

    Chiang, Kevin; Shu, Jiang; Zempleni, Janos; Cui, Juan

    2015-01-01

    With the advent of high throughput technology, a huge amount of microRNA information has been added to the growing body of knowledge for non-coding RNAs. Here we present the Dietary MicroRNA Databases (DMD), the first repository for archiving and analyzing the published and novel microRNAs discovered in dietary resources. Currently there are fifteen types of dietary species, such as apple, grape, cow milk, and cow fat, included in the database originating from 9 plant and 5 animal species. Annotation for each entry, a mature microRNA indexed as DM0000*, covers information of the mature sequences, genome locations, hairpin structures of parental pre-microRNAs, cross-species sequence comparison, disease relevance, and the experimentally validated gene targets. Furthermore, a few functional analyses including target prediction, pathway enrichment and gene network construction have been integrated into the system, which enable users to generate functional insights through viewing the functional pathways and building protein-protein interaction networks associated with each microRNA. Another unique feature of DMD is that it provides a feature generator where a total of 411 descriptive attributes can be calculated for any given microRNAs based on their sequences and structures. DMD would be particularly useful for research groups studying microRNA regulation from a nutrition point of view. The database can be accessed at http://sbbi.unl.edu/dmd/. PMID:26030752

  8. MicroRNA control of myelopoiesis and the differentiation block in acute myeloid leukaemia

    PubMed Central

    Palma, Catalina A; Tonna, Elise J; Ma, David F; Lutherborrow, Mark A

    2012-01-01

    Abstract In the relatively short period of time since their discovery, microRNAs have been shown to control many important cellular functions such as cell differentiation, growth, proliferation and apoptosis. In addition, microRNAs have been demonstrated as key drivers of many malignancies and can function as either tumour suppressors or oncogenes. The haematopoietic system is not outside the realm of microRNA control with microRNAs controlling aspects of stem cell and progenitor self-renewal and differentiation, with many, if not all, haematological disorders associated with aberrant microRNA expression and function. In this review, we focus on the current understanding of microRNA control of haematopoiesis and detail the evidence for the contribution and clinical relevance of aberrant microRNA function to the characteristic block of differentiation in acute myeloid leukaemia. PMID:22225649

  9. An Improved microRNA Annotation of the Canine Genome.

    PubMed

    Penso-Dolfin, Luca; Swofford, Ross; Johnson, Jeremy; Alföldi, Jessica; Lindblad-Toh, Kerstin; Swarbreck, David; Moxon, Simon; Di Palma, Federica

    2016-01-01

    The domestic dog, Canis familiaris, is a valuable model for studying human diseases. The publication of the latest Canine genome build and annotation, CanFam3.1 provides an opportunity to enhance our understanding of gene regulation across tissues in the dog model system. In this study, we used the latest dog genome assembly and small RNA sequencing data from 9 different dog tissues to predict novel miRNAs in the dog genome, as well as to annotate conserved miRNAs from the miRBase database that were missing from the current dog annotation. We used both miRCat and miRDeep2 algorithms to computationally predict miRNA loci. The resulting, putative hairpin sequences were analysed in order to discard false positives, based on predicted secondary structures and patterns of small RNA read alignments. Results were further divided into high and low confidence miRNAs, using the same criteria. We generated tissue specific expression profiles for the resulting set of 811 loci: 720 conserved miRNAs, (207 of which had not been previously annotated in the dog genome) and 91 novel miRNA loci. Comparative analyses revealed 8 putative homologues of some novel miRNA in ferret, and one in microbat. All miRNAs were also classified into the genic and intergenic categories, based on the Ensembl RefSeq gene annotation for CanFam3.1. This additionally allowed us to identify four previously undescribed MiRtrons among our total set of miRNAs. We additionally annotated piRNAs, using proTRAC on the same input data. We thus identified 263 putative clusters, most of which (211 clusters) were found to be expressed in testis. Our results represent an important improvement of the dog genome annotation, paving the way to further research on the evolution of gene regulation, as well as on the contribution of post-transcriptional regulation to pathological conditions. PMID:27119849

  10. An Improved microRNA Annotation of the Canine Genome

    PubMed Central

    Swofford, Ross; Johnson, Jeremy; Alföldi, Jessica; Lindblad-Toh, Kerstin; Swarbreck, David; Moxon, Simon; Di Palma, Federica

    2016-01-01

    The domestic dog, Canis familiaris, is a valuable model for studying human diseases. The publication of the latest Canine genome build and annotation, CanFam3.1 provides an opportunity to enhance our understanding of gene regulation across tissues in the dog model system. In this study, we used the latest dog genome assembly and small RNA sequencing data from 9 different dog tissues to predict novel miRNAs in the dog genome, as well as to annotate conserved miRNAs from the miRBase database that were missing from the current dog annotation. We used both miRCat and miRDeep2 algorithms to computationally predict miRNA loci. The resulting, putative hairpin sequences were analysed in order to discard false positives, based on predicted secondary structures and patterns of small RNA read alignments. Results were further divided into high and low confidence miRNAs, using the same criteria. We generated tissue specific expression profiles for the resulting set of 811 loci: 720 conserved miRNAs, (207 of which had not been previously annotated in the dog genome) and 91 novel miRNA loci. Comparative analyses revealed 8 putative homologues of some novel miRNA in ferret, and one in microbat. All miRNAs were also classified into the genic and intergenic categories, based on the Ensembl RefSeq gene annotation for CanFam3.1. This additionally allowed us to identify four previously undescribed MiRtrons among our total set of miRNAs. We additionally annotated piRNAs, using proTRAC on the same input data. We thus identified 263 putative clusters, most of which (211 clusters) were found to be expressed in testis. Our results represent an important improvement of the dog genome annotation, paving the way to further research on the evolution of gene regulation, as well as on the contribution of post-transcriptional regulation to pathological conditions. PMID:27119849

  11. Intact MicroRNA Analysis Using High Resolution Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Kullolli, Majlinda; Knouf, Emily; Arampatzidou, Maria; Tewari, Muneesh; Pitteri, Sharon J.

    2014-01-01

    MicroRNAs (miRNAs) are small single-stranded non-coding RNAs that post-transcriptionally regulate gene expression, and play key roles in the regulation of a variety of cellular processes and in disease. New tools to analyze miRNAs will add understanding of the physiological origins and biological functions of this class of molecules. In this study, we investigate the utility of high resolution mass spectrometry for the analysis of miRNAs through proof-of-concept experiments. We demonstrate the ability of mass spectrometry to resolve and separate miRNAs and corresponding 3' variants in mixtures. The mass accuracy of the monoisotopic deprotonated peaks from various miRNAs is in the low ppm range. We compare fragmentation of miRNA by collision-induced dissociation (CID) and by higher-energy collisional dissociation (HCD) which yields similar sequence coverage from both methods but additional fragmentation by HCD versus CID. We measure the linear dynamic range, limit of detection, and limit of quantitation of miRNA loaded onto a C18 column. Lastly, we explore the use of data-dependent acquisition of MS/MS spectra of miRNA during online LC-MS and demonstrate that multiple charge states can be fragmented, yielding nearly full sequence coverage of miRNA on a chromatographic time scale. We conclude that high resolution mass spectrometry allows the separation and measurement of miRNAs in mixtures and a standard LC-MS setup can be adapted for online analysis of these molecules.

  12. MicroRNA regulation and dysregulation in epilepsy

    PubMed Central

    Dogini, Danyella B.; Avansini, Simoni H.; Vieira, Andre S.; Lopes-Cendes, Iscia

    2013-01-01

    Epilepsy, one of the most frequent neurological disorders, represents a group of diseases that have in common the clinical occurrence of seizures. The pathogenesis of different types of epilepsy involves many important biological pathways; some of which have been shown to be regulated by microRNAs (miRNAs). In this paper, we will critically review relevant studies regarding the role of miRNAs in epilepsy. Overall, the most common type of epilepsy in the adult population is temporal lobe epilepsy (TLE), and the form associated with mesial temporal sclerosis (MTS), called mesial TLE, is particularly relevant due to the high frequency of resistance to clinical treatment. There are several target studies, as well few genome-wide miRNA expression profiling studies reporting abnormal miRNA expression in tissue with MTS, both in patients and in animal models. Overall, these studies show a fine correlation between miRNA regulation/dysregulation and inflammation, seizure-induced neuronal death and other relevant biological pathways. Furthermore, expression of many miRNAs is dynamically regulated during neurogenesis and its dysregulation may play a role in the process of cerebral corticogenesis leading to malformations of cortical development (MCD), which represent one of the major causes of drug-resistant epilepsy. In addition, there are reports of miRNAs involved in cell proliferation, fate specification, and neuronal maturation and these processes are tightly linked to the pathogenesis of MCD. Large-scale analyzes of miRNA expression in animal models with induced status epilepticus have demonstrated changes in a selected group of miRNAs thought to be involved in the regulation of cell death, synaptic reorganization, neuroinflammation, and neural excitability. In addition, knocking-down specific miRNAs in these animals have demonstrated that this may consist in a promising therapeutic intervention. PMID:24109432

  13. Functions of microRNA in response to cocaine stimulation.

    PubMed

    Xu, L-F; Wang, J; Lv, F B; Song, Q

    2013-01-01

    MicroRNAs (miRNAs) are a type of non-protein-coding single-stranded RNA, which are typically 20-25 nt in length. miRNAs play important roles in various biological processes, including development, cell proliferation, differentiation, and apoptosis. We aimed to detect the miRNA response to cocaine stimulations and their target genes. Using the miRNA expression data GSE21901 downloaded from the Gene Expression Omnibus database, we screened out the differentially expressed miRNA after short-term (1 h) and longer-term (6 h) cocaine stimulations based on the fold change >1.2. Target genes of differentially expressed miRNAs were retrieved from TargetScan database with the context score -0.3. Functional annotation enrichment analysis was performed for all the target genes with DAVID. A total of 121 differentially expressed miRNAs between the 1-h treatment and the control samples, 58 between the 6-h treatment and the control samples, and 69 between the 1-h and the 6-h treatment samples. Among them, miR-212 results of particular interest, since its expression level was constantly elevated responding to cocaine treatment. After functional and pathway annotations of target genes, we proved that miR-212 was a critical element in cocaine-addiction, because of its involvement in regulating several important cell cycle events. The results may pave the way for further understanding the regulatory mechanisms of cocaine-response in human bodies. PMID:24338410

  14. Arabidopsis microRNA expression regulation in a wide range of abiotic stress responses

    PubMed Central

    Barciszewska-Pacak, Maria; Milanowska, Kaja; Knop, Katarzyna; Bielewicz, Dawid; Nuc, Przemyslaw; Plewka, Patrycja; Pacak, Andrzej M.; Vazquez, Franck; Karlowski, Wojciech; Jarmolowski, Artur; Szweykowska-Kulinska, Zofia

    2015-01-01

    Arabidopsis microRNA expression regulation was studied in a wide array of abiotic stresses such as drought, heat, salinity, copper excess/deficiency, cadmium excess, and sulfur deficiency. A home-built RT-qPCR mirEX platform for the amplification of 289 Arabidopsis microRNA transcripts was used to study their response to abiotic stresses. Small RNA sequencing, Northern hybridization, and TaqMan® microRNA assays were performed to study the abundance of mature microRNAs. A broad response on the level of primary miRNAs (pri-miRNAs) was observed. However, stress response at the level of mature microRNAs was rather confined. The data presented show that in most instances, the level of a particular mature miRNA could not be predicted based on the level of its pri-miRNA. This points to an essential role of posttranscriptional regulation of microRNA expression. New Arabidopsis microRNAs responsive to abiotic stresses were discovered. Four microRNAs: miR319a/b, miR319b.2, and miR400 have been found to be responsive to several abiotic stresses and thus can be regarded as general stress-responsive microRNA species. PMID:26089831

  15. MicroRNA 140 Promotes Expression of Long Noncoding RNA NEAT1 in Adipogenesis.

    PubMed

    Gernapudi, Ramkishore; Wolfson, Benjamin; Zhang, Yongshu; Yao, Yuan; Yang, Peixin; Asahara, Hiroshi; Zhou, Qun

    2016-01-01

    More than 40% of the U.S. population are clinically obese and suffer from metabolic syndrome with an increased risk of postmenopausal estrogen receptor-positive breast cancer. Adipocytes are the primary component of adipose tissue and are formed through adipogenesis from precursor mesenchymal stem cells. While the major molecular pathways of adipogenesis are understood, little is known about the noncoding RNA signaling networks involved in adipogenesis. Using adipocyte-derived stem cells (ADSCs) isolated from wild-type and microRNA 140 (miR-140) knockout mice, we identify a novel miR-140/long noncoding RNA (lncRNA) NEAT1 signaling network necessary for adipogenesis. miR-140 knockout ADSCs have dramatically decreased adipogenic capabilities associated with downregulation of NEAT1 expression. We identified a miR-140 binding site in NEAT1 and found that mature miR-140 in the nucleus can physically interact with NEAT1, leading to increased NEAT1 expression. We demonstrated that reexpression of NEAT1 in miR-140 knockout ADSCs is sufficient to restore their ability to undergo differentiation. Our results reveal an exciting new noncoding RNA signaling network that regulates adipogenesis and that is a potential new target in the prevention or treatment of obesity. PMID:26459763

  16. MicroRNA 140 Promotes Expression of Long Noncoding RNA NEAT1 in Adipogenesis

    PubMed Central

    Gernapudi, Ramkishore; Wolfson, Benjamin; Zhang, Yongshu; Yao, Yuan; Yang, Peixin; Asahara, Hiroshi

    2015-01-01

    More than 40% of the U.S. population are clinically obese and suffer from metabolic syndrome with an increased risk of postmenopausal estrogen receptor-positive breast cancer. Adipocytes are the primary component of adipose tissue and are formed through adipogenesis from precursor mesenchymal stem cells. While the major molecular pathways of adipogenesis are understood, little is known about the noncoding RNA signaling networks involved in adipogenesis. Using adipocyte-derived stem cells (ADSCs) isolated from wild-type and microRNA 140 (miR-140) knockout mice, we identify a novel miR-140/long noncoding RNA (lncRNA) NEAT1 signaling network necessary for adipogenesis. miR-140 knockout ADSCs have dramatically decreased adipogenic capabilities associated with downregulation of NEAT1 expression. We identified a miR-140 binding site in NEAT1 and found that mature miR-140 in the nucleus can physically interact with NEAT1, leading to increased NEAT1 expression. We demonstrated that reexpression of NEAT1 in miR-140 knockout ADSCs is sufficient to restore their ability to undergo differentiation. Our results reveal an exciting new noncoding RNA signaling network that regulates adipogenesis and that is a potential new target in the prevention or treatment of obesity. PMID:26459763

  17. Carbonic Anhydrase 9 mRNA/microRNA34a Interplay in Hypoxic Human Mammospheres.

    PubMed

    De Carolis, Sabrina; Bertoni, Sara; Nati, Marina; D'Anello, Laura; Papi, Alessio; Tesei, Anna; Cricca, Monica; Bonafé, Massimiliano

    2016-07-01

    The hypoxic environment is a crucial component of the cancer stem cell niche and it is capable of eliciting stem cell features in cancer cells. We previously reported that SNAI2 up-regulates the expression of Carbonic Anhydrase iso-enzyme 9 (CA9) in hypoxic MCF7 cells. Here we show that SNAI2 down-regulates miR34a expression in hypoxic MCF7 cell-derived mammospheres. Next, we report on the capability of miR34a to decrease CA9 mRNA stability and CA9 protein expression. We also convey that the over-expression of cloned CA9-mRNA-3'UTR increases the mRNA half-life and protein levels of two miR34a targets JAGGED1 and NOTCH3. The data here reported shows that the SNAI2-dependent down-regulation of miR34a substantially contributes to the post-transcriptional up-regulation of CA9, and that CA9-mRNA-3'UTR acts as an endogenous microRNA sponge. We conclude that CA9/miR34 interplay shares in the hypoxic regulation of mammospheres and therefore, may play a relevant role in the hypoxic breast cancer stem cell niche. PMID:26553365

  18. mirConnX: condition-specific mRNA-microRNA network integrator.

    PubMed

    Huang, Grace T; Athanassiou, Charalambos; Benos, Panayiotis V

    2011-07-01

    mirConnX is a user-friendly web interface for inferring, displaying and parsing mRNA and microRNA (miRNA) gene regulatory networks. mirConnX combines sequence information with gene expression data analysis to create a disease-specific, genome-wide regulatory network. A prior, static network has been constructed for all human and mouse genes. It consists of computationally predicted transcription factor (TF)-gene associations and miRNA target predictions. The prior network is supplemented with known interactions from the literature. Dynamic TF- and miRNA-gene associations are inferred from user-provided expression data using an association measure of choice. The static and dynamic networks are then combined using an integration function with user-specified weights. Visualization of the network and subsequent analysis are provided via a very responsive graphic user interface. Two organisms are currently supported: Homo sapiens and Mus musculus. The intuitive user interface and large database make mirConnX a useful tool for clinical scientists for hypothesis generation and explorations. mirConnX is freely available for academic use at http://www.benoslab.pitt.edu/mirconnx. PMID:21558324

  19. MicroRNA Gene Expression Signature Driven by miR-9 Overexpression in Ovarian Clear Cell Carcinoma.

    PubMed

    Yanaihara, Nozomu; Noguchi, Yukiko; Saito, Misato; Takenaka, Masataka; Takakura, Satoshi; Yamada, Kyosuke; Okamoto, Aikou

    2016-01-01

    Previous studies have identified microRNA (miRNA) involvement in human cancers. This study aimed to elucidate potential clinical and biological associations of ovarian cancer-related miRNA gene expression profiles in high-grade serous carcinoma (HGSC) and ovarian clear cell carcinoma (OCCC). Accordingly, we investigated 27 patients with ovarian cancer (12 HGSC and 15 OCCC cases) using quantitative real-time reverse transcription polymerase chain reaction to determine the cancer-related miRNA expressions. Gene Cluster 3.0 was used for hierarchical clustering analysis, and differentially expressed miRNAs between HGSC and OCCC were identified by the class comparison analysis using BRB-ArrayTools. An unsupervised hierarchical clustering analysis identified two distinct miRNA expression clusters, with histological subtype-related significant differences in the associations between clusters and clinicopathological features. A comparison of miRNA expression in HGSCs and OCCCs identified five miRNAs (miR-132, miR-9, miR-126, miR-34a, and miR-21), with OCCCs demonstrating a statistically higher expression. Further investigation of the biological significance of miR-9 overexpression in OCCC revealed that miR-9 inhibition reduced the cell invasion ability and upregulated E-cadherin expression. Using a luciferase reporter assay, we further demonstrated the direct binding of miR-9 to E-cadherin. Global cancer-related miRNA expression analysis identified statistically unique profiles that could discriminate ovarian cancer histotypes. In OCCC, miR-9 overexpression may affect pathogenesis by targeting E-cadherin, thereby inducing an epithelial-mesenchymal transition. Therefore, miR-9 may be a promising therapeutic target strategy for OCCC. PMID:27612152

  20. Identification of cytokine-induced modulation of microRNA expression and secretion as measured by a novel microRNA specific qPCR assay.

    PubMed

    Benes, Vladimir; Collier, Paul; Kordes, Claus; Stolte, Jens; Rausch, Tobias; Muckentaler, Martina U; Häussinger, Dieter; Castoldi, Mirco

    2015-01-01

    microRNAs are an abundant class of small non-coding RNAs that control gene expression post-transcriptionally. Importantly, microRNA activity participates in the regulation of cellular processes and is a potentially valuable source of biomarkers in the diagnosis and prognosis of human diseases. Here we introduce miQPCR, an innovative method to quantify microRNAs expression by using Real-Time PCR. miQPCR exploits T4 RNA ligase activities to extend uniformly microRNAs' 3'-ends by addition of a linker-adapter. The adapter is then used as 'anchor' to prime cDNA synthesis and throughout qPCR to amplify specifically target amplicons. miQPCR is an open, adaptable and cost-effective procedure, which offers the following advantages; i) universal elongation and reverse transcription of all microRNAs; ii) Tm-adjustment of microRNA-specific primers; iii) high sensitivity and specificity in discriminating among closely related sequences and; iv) suitable for the analysis of cellular and cell-free circulating microRNAs. Analysis of cellular and cell-free circulating microRNAs secreted by rat primary hepatocytes stimulated with cytokines and growth factors identifies for the first time a widespread modulation of both microRNAs expression and secretion. Altogether, our findings suggest that the pleiotropic activity of humoral factors on microRNAs may extensively affect liver function in response to injury and regeneration. PMID:26108880

  1. Identification of cytokine-induced modulation of microRNA expression and secretion as measured by a novel microRNA specific qPCR assay

    PubMed Central

    Benes, Vladimir; Collier, Paul; Kordes, Claus; Stolte, Jens; Rausch, Tobias; Muckentaler, Martina U.; Häussinger, Dieter; Castoldi, Mirco

    2015-01-01

    microRNAs are an abundant class of small non-coding RNAs that control gene expression post-transcriptionally. Importantly, microRNA activity participates in the regulation of cellular processes and is a potentially valuable source of biomarkers in the diagnosis and prognosis of human diseases. Here we introduce miQPCR, an innovative method to quantify microRNAs expression by using Real-Time PCR. miQPCR exploits T4 RNA ligase activities to extend uniformly microRNAs’ 3′-ends by addition of a linker-adapter. The adapter is then used as ‘anchor’ to prime cDNA synthesis and throughout qPCR to amplify specifically target amplicons. miQPCR is an open, adaptable and cost-effective procedure, which offers the following advantages; i) universal elongation and reverse transcription of all microRNAs; ii) Tm-adjustment of microRNA-specific primers; iii) high sensitivity and specificity in discriminating among closely related sequences and; iv) suitable for the analysis of cellular and cell-free circulating microRNAs. Analysis of cellular and cell-free circulating microRNAs secreted by rat primary hepatocytes stimulated with cytokines and growth factors identifies for the first time a widespread modulation of both microRNAs expression and secretion. Altogether, our findings suggest that the pleiotropic activity of humoral factors on microRNAs may extensively affect liver function in response to injury and regeneration. PMID:26108880

  2. Role of microRNA-7 in digestive system malignancy

    PubMed Central

    Chen, Wan-Qun; Hu, Ling; Chen, Geng-Xin; Deng, Hai-Xia

    2016-01-01

    There are several malignancies of the digestive system (including gastric, pancreatic and colorectal cancers, and hepatocellular carcinoma), which are the most common types of cancer and a major cause of death worldwide. MicroRNA (miR)-7 is abundant in the pancreas, playing an important role in pancreatic development and endocrine function. Expression of miR-7 is downregulated in digestive system malignancies compared with normal tissue. Although there are contrasting results for miR-7 expression, almost all research reveals that miR-7 is a tumor suppressor, by targeting various genes in specific pathways. Moreover, miR-7 can target different genes simultaneously in different malignancies of the digestive system. By acting on many cytokines, miR-7 is also involved in many gastrointestinal inflammatory diseases as a significant carcinogenic factor. Consequently, miR-7 might be a biomarker or therapeutic target gene in digestive system malignancies. PMID:26798443

  3. MicroRNA-9 controls dendritic development by targeting REST

    PubMed Central

    Giusti, Sebastian A; Vogl, Annette M; Brockmann, Marisa M; Vercelli, Claudia A; Rein, Martin L; Trümbach, Dietrich; Wurst, Wolfgang; Cazalla, Demian; Stein, Valentin; Deussing, Jan M; Refojo, Damian

    2014-01-01

    MicroRNAs (miRNAs) are conserved noncoding RNAs that function as posttranscriptional regulators of gene expression. miR-9 is one of the most abundant miRNAs in the brain. Although the function of miR-9 has been well characterized in neural progenitors, its role in dendritic and synaptic development remains largely unknown. In order to target miR-9 in vivo, we developed a transgenic miRNA sponge mouse line allowing conditional inactivation of the miR-9 family in a spatio-temporal-controlled manner. Using this novel approach, we found that miR-9 controls dendritic growth and synaptic transmission in vivo. Furthermore, we demonstrate that miR-9-mediated downregulation of the transcriptional repressor REST is essential for proper dendritic growth. DOI: http://dx.doi.org/10.7554/eLife.02755.001 PMID:25406064

  4. MicroRNA signatures highlight new breast cancer subtypes.

    PubMed

    Bhattacharyya, Malay; Nath, Joyshree; Bandyopadhyay, Sanghamitra

    2015-02-10

    MicroRNAs (miRNAs) are a kind of short non-coding RNAs, of about 22 nucleotides in length, which modulate and sometimes degrade the target mRNAs thereby regulating a number of cellular functions. Recent research in this area establishes the involvement of miRNAs in various disease progressions, including certain types of cancer development. Further, genome-wide expression profiling of miRNAs has been proven to be useful for differentiating various cancer types. In this paper, we have used miRNA expression profiles over a large set of breast cancer tumor samples for identifying subtypes of breast cancers. The experimental results demonstrate that miRNAs carry a unique signature that distinguishes cancer subtypes and reveal new cancer subtypes. Additional survival analyses based on clinical data also strengthen this claim. PMID:25485717

  5. A microRNA code for prostate cancer metastasis

    PubMed Central

    Bonci, D; Coppola, V; Patrizii, M; Addario, A; Cannistraci, A; Francescangeli, F; Pecci, R; Muto, G; Collura, D; Bedini, R; Zeuner, A; Valtieri, M; Sentinelli, S; Benassi, M S; Gallucci, M; Carlini, P; Piccolo, S; De Maria, R

    2016-01-01

    Although the development of bone metastasis is a major detrimental event in prostate cancer, the molecular mechanisms responsible for bone homing and destruction remain largely unknown. Here we show that loss of miR-15 and miR-16 in cooperation with increased miR-21 expression promote prostate cancer spreading and bone lesions. This combination of microRNA endows bone-metastatic potential to prostate cancer cells. Concomitant loss of miR-15/miR-16 and gain of miR-21 aberrantly activate TGF-β and Hedgehog signaling, that mediate local invasion, distant bone marrow colonization and osteolysis by prostate cancer cells. These findings establish a new molecular circuitry for prostate cancer metastasis that was validated in patients' cohorts. Our data indicate a network of biomarkers and druggable pathways to improve patient treatment. PMID:26073083

  6. A Fleeting Glimpse Inside microRNA, Epigenetics, and Micropeptidomics

    PubMed Central

    2016-01-01

    MicroRNAs (miRs) are important regulators of gene expression in numerous biological processes. Their maturation process is herein described, including the most updated insights from the current literature. Circa 2000 miR sequences have been identified in the human genome, with over 50,000 miR-target interactions, including enzymes involved in epigenetic modulation of gene expression. Moreover, some “pieces of RNA” previously annotated as noncoding have been recently found to encode micropeptides that carry out critical mechanistic functions in the cell. Advanced techniques now available will certainly allow a precise scanning of the genome looking for micropeptides hidden within the “noncoding” RNA. PMID:26662983

  7. The MicroRNA-21 in Autoimmune Diseases

    PubMed Central

    Wang, Shaowen; Wan, Xiaochun; Ruan, Qingguo

    2016-01-01

    MicroRNA-21 (miR-21) is an oncomiR and significantly upregulated in a wide range of cancers. It is strongly involved in apoptosis and oncogenesis, since most of its reported targets are tumor suppressors. Recently, miR-21 was found to be correlated with the pathogenesis of autoimmune diseases and may play an essential role in regulating autoimmune responses. In particular, miR-21 promotes Th17 cell differentiation, which mediates the development of multiple autoimmune diseases. In this article, we review the current research on the mechanisms that regulate miR-21 expression, the potential of miR-21 as a diagnostic biomarker for autoimmune disease and the mechanisms by which miR-21 promotes the development of autoimmune disease. We also discussed the therapeutic potential of targeting miR-21 in treating patients with autoimmune disease. PMID:27271606

  8. MicroRNA Regulatory Networks in Cardiovascular Development

    PubMed Central

    Liu, Ning; Olson, Eric N.

    2010-01-01

    The heart, more than any other organ, requires precise function on a second-to-second basis throughout the lifespan of the organism. Even subtle perturbations in cardiac structure or function have catastrophic consequences, resulting in lethal forms of congenital and adult heart disease. Such intolerance of the heart to variability necessitates especially robust regulatory mechanisms to govern cardiac gene expression. Recent studies have revealed central roles for microRNAs (miRNAs) as governors of gene expression during cardiovascular development and disease. The integration of miRNAs into the genetic circuitry of the heart provides a rich and robust array of regulatory interactions to control cardiac gene expression. miRNA regulatory networks also offer opportunities for therapeutically modulating cardiac function through the manipulation of pathogenic and protective miRNAs. We discuss the roles of miRNAs as regulators of cardiac form and function, unresolved questions in the field, and issues for the future. PMID:20412767

  9. Involvement and Clinical Aspects of MicroRNA in Osteosarcoma.

    PubMed

    Ram Kumar, Ram Mohan; Boro, Aleksandar; Fuchs, Bruno

    2016-01-01

    Osteosarcoma (OS) is the most common primary bone cancer in children and adolescents, but its pathogenesis has been difficult to establish because of its well-known heterogeneous nature. OS has been associated with genetic and cytogenetic abnormalities, which include function-impairing mutations in tumor suppressors and the activation of oncogenes. OS tumorigenesis has been linked to alterations of several genes characterized by a high level of genetic instability and recurrent DNA amplifications and deletions. MicroRNAs (miRNAs), 18-25-nucleotide noncoding RNAs, are critical for various biological processes like differentiation, cell growth and cell death. Dysregulation of miRNA expression leads to phenotypic and genotypic changes in cells, which leads to cancer. Studies on miRNAs have initiated a significant effect in both diagnosis and treatment of cancer. This review focuses on the current knowledge of clinical applications of miRNAs for the better diagnosis and management of OS. PMID:27271607

  10. Involvement and Clinical Aspects of MicroRNA in Osteosarcoma

    PubMed Central

    Ram Kumar, Ram Mohan; Boro, Aleksandar; Fuchs, Bruno

    2016-01-01

    Osteosarcoma (OS) is the most common primary bone cancer in children and adolescents, but its pathogenesis has been difficult to establish because of its well-known heterogeneous nature. OS has been associated with genetic and cytogenetic abnormalities, which include function-impairing mutations in tumor suppressors and the activation of oncogenes. OS tumorigenesis has been linked to alterations of several genes characterized by a high level of genetic instability and recurrent DNA amplifications and deletions. MicroRNAs (miRNAs), 18–25-nucleotide noncoding RNAs, are critical for various biological processes like differentiation, cell growth and cell death. Dysregulation of miRNA expression leads to phenotypic and genotypic changes in cells, which leads to cancer. Studies on miRNAs have initiated a significant effect in both diagnosis and treatment of cancer. This review focuses on the current knowledge of clinical applications of miRNAs for the better diagnosis and management of OS. PMID:27271607

  11. MicroRNA transcriptome profiles during swine skeletal muscle development

    PubMed Central

    McDaneld, Tara G; Smith, Timothy PL; Doumit, Matthew E; Miles, Jeremy R; Coutinho, Luiz L; Sonstegard, Tad S; Matukumalli, Lakshmi K; Nonneman, Dan J; Wiedmann, Ralph T

    2009-01-01

    Background MicroRNA (miR) are a class of small RNAs that regulate gene expression by inhibiting translation of protein encoding transcripts. To evaluate the role of miR in skeletal muscle of swine, global microRNA abundance was measured at specific developmental stages including proliferating satellite cells, three stages of fetal growth, day-old neonate, and the adult. Results Twelve potential novel miR were detected that did not match previously reported sequences. In addition, a number of miR previously reported to be expressed in mammalian muscle were detected, having a variety of abundance patterns through muscle development. Muscle-specific miR-206 was nearly absent in proliferating satellite cells in culture, but was the highest abundant miR at other time points evaluated. In addition, miR-1 was moderately abundant throughout developmental stages with highest abundance in the adult. In contrast, miR-133 was moderately abundant in adult muscle and either not detectable or lowly abundant throughout fetal and neonate development. Changes in abundance of ubiquitously expressed miR were also observed. MiR-432 abundance was highest at the earliest stage of fetal development tested (60 day-old fetus) and decreased throughout development to the adult. Conversely, miR-24 and miR-27 exhibited greatest abundance in proliferating satellite cells and the adult, while abundance of miR-368, miR-376, and miR-423-5p was greatest in the neonate. Conclusion These data present a complete set of transcriptome profiles to evaluate miR abundance at specific stages of skeletal muscle growth in swine. Identification of these miR provides an initial group of miR that may play a vital role in muscle development and growth. PMID:19208255

  12. MicroRNA levels quantified in whole blood varies from PBMCs

    PubMed Central

    Atarod, Sadaf; Smith, Hannah; Dickinson, Anne; Wang, Xiao-Nong

    2015-01-01

    MicroRNAs are non-coding RNAs that negatively regulate mRNA expression and play significant roles in both health and disease. Differential microRNA expression has been used to aid diagnosis and discriminate disease stages. The accuracy and reliability of microRNA expression measurement is of utmost importance. Quantification of microRNA expression in human peripheral blood is commonly detected using total RNA extracted via different methods. To date, no convincing data are available showing whether microRNA quantification results can be influenced by the use of total RNA extracted from whole blood or peripheral blood mononuclear cells (PBMCs). This study examined miR-146a-5p and miR-155-5p expression using total RNA extracted in parallel from whole blood and PBMCs of 14 healthy volunteers. The data showed that the quantification of miRNA using total RNA extracted from whole blood varied from that of PBMCs, indicating that the miRNA expression was a result of all the different cell-types present in whole blood. Our results suggested that the source of total RNA and the statistical analyses performed are crucial considerations when designing miRNA research. PMID:26500764

  13. Amplification-based method for microRNA detection.

    PubMed

    Shen, Yanting; Tian, Fei; Chen, Zhenzhu; Li, Rui; Ge, Qinyu; Lu, Zuhong

    2015-09-15

    Over the last two decades, the study of miRNAs has attracted tremendous attention since they regulate gene expression post-transcriptionally and have been demonstrated to be dysregulated in many diseases. Detection methods with higher sensitivity, specificity and selectivity between precursors and mature microRNAs are urgently needed and widely studied. This review gave an overview of the amplification-based technologies including traditional methods, current modified methods and the cross-platforms of them combined with other techniques. Many progresses were found in the modified amplification-based microRNA detection methods, while traditional platforms could not be replaced until now. Several sample-specific normalizers had been validated, suggesting that the different normalizers should be established for different sample types and the combination of several normalizers might be more appropriate than a single universal normalizer. This systematic overview would be useful to provide comprehensive information for subsequent related studies and could reduce the un-necessary repetition in the future. PMID:25930002

  14. The multiple roles of microRNA-155 in oncogenesis.

    PubMed

    Higgs, Gadareth; Slack, Frank

    2013-01-01

    The microRNA miR-155 is prominent in cancer biology. Among microRNAs that have been linked to cancer, it is the most commonly overexpressed in malignancies (PNAS 109:20047-20052, 2012). Since its discovery, miR-155 has been implicated in promoting cancers of the breast, lung, liver, and lymphatic system. As such, targeted therapies may prove beneficial to cancer treatment. This review discusses the important role of miR-155 in oncogenesis. It synthesizes information from ten recent papers on miR-155, and includes an analysis and discussion of its association with cancer, interactions with other miRNAs, mechanisms of action, and the most promising available treatment options.Current debates in the field include the importance of miRNAs in general and their utility as targets in preventing tumorigenesis (Blood 119:513-520, 2012). Most of the papers being reviewed here confirm the role of miR-155 in oncogenesis (EMBO Mol Med 1:288-295, 2009). While there is some controversy surrounding recent research that claims that miR-155 may display anti-oncogenic or pro-immunological benefits (Cell Rep 2:1697-1709, 2012), most research seems to point to the importance of anti-miRs, with anti-miR-155 in particular, for cancer therapy. PMID:24073882

  15. Effects of simulated microgravity on microRNA and mRNA expression profile of rat soleus

    NASA Astrophysics Data System (ADS)

    Dai, Zhongquan; Wu, Feng; Qu, Lina

    Abstract Spaceflight induces muscle atrophy but mechanism is not well understood. Here, we quantified microRNAs (miRNAs) and mRNA shifts of rat soleus after 7, 14 and 28 days tail suspension (TS). Microarray data revealed that TS altered 23 miRNAs and 1313 mRNAs at least 2-fold change. QRT-PCR confirmed changes of miRNAs and mRNAs related to muscle atrophy. MiR-214, miR-486-5p and miR-320 family decreased, but Let-7e increased. Actn3 and myh4 displayed abundant upregulation and a3galt2 downregulated. Predicted targeted genes (whyz, ywhaz and SFRP2) of altered miRNAs decreased. Further analysis of gene functional annotation confirmed consistency of alteration profile between miRNAs and mRNA and enrichment of main clusters in regulation of muscle metabolism. Our results highlight the importance of miR-214, miR-486-5p, miR-320 and Let-7e in muscle atrophy process induced by microgravity.

  16. MicroRNA GENE EXPRESSION SIGNATURES IN THE DEVELOPING NEURAL TUBE

    PubMed Central

    Mukhopadhyay, Partha; Brock, Guy; Appana, Savitri; Webb, Cynthia; Greene, Robert M.; Pisano, M. Michele

    2011-01-01

    BACKGROUND Neurulation requires precise, spatio-temporal expression of numerous genes and coordinated interaction of signal transduction and gene regulatory networks, disruption of which may contribute to the etiology of neural tube (NT) defects. MicroRNAs are key modulators of cell and tissue differentiation. In order to define potential roles of miRNAs in development of the murine NT, miRNA microarray analysis was conducted to establish expression profiles, and identify miRNA target genes and functional gene networks. METHODS miRNA expression profiles in murine embryonic NTs derived from gestational days 8.5, 9.0 and 9.5 were defined and compared utilizing miRXplore™ microarrays from Miltenyi Biotech GmbH. Gene expression changes were verified by TaqMan™ quantitative Real-Time PCR. clValid R package and the UPGMA (hierarchical) clustering method were utilized for cluster analysis of the microarray data. Functional associations among selected miRNAs were examined via Ingenuity Pathway Analysis. RESULTS miRXplore™ chips enabled examination of 609 murine miRNAs. Expression of approximately 12% of these was detected in murine embryonic NTs. Clustering analysis revealed several developmentally regulated expression clusters among these expressed genes. Target analysis of differentially expressed miRNAs enabled identification of numerous target genes associated with cellular processes essential for normal NT development. Utilization of Ingenuity Pathway Analysis revealed interactive biological networks which connected differentially expressed miRNAs with their target genes, and highlighted functional relationships. CONCLUSIONS The present study defined unique gene expression signatures of a range of miRNAs in the developing NT during the critical period of NT morphogenesis. Analysis of miRNA target genes and gene interaction pathways revealed that specific miRNAs may direct expression of numerous genes encoding proteins which have been shown to be indispensable

  17. Potential microRNA biomarkers for acute ischemic stroke.

    PubMed

    Zeng, Ye; Liu, Jing-Xia; Yan, Zhi-Ping; Yao, Xing-Hong; Liu, Xiao-Heng

    2015-12-01

    Acute ischemic stroke is a significant cause of high morbidity and mortality in the aging population globally. However, current therapeutic strategies for acute ischemic stroke are limited. Atherosclerotic plaque is considered an independent risk factor for acute ischemic stroke. To identify biomarkers for carotid atheromatous plaque, bioinformatics analysis of the gene microarray data of plaque and intact tissue from individuals was performed. Differentially expressed genes (DEGs) were identified using the Multtest and Limma packages of R language, including 56 downregulated and 69 upregulated DEGs. Enriched microRNA (miRNA or miR) DEGs networks were generated using WebGestalt software and the STRING databases, and the miRNAs were validated using serum from acute ischemic stroke patients with reverse transcription quantitative PCR (RT‑qPCR). Four confirmed differentially expressed miRNAs (miR‑9, ‑22, ‑23 and ‑125) were associated with 28 upregulated DEGs, and 7 miRNAs (miR‑9, ‑30, ‑33, ‑124, ‑181, ‑218 and ‑330) were associated with 25 downregulated DEGs. Gene ontology (GO) function suggested that the confirmed miRNA‑targeted DEGs predominantly associated with signal transduction, the circulatory system, biological adhesion, striated muscle contraction, wound healing and the immune system. The confirmed miRNA‑targeted genes identified serve as potential therapeutic targets for acute ischemic stroke. PMID:26459744

  18. Roles of microRNA on cancer cell metabolism

    PubMed Central

    2012-01-01

    Advanced studies of microRNAs (miRNAs) have revealed their manifold biological functions, including control of cell proliferation, cell cycle and cell death. However, it seems that their roles as key regulators of metabolism have drawn more and more attention in the recent years. Cancer cells display increased metabolic autonomy in comparison to non-transformed cells, taking up nutrients and metabolizing them in pathways that support growth and proliferation. MiRNAs regulate cell metabolic processes through complicated mechanisms, including directly targeting key enzymes or transporters of metabolic processes and regulating transcription factors, oncogenes / tumor suppressors as well as multiple oncogenic signaling pathways. MiRNAs like miR-375, miR-143, miR-14 and miR-29b participate in controlling cancer cell metabolism by regulating the expression of genes whose protein products either directly regulate metabolic machinery or indirectly modulate the expression of metabolic enzymes, serving as master regulators, which will hopefully lead to a new therapeutic strategy for malignant cancer. This review focuses on miRNA regulations of cancer cell metabolism,including glucose uptake, glycolysis, tricarboxylic acid cycle and insulin production, lipid metabolism and amino acid biogenesis, as well as several oncogenic signaling pathways. Furthermore, the challenges of miRNA-based strategies for cancer diagnosis, prognosis and therapeutics have been discussed. PMID:23164426

  19. MicroRNA Biomarkers of Toxicity in Biological Matrices.

    PubMed

    Harrill, Alison H; McCullough, Shaun D; Wood, Charles E; Kahle, Juliette J; Chorley, Brian N

    2016-08-01

    Biomarker measurements that reliably correlate with tissue injury and that can be measured within accessible biofluids offer benefits in terms of cost, time, and convenience when assessing chemical and drug-induced toxicity in model systems or human cohorts. MicroRNAs (miRNAs) have emerged in recent years as a promising new class of biomarker for monitoring toxicity. Recent enthusiasm for miRNA biomarker research has been fueled by evidence that certain miRNAs are cell-type specific and are released during injury, thus raising the possibility of using biofluid-based miRNAs as a "liquid biopsy" that may be obtained by sampling extracellular fluids. As biomarkers, miRNAs demonstrate improved stability as compared with many protein markers and sequences are largely conserved across species, simplifying analytical techniques. Recent efforts have sought to identify miRNAs that are released into accessible biofluids following xenobiotic exposure, using compounds that target specific organs. Whereas still early in the discovery phase, miRNA biomarkers will have an increasingly important role in the assessment of adverse effects of both environmental chemicals and pharmaceutical drugs. Here, we review the current findings of biofluid-based miRNAs, as well as highlight technical challenges in assessing toxicologic pathology using these biomarkers. PMID:27462126

  20. MicroRNA and Epigenetics: Diagnostic and Therapeutic Opportunities

    PubMed Central

    Monroig, Paloma del C.; Calin, George A.

    2013-01-01

    MicroRNAs (miRNAs) are a large family of post-transcriptional regulators of gene expression that control cellular and developmental processes by targeting messenger RNAs (mRNA). These small non-coding RNAs (ncRNAs) are aberrantly expressed in cancer, and are known to contribute to tumorigenesis and disease progression. Therapeutic strategies based on modulating miRNAs activity are emerging due to the ability of these ncRNAs to influence cellular behavior. MiRNA levels predict disease prognosis and overall patient survival, and reconstituting their basal levels has been proven to inhibit tumor growth and metastasis. Different delivery mechanisms have been tested in vivo, however many challenges need to be overcome before their utilization in the clinic. Moreover, it has been found that circulating miRNAs in body fluids have the potential to reshape cancer diagnosis and prognosis by functioning as biomarkers and indicators of progression and metastasis. These miRNAs as biofluids-based biomarkers provide an alternative strategy for early diagnosis and treatment of cancer patients. PMID:23515489

  1. MicroRNA and MET in lung cancer

    PubMed Central

    2015-01-01

    MicroRNAs (miRNAs) are a class of small non-protein coding RNAs that modulate important cellular functions via their post-transcriptional regulation of messenger RNAs (mRNAs). Recent evidences from multiple tumor types and model systems implicate miRNA dysregulation as a common mechanism of tumorigenesis, cancer progression and resistance to therapy. Several miRNAs are dysregulated in cancers and a single miRNA can have multiple targets involved in different oncogenic pathways. MET, the tyrosine kinase receptor for hepatocyte growth factor (HGF), has a central role in lung cancer development and in acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors; it has been predicted and shown to be the target gene of multiple miRNAs, which play a crucial role in controlling its activity in a stimulatory or inhibitory sense. In this review we will focus on the most important and recent studies about the role of miRNAs in the control of MET expression, reporting also the progress made using miRNAs for therapy of lung cancer. PMID:25992367

  2. MicroRNA in neurodegenerative drug discovery: the way forward?

    PubMed

    Campbell, Kristyn; Booth, Stephanie A

    2015-01-01

    Neurodegenerative diseases occur when neuronal cells in the brain or spinal cord progressively lose function and eventually die. Pathological analysis of these tissues reveals changes that include the loss of synapses, tangles of misfolded protein and immune cell activation, even during very early stages of disease well before debilitating clinical signs are apparent. This suggests that if neurodegeneration is treated early enough, drugs designed to delay the progress of these diseases by either repairing the early damage and loss of neurons, or protecting neuron functionality from further insult, may be efficacious. MicroRNAs (miRNAs) are small non-coding RNAs that can post-transcriptionally regulate gene expression. They are particularly numerous within neurons where many are expressed with high specificity, which suggests that they have important roles in the healthy brain. Indeed, miRNAs are essential for the post-mitotic survival of neurons, implying a crucial role in survival and neuroprotection. This has focused attention on exploring the use of miRNA-based drugs as a means to correct cellular abnormalities and maintain neuronal function in neurodegenerative diseases. These efforts are spurred on by the rapid progress to clinical trials for a number of miRNA-based therapies for other diseases such as cardiovascular diseases, fibrosis and cancer. PMID:25405898

  3. Principles of microRNA involvement in human cancers

    PubMed Central

    Ling, Hui; Zhang, Wei; Calin, George A.

    2011-01-01

    Naturally occurring microRNAs (miRNAs), small non-coding RNAs of 19 to 24 nucleotides (nt), are encoded in the genomes of invertebrates, vertebrates, and plants. miRNAs act as regulators of gene expression during development and differentiation at the transcriptional, posttranscriptional, and/or translational levels, although most target genes are still elusive. Many miRNAs are conserved in sequence between distantly related organisms, suggesting that these molecules participate in essential processes. In this review, we present principles related to the basic and translational research that has emerged in the last decade, a period that can be truly considered the “miRNA revolution” in molecular oncology. These principles include the regulation mechanism of miRNA expression, functions of miRNAs in cancers, diagnostic values and therapeutic potentials of miRNAs. Furthermore, we present a compendium of information about the main miRNAs that have been identified in the last several years as playing important roles in cancers. Also, we orient the reader to several additional reviews that may provide a deeper understanding of this new and exciting field of research. PMID:22035854

  4. MicroRNA-7a regulates pancreatic β cell function

    PubMed Central

    Latreille, Mathieu; Hausser, Jean; Stützer, Ina; Zhang, Quan; Hastoy, Benoit; Gargani, Sofia; Kerr-Conte, Julie; Pattou, Francois; Zavolan, Mihaela; Esguerra, Jonathan L.S.; Eliasson, Lena; Rülicke, Thomas; Rorsman, Patrik; Stoffel, Markus

    2014-01-01

    Dysfunctional microRNA (miRNA) networks contribute to inappropriate responses following pathological stress and are the underlying cause of several disease conditions. In pancreatic β cells, miRNAs have been largely unstudied and little is known about how specific miRNAs regulate glucose-stimulated insulin secretion (GSIS) or impact the adaptation of β cell function to metabolic stress. In this study, we determined that miR-7 is a negative regulator of GSIS in β cells. Using Mir7a2 deficient mice, we revealed that miR-7a2 regulates β cell function by directly regulating genes that control late stages of insulin granule fusion with the plasma membrane and ternary SNARE complex activity. Transgenic mice overexpressing miR-7a in β cells developed diabetes due to impaired insulin secretion and β cell dedifferentiation. Interestingly, perturbation of miR-7a expression in β cells did not affect proliferation and apoptosis, indicating that miR-7 is dispensable for the maintenance of endocrine β cell mass. Furthermore, we found that miR-7a levels are decreased in obese/diabetic mouse models and human islets from obese and moderately diabetic individuals with compensated β cell function. Our results reveal an interconnecting miR-7 genomic circuit that regulates insulin granule exocytosis in pancreatic β cells and support a role for miR-7 in the adaptation of pancreatic β cell function in obesity and type 2 diabetes. PMID:24789908

  5. [Digital droplet PCR - a prospective technological approach to quantitative profiling of microRNA].

    PubMed

    Kiseleva, Y Y; Ptitsyn, K G; Radko, S P; Zgoda, V G; Archakov, A I

    2016-05-01

    MicroRNA is a special type of regulatory molecules governing gene expression. Circulating microRNAs found in blood and other biological fluids are considered today as potential biomarkers of human pathology. Presently, quantitative alterations of particular microRNAs are revealed for a large number of oncological diseases and other disorders. The recently emerged method of digital droplet PCR (ddPCR) possesses a number of advantages making this method the most suitable for verification and validation of perspective microRNA markers of human pathologies. Among these advantages are the high accuracy and reproducibility of microRNA quantification as well as the capability to directly measure the absolute number of microRNA copies with the large dynamic range and a high throughput. The paper reviews microRNA biogenesis, the origin of circulating microRNAs, and methods used for their quantification. The special technical features of ddPCR, which make it an attractive method both for studying microRNAs as biomarkers of human pathologies and for basic research devoted to aspects of gene regulation by microRNA molecules, are also discussed. PMID:27562993

  6. Altered microRNA expression profiles in a rat model of spina bifida.

    PubMed

    Qin, Pan; Li, Lin; Zhang, Da; Liu, Qiu-Liang; Chen, Xin-Rang; Yang, He-Ying; Fan, Ying-Zhong; Wang, Jia-Xiang

    2016-03-01

    MicroRNAs (miRNAs) are dynamically regulated during neurodevelopment, yet few reports have examined their role in spina bifida. In this study, we used an established fetal rat model of spina bifida induced by intragastrically administering olive oil-containing all-trans retinoic acid to dams on day 10 of pregnancy. Dams that received intragastric administration of all-trans retinoic acid-free olive oil served as controls. The miRNA expression profile in the amniotic fluid of rats at 20 days of pregnancy was analyzed using an miRNA microarray assay. Compared with that in control fetuses, the expression of miRNA-9, miRNA-124a, and miRNA-138 was significantly decreased (> 2-fold), whereas the expression of miRNA-134 was significantly increased (> 4-fold) in the amniotic fluid of rats with fetuses modeling spina bifida. These results were validated using real-time quantitative reverse-transcription polymerase chain reaction. Hierarchical clustering analysis of the microarray data showed that these differentially expressed miRNAs could distinguish fetuses modeling spina bifida from control fetuses. Our bioinformatics analysis suggested that these differentially expressed miRNAs were associated with many cytological pathways, including a nervous system development signaling pathway. These findings indicate that further studies are warranted examining the role of miRNAs through their regulation of a variety of cell functional pathways in the pathogenesis of spina bifida. Such studies may provide novel targets for the early diagnosis and treatment of spina bifida. PMID:27127493

  7. Altered microRNA expression profiles in a rat model of spina bifida

    PubMed Central

    Qin, Pan; Li, Lin; Zhang, Da; Liu, Qiu-liang; Chen, Xin-rang; Yang, He-ying; Fan, Ying-zhong; Wang, Jia-xiang

    2016-01-01

    MicroRNAs (miRNAs) are dynamically regulated during neurodevelopment, yet few reports have examined their role in spina bifida. In this study, we used an established fetal rat model of spina bifida induced by intragastrically administering olive oil-containing all-trans retinoic acid to dams on day 10 of pregnancy. Dams that received intragastric administration of all-trans retinoic acid-free olive oil served as controls. The miRNA expression profile in the amniotic fluid of rats at 20 days of pregnancy was analyzed using an miRNA microarray assay. Compared with that in control fetuses, the expression of miRNA-9, miRNA-124a, and miRNA-138 was significantly decreased (> 2-fold), whereas the expression of miRNA-134 was significantly increased (> 4-fold) in the amniotic fluid of rats with fetuses modeling spina bifida. These results were validated using real-time quantitative reverse-transcription polymerase chain reaction. Hierarchical clustering analysis of the microarray data showed that these differentially expressed miRNAs could distinguish fetuses modeling spina bifida from control fetuses. Our bioinformatics analysis suggested that these differentially expressed miRNAs were associated with many cytological pathways, including a nervous system development signaling pathway. These findings indicate that further studies are warranted examining the role of miRNAs through their regulation of a variety of cell functional pathways in the pathogenesis of spina bifida. Such studies may provide novel targets for the early diagnosis and treatment of spina bifida. PMID:27127493

  8. Cocaine Enhances HIV-1 Infectivity in Monocyte Derived Dendritic Cells by Suppressing microRNA-155

    PubMed Central

    Napuri, Jessica; Pilakka-Kanthikeel, Sudheesh; Raymond, Andrea; Agudelo, Marisela; Yndart-Arias, Adriana; Saxena, Shailendra K.; Nair, Madhavan

    2013-01-01

    Cocaine and other drugs of abuse increase HIV-induced immunopathogenesis; and neurobiological mechanisms of cocaine addiction implicate a key role for microRNAs (miRNAs), single-stranded non-coding RNAs that regulate gene expression and defend against viruses. In fact, HIV defends against miRNAs by actively suppressing the expression of polycistronic miRNA cluster miRNA-17/92, which encodes miRNAs including miR-20a. IFN-g production by natural killer cells is regulated by miR-155 and this miRNA is also critical to dendritic cell (DC) maturation. However, the impact of cocaine on miR-155 expression and subsequent HIV replication is unknown. We examined the impact of cocaine on two miRNAs, miR-20a and miR-155, which are integral to HIV replication, and immune activation. Using miRNA isolation and analysis, RNA interference, quantitative real time PCR, and reporter assays we explored the effects of cocaine on miR-155 and miR-20 in the context of HIV infection. Here we demonstrate using monocyte-derived dendritic cells (MDCCs) that cocaine significantly inhibited miR-155 and miR-20a expression in a dose dependent manner. Cocaine and HIV synergized to lower miR-155 and miR-20a in MDDCs by 90%. Cocaine treatment elevated LTR-mediated transcription and PU.1 levels in MDCCs. But in context of HIV infection, PU.1 was reduced in MDDCs regardless of cocaine presence. Cocaine increased DC-SIGN and and decreased CD83 expression in MDDC, respectively. Overall, we show that cocaine inhibited miR-155 and prevented maturation of MDDCs; potentially, resulting in increased susceptibility to HIV-1. Our findings could lead to the development of novel miRNA-based therapeutic strategies targeting HIV infected cocaine abusers. PMID:24391808

  9. MicroRNA and gene expression patterns in the differentiation of human embryonic stem cells

    PubMed Central

    Ren, Jiaqiang; Jin, Ping; Wang, Ena; Marincola, Francesco M; Stroncek, David F

    2009-01-01

    Background The unique features of human embryonic stem (hES) cells make them the best candidate resource for both cell replacement therapy and development research. However, the molecular mechanisms responsible for the simultaneous maintenance of their self-renewal properties and undifferentiated state remain unclear. Non-coding microRNAs (miRNA) which regulate mRNA cleavage and inhibit encoded protein translation exhibit temporal or tissue-specific expression patterns and they play an important role in development timing. Results In this study, we analyzed miRNA and gene expression profiles among samples from 3 hES cell lines (H9, I6 and BG01v), differentiated embryoid bodies (EB) derived from H9 cells at different time points, and 5 adult cell types including Human Microvascular Endothelial Cells (HMVEC), Human Umbilical Vein Endothelial Cells (HUVEC), Umbilical Artery Smooth Muscle Cells (UASMC), Normal Human Astrocytes (NHA), and Lung Fibroblasts (LFB). This analysis rendered 104 miRNAs and 776 genes differentially expressed among the three cell types. Selected differentially expressed miRNAs and genes were further validated and confirmed by quantitative real-time-PCR (qRT-PCR). Especially, members of the miR-302 cluster on chromosome 4 and miR-520 cluster on chromosome 19 were highly expressed in undifferentiated hES cells. MiRNAs in these two clusters displayed similar expression levels. The members of these two clusters share a consensus 7-mer seed sequence and their targeted genes had overlapping functions. Among the targeted genes, genes with chromatin structure modification function are enriched suggesting a role in the maintenance of chromatin structure. We also found that the expression level of members of the two clusters, miR-520b and miR-302c, were negatively correlated with their targeted genes based on gene expression analysis Conclusion We identified the expression patterns of miRNAs and gene transcripts in the undifferentiation of human embryonic

  10. The use of artificial microRNA technology to control gene expression in Arabidopsis thaliana.

    PubMed

    Eamens, Andrew L; McHale, Marcus; Waterhouse, Peter M

    2014-01-01

    In plants, double-stranded RNA (dsRNA) is an effective trigger of RNA silencing, and several classes of endogenous small RNA (sRNA), processed from dsRNA substrates by DICER-like (DCL) endonucleases, are essential in controlling gene expression. One such sRNA class, the microRNAs (miRNAs) control the expression of closely related genes to regulate all aspects of plant development, including the determination of leaf shape, leaf polarity, flowering time, and floral identity. A single miRNA sRNA silencing signal is processed from a long precursor transcript of nonprotein-coding RNA, termed the primary miRNA (pri-miRNA). A region of the pri-miRNA is partially self-complementary allowing the transcript to fold back onto itself to form a stem-loop structure of imperfectly dsRNA. Artificial miRNA (amiRNA) technology uses endogenous pri-miRNAs, in which the miRNA and miRNA* (passenger strand of the miRNA duplex) sequences have been replaced with corresponding amiRNA/amiRNA* sequences that direct highly efficient RNA silencing of the targeted gene. Here, we describe the rules for amiRNA design, as well as outline the PCR and bacterial cloning procedures involved in the construction of an amiRNA plant expression vector to control target gene expression in Arabidopsis thaliana. PMID:24057368

  11. MicroRNA binding sites in C. elegans 3' UTRs.

    PubMed

    Liu, Chaochun; Rennie, William A; Mallick, Bibekanand; Kanoria, Shaveta; Long, Dang; Wolenc, Adam; Carmack, C Steven; Ding, Ye

    2014-01-01

    MicroRNAs (miRNAs) are post-transcriptional regulators of gene expression. Since the discovery of lin-4, the founding member of the miRNA family, over 360 miRNAs have been identified for Caenorhabditis elegans (C. elegans). Prediction and validation of targets are essential for elucidation of regulatory functions of these miRNAs. For C. elegans, crosslinking immunoprecipitation (CLIP) has been successfully performed for the identification of target mRNA sequences bound by Argonaute protein ALG-1. In addition, reliable annotation of the 3' untranslated regions (3' UTRs) as well as developmental stage-specific expression profiles for both miRNAs and 3' UTR isoforms are available. By utilizing these data, we developed statistical models and bioinformatics tools for both transcriptome-scale and developmental stage-specific predictions of miRNA binding sites in C. elegans 3' UTRs. In performance evaluation via cross validation on the ALG-1 CLIP data, the models were found to offer major improvements over established algorithms for predicting both seed sites and seedless sites. In particular, our top-ranked predictions have a substantially higher true positive rate, suggesting a much higher likelihood of positive experimental validation. A gene ontology analysis of stage-specific predictions suggests that miRNAs are involved in dynamic regulation of biological functions during C. elegans development. In particular, miRNAs preferentially target genes related to development, cell cycle, trafficking, and cell signaling processes. A database for both transcriptome-scale and stage-specific predictions and software for implementing the prediction models are available through the Sfold web server at http://sfold.wadsworth.org. PMID:24827614

  12. Investigation on torquetenovirus (TTV) microRNA transcriptome in vivo.

    PubMed

    Vignolini, Tiziano; Macera, Lisa; Antonelli, Guido; Pistello, Mauro; Maggi, Fabrizio; Giannecchini, Simone

    2016-06-01

    Torquetenovirus (TTV) is a widespread anellovirus that establishes persistent infections in human showing an increased viremia in immunosuppressed patients. TTV possesses microRNA (miRNA)-coding sequences that might be involved in viral immune evasion. Here, the presence of TTV DNA and miRNAs expression was investigated in plasma samples of 77 diseased (20 infected with human immunodeficiency virus (HIV), 18 infected with hepatitis B (HBV) virus, 18 infected with hepatitis C (HCV) virus, 21 solid organ transplanted) patients, and 25 healthy controls. TTV prevalence was significantly different in healthy controls (60%, 15/25) versus diseased patients (80%, 62/77), showing the highest TTV loads in transplant recipients. Genetic TTV analysis showed the highest prevalence of group 1, followed by groups 3, 4 and 5, and a lack of isolates of group 2. The expression of at least one TTV miRNAs of group 1, 3 and 5 was found in exosomes of plasma of the great majority of individuals (96%, 98/102 subjects) showing the higher prevalence of miRNAs of TTV group 3 (90%, 92/102), followed by miRNAs of group 1 (66%, 67/102), and miRNA of group 5 (49%, 50/102). TTV miRNAs expression and TTV viremia were not always directly correlated, and significant differences appeared in production of some TTV miRNAs between healthy controls and diseased patients. The reported TTV miRNAs status in exosomes encourages further investigation to understand their potential role in the expansion of anelloviruses upon immunosuppression. PMID:26959653

  13. microRNA and gene networks in human pancreatic cancer.

    PubMed

    Zhu, Minghui; Xu, Zhiwen; Wang, Kunhao; Wang, Ning; Li, Yang

    2013-10-01

    To date, scientists have obtained a substantial amount of knowledge with regard to genes and microRNAs (miRNAs) in pancreatic cancer (PC). However, deciphering the regulatory mechanism of these genes and miRNAs remains difficult. In the present study, three regulatory networks consisting of a differentially-expressed network, a related network and a global network, were constructed in order to identify the mechanisms and certain key miRNA and gene pathways in PC. The interactions between transcription factors (TFs) and miRNAs, miRNAs and target genes and an miRNA and its host gene were investigated. The present study compared and analyzed the similarities and differences between the three networks in order to distinguish the key pathways. Certain pathways involving the differentially-expressed genes and miRNAs demonstrated specific features. TP53 and hsa-miR-125b were observed to form a self-adaptation association. A further 16 significant differentially-expressed miRNAs were obtained and it was observed that an miRNA and its host gene exhibit specific features in PC, for example, hsa-miR-196a-1 and its host gene, HOXB7, form a self-adaptation association. The differentially-expressed network partially illuminated the mechanism of PC. The present study provides comprehensive data that is associated with PC and may aid future studies in obtaining pertinent data results with regards to PC. In the future, an improved understanding of PC may be obtained through an increased knowledge of the occurrence, mechanism, improvement, metastasis and treatment of the disease. PMID:24137477

  14. MicroRNA Signatures of Drought Signaling in Rice Root

    PubMed Central

    Nikpay, Nava; Ebrahimi, Mohammad Ali; Bihamta, Mohammad Reza; Mardi, Mohsen; Salekdeh, Ghasem Hosseini

    2016-01-01

    Background Drought stress is one of the most important abiotic stresses and the main constraint to rice agriculture. MicroRNA-mediated post-transcriptional gene regulation is one of the ways to establish drought stress tolerance in plants. MiRNAs are 20–24-nt regulatory RNAs that play an important role in regulating plant gene expression upon exposure to biotic and abiotic stresses. Methodology/Principal Findings In this study, we applied a partial root drying system as well as a complete root drying system to identify miRNAs involved in conditions of drought stress, drought signaling and wet signaling using high-throughput sequencing. To this end, we produced four small RNA libraries: (1) fully-watered (WW), (2) fully-droughted (WD), and split-root systems where (3) one-half was well watered (SpWW) and (4) the other half was water-deprived (SpWD). Our analysis revealed 10,671 and 783 unique known and novel miRNA reads in all libraries, respectively. We identified, 65 (52 known + 13 novel), 72 (61 known + 11 novel) and 51 (38 known + 13 novel) miRNAs that showed differential expression under conditions of drought stress, drought signaling and wet signaling, respectively. The results of quantitative real-time PCR showed expression patterns similar to the high-throughput sequencing results. Furthermore, our target prediction led to the identification of 244, 341 and 239 unique target genes for drought-stress-, drought-signaling- and wet-signaling-responsive miRNAs, respectively. Conclusions/Significance Our results suggest that miRNAs that are responsive under different conditions could play different roles in the regulation of abscisic acid signaling, calcium signaling, detoxification and lateral root formation. PMID:27276090

  15. MicroRNA: Small RNA mediators of the brains genomic response to environmental stress.

    PubMed

    Hollins, Sharon L; Cairns, Murray J

    2016-08-01

    The developmental processes that establish the synaptic architecture of the brain while retaining capacity for activity-dependent remodeling, are complex and involve a combination of genetic and epigenetic influences. Dysregulation of these processes can lead to problems with neural circuitry which manifest in humans as a range of neurodevelopmental syndromes, such as schizophrenia, bipolar disorder and fragile X mental retardation. Recent studies suggest that prenatal, postnatal and intergenerational environmental factors play an important role in the aetiology of stress-related psychopathology. A number of these disorders have been shown to display epigenetic changes in the postmortem brain that reflect early life experience. These changes affect the regulation of gene expression though chromatin remodeling (transcriptional) and post-transcriptional influences, especially small noncoding microRNA (miRNA). These dynamic and influential molecules appear to play an important function in both brain development and its adaption to stress. In this review, we examine the role of miRNA in mediating the brain's response to both prenatal and postnatal environmental perturbations and explore how stress- induced alterations in miRNA expression can regulate the stress response via modulation of the immune system. Given the close relationship between environmental stress, miRNA, and brain development/function, we assert that miRNA hold a significant position at the molecular crossroads between neural development and adaptations to environmental stress. A greater understanding of the dynamics that mediate an individual's predisposition to stress-induced neuropathology has major human health benefits and is an important area of research. PMID:27317386

  16. Mutual antagonism between hepatitis B viral mRNA and host microRNA let-7

    PubMed Central

    Takata, Akemi; Otsuka, Motoyuki; Ohno, Motoko; Kishikawa, Takahiro; Yoshikawa, Takeshi; Koike, Kazuhiko

    2016-01-01

    The interplay between viral and host factors plays a major role in viral pathogenesis. Hepatitis B virus (HBV) infection is a global health problem that leads to liver cirrhosis and hepatocellular carcinoma (HCC). Although HBV proteins have been studied extensively about their implication in hepatocarcinogenesis, the molecular mechanisms of oncogenesis are still largely unknown. A recent concept in gene regulation, in which competitive endogenous RNAs compete for common microRNAs (miRNAs), suggests that mRNA targets are key elements in the regulation of miRNA availability. Here, we show that HBV mRNA in the preS2 region can be targeted by host miRNA let-7 g. This leads to the sequestration of let-7 g and inhibition of let-7 g function. The expression of HBV transcripts, including the preS2 region, de-repressed let-7 g targets, which may contribute to long-term oncogenesis. HBV transcript-expressing transgenic mice, but not non-targeted transcript-expressing mice, were more prone to chemically induced hepatoocarcinogenesis. Let-7 target protein expression was upregulated in human HCC tissues derived from HBV-infected patients. On the other hand, let-7 g inhibited HBV preS2 protein expression and viral products. These results suggest that the interplay between viral intermediate transcripts during HBV replication and host miRNAs is crucial to the pathogenesis of chronic viral infection. PMID:26979389

  17. Mutual antagonism between hepatitis B viral mRNA and host microRNA let-7.

    PubMed

    Takata, Akemi; Otsuka, Motoyuki; Ohno, Motoko; Kishikawa, Takahiro; Yoshikawa, Takeshi; Koike, Kazuhiko

    2016-01-01

    The interplay between viral and host factors plays a major role in viral pathogenesis. Hepatitis B virus (HBV) infection is a global health problem that leads to liver cirrhosis and hepatocellular carcinoma (HCC). Although HBV proteins have been studied extensively about their implication in hepatocarcinogenesis, the molecular mechanisms of oncogenesis are still largely unknown. A recent concept in gene regulation, in which competitive endogenous RNAs compete for common microRNAs (miRNAs), suggests that mRNA targets are key elements in the regulation of miRNA availability. Here, we show that HBV mRNA in the preS2 region can be targeted by host miRNA let-7 g. This leads to the sequestration of let-7 g and inhibition of let-7 g function. The expression of HBV transcripts, including the preS2 region, de-repressed let-7 g targets, which may contribute to long-term oncogenesis. HBV transcript-expressing transgenic mice, but not non-targeted transcript-expressing mice, were more prone to chemically induced hepatoocarcinogenesis. Let-7 target protein expression was upregulated in human HCC tissues derived from HBV-infected patients. On the other hand, let-7 g inhibited HBV preS2 protein expression and viral products. These results suggest that the interplay between viral intermediate transcripts during HBV replication and host miRNAs is crucial to the pathogenesis of chronic viral infection. PMID:26979389

  18. From microRNA functions to microRNA therapeutics: Novel targets and novel drugs in breast cancer research and treatment

    PubMed Central

    PIVA, ROBERTA; SPANDIDOS, DEMETRIOS A.; GAMBARI, ROBERTO

    MicroRNAs (miRNAs or miRs) are a family of small non-coding RNAs that regulate gene expression by the sequence-selective targeting of mRNAs, leading to translational repression or mRNA degradation, depending on the degree of complementarity with target mRNA sequences. miRNAs play a crucial role in cancer. In the case of breast tumors, several studies have demonstrated a correlation between: i) the expression profile of oncogenic miRNAs (oncomiRs) or tumor suppressor miRNAs and ii) the tumorigenic potential of triple-negative [estrogen receptor (ER), progesterone receptor (PR) and Her2/neu] primary breast cancers. Among the miRNAs involved in breast cancer, miR-221 plays a crucial role for the following reasons: i) miR-221 is significantly overexpressed in triple-negative primary breast cancers; ii) the oncosuppressor p27 Kip1 , a validated miR-221 target, is downregulated in aggressive cancer cell lines; and iii) the upregulation of a key transcription factor, Slug, appears to be crucial, since it binds to the miR-221/miR-222 promoter and is responsible for the high expression of the miR-221/miR-222 cluster in breast cancer cells. A Slug/miR-221 network has been suggested, linking miR-221 activity with the downregulation of a Slug repressor, leading to Slug/miR-221 upregulation and p27 Kip1 downregulation. Interference with this process can be achieved using antisense miRNA (antagomiR) molecules targeting miR-221, inducing the down-regulation of Slug and the upregulation of p27 Kip1 . PMID:23939688

  19. Inhibiting the oncogenic mir-221 by microRNA sponge: toward microRNA-based therapeutics for hepatocellular carcinoma

    PubMed Central

    Moshiri, Farzaneh; Callegari, Elisa; D'Abundo, Lucilla; Corrà, Fabio; Lupini, Laura; Sabbioni, Silvia

    2014-01-01

    Aim We evaluated the capability of “microRNA sponges” in sequestering and inhibiting the over-expressed miR-221 in HCC cell lines. Background Advanced hepatocellular carcinoma (HCC) is a serious public health problem, with no effective cure at present. It has been demonstrated that the deregulation of microRNAs expression contributes to tumorigenesis. In HCC, miR-221 was shown to be up-regulated in more than 70% of the cases and was associated with higher tumor stage, metastasis and a shorter time to recurrence after surgery, suggesting an important pathogenic role. A tumor promoting function of miR-221 was proved in a transgenic mouse model, which was predisposed to the development of liver cancers. These findings suggested that miR-221 could represent a potential target for anti-tumor approaches. Material and Methods Novel adeno and adeno-associated viral vectors (AAVs) were developed: they were genetically modified to drive the expression of multiple binding sites for miR-221, the “miR-221 sponge”, which was designed to sequester miR-221 cellular molecules. Results Analysis of viral vectors activity in HCC cells revealed their capability to reduce miR-221 endogenous levels, which was accompanied by the increase in CDKN1B/ p27 protein, a known target of miR-221. An increase in apoptosis was also measured in Hep3B cells after infection with any of the two viral vectors in comparison with control vectors, with stronger effects induced by adenovirus compared to AAV vectors. Conclusion The depletion of oncogenic microRNAs represents a potential anti-cancer approach that needs to be tested for safety and efficacy. Here, we describe the development of novel “miR-221 sponge” vectors, which can reduce miR-221 activity in vitro and may be used for in vivo delivery. PMID:25436097

  20. Discordant Expression of Circulating microRNA from Cellular and Extracellular Sources

    PubMed Central

    Levy, Daniel; Larson, Martin; Gerstein, Mark; Mick, Eric; Rozowsky, Joel; Kitchen, Robert; Murthy, Venkatesh; Mikalev, Ekaterina; Freedman, Jane E.

    2016-01-01

    MicroRNA (miRNA) expression has rapidly grown into one of the largest fields for disease characterization and development of clinical biomarkers. Consensus is lacking in regards to the optimal sample source or if different circulating sources are concordant. Here, using miRNA measurements from contemporaneously obtained whole blood- and plasma-derived RNA from 2391 individuals, we demonstrate that plasma and blood miRNA levels are divergent and may reflect different biological processes and disease associations. PMID:27123852

  1. Composition and Expression of Conserved MicroRNA Genes in Diploid Cotton (Gossypium) Species

    PubMed Central

    Gong, Lei; Kakrana, Atul; Arikit, Siwaret; Meyers, Blake C.; Wendel, Jonathan F.

    2013-01-01

    MicroRNAs are ubiquitous in plant genomes but vary greatly in their abundance within and conservation among plant lineages. To gain insight into the evolutionary birth/death dynamics of microRNA families, we sequenced small RNA and 5′-end PARE libraries generated from two closely related species of Gossypium. Here, we demonstrate that 33 microRNA families, with similar copy numbers and average evolutionary rates, are conserved in the two congeneric cottons. Analysis of the presence/absence of these microRNA families in other land plants sheds light on their depth of phylogenetic origin and lineage-specific loss/gain. Conserved microRNA families in Gossypium exhibit a striking interspecific asymmetry in expression, potentially connected to relative proximity to neighboring transposable elements. A complex correlated expression pattern of microRNA target genes with their controlling microRNAs indicates that possible functional divergence of conserved microRNA families can also exist even within a single plant genus. PMID:24281048

  2. Prediction of MicroRNA-Disease Associations Based on Social Network Analysis Methods.

    PubMed

    Zou, Quan; Li, Jinjin; Hong, Qingqi; Lin, Ziyu; Wu, Yun; Shi, Hua; Ju, Ying

    2015-01-01

    MicroRNAs constitute an important class of noncoding, single-stranded, ~22 nucleotide long RNA molecules encoded by endogenous genes. They play an important role in regulating gene transcription and the regulation of normal development. MicroRNAs can be associated with disease; however, only a few microRNA-disease associations have been confirmed by traditional experimental approaches. We introduce two methods to predict microRNA-disease association. The first method, KATZ, focuses on integrating the social network analysis method with machine learning and is based on networks derived from known microRNA-disease associations, disease-disease associations, and microRNA-microRNA associations. The other method, CATAPULT, is a supervised machine learning method. We applied the two methods to 242 known microRNA-disease associations and evaluated their performance using leave-one-out cross-validation and 3-fold cross-validation. Experiments proved that our methods outperformed the state-of-the-art methods. PMID:26273645

  3. Identification of clustered microRNAs using an ab initio prediction method

    PubMed Central

    Sewer, Alain; Paul, Nicodème; Landgraf, Pablo; Aravin, Alexei; Pfeffer, Sébastien; Brownstein, Michael J; Tuschl, Thomas; van Nimwegen, Erik; Zavolan, Mihaela

    2005-01-01

    Background MicroRNAs (miRNAs) are endogenous 21 to 23-nucleotide RNA molecules that regulate protein-coding gene expression in plants and animals via the RNA interference pathway. Hundreds of them have been identified in the last five years and very recent works indicate that their total number is still larger. Therefore miRNAs gene discovery remains an important aspect of understanding this new and still widely unknown regulation mechanism. Bioinformatics approaches have proved to be very useful toward this goal by guiding the experimental investigations. Results In this work we describe our computational method for miRNA prediction and the results of its application to the discovery of novel mammalian miRNAs. We focus on genomic regions around already known miRNAs, in order to exploit the property that miRNAs are occasionally found in clusters. Starting with the known human, mouse and rat miRNAs we analyze 20 kb of flanking genomic regions for the presence of putative precursor miRNAs (pre-miRNAs). Each genome is analyzed separately, allowing us to study the species-specific identity and genome organization of miRNA loci. We only use cross-species comparisons to make conservative estimates of the number of novel miRNAs. Our ab initio method predicts between fifty and hundred novel pre-miRNAs for each of the considered species. Around 30% of these already have experimental support in a large set of cloned mammalian small RNAs. The validation rate among predicted cases that are conserved in at least one other species is higher, about 60%, and many of them have not been detected by prediction methods that used cross-species comparisons. A large fraction of the experimentally confirmed predictions correspond to an imprinted locus residing on chromosome 14 in human, 12 in mouse and 6 in rat. Our computational tool can be accessed on the world-wide-web. Conclusion Our results show that the assumption that many miRNAs occur in clusters is fruitful for the discovery of

  4. Detecting pan-cancer conserved microRNA modules from microRNA expression profiles across multiple cancers.

    PubMed

    Liu, Zhaowen; Zhang, Junying; Yuan, Xiguo; Liu, Baobao; Liu, Yajun; Li, Aimin; Zhang, Yuanyuan; Sun, Xiaohan; Tuo, Shouheng

    2015-08-01

    MicroRNAs (miRNAs) play an indispensable role in cancer initiation and progression. Different cancers have some common hallmarks in general. Analyzing miRNAs that consistently contribute to different cancers can help us to discover the relationship between miRNAs and traits shared by cancers. Most previous works focus on analyzing single miRNA. However, dysregulation of a single miRNA is generally not sufficient to contribute to complex cancer processes. In this study, we put emphasis on analyzing cooperation of miRNAs across cancers. We assume that miRNAs can cooperatively regulate oncogenic pathways and contribute to cancer hallmarks. Such a cooperation is modeled by a miRNA module referred to as a pan-cancer conserved miRNA module. The module consists of miRNAs which simultaneously regulate cancers and are significantly intra-correlated. A novel computational workflow for the module discovery is presented. Multiple modules are discovered from miRNA expression profiles using the method. The function of top two ranked modules are analyzed using the mRNAs which correlate to all the miRNAs in a module across cancers, inferring that the two modules function in regulating the cell cycle which relates to cancer hallmarks as self sufficiency in growth signals and insensitivity to antigrowth signals. Additionally, two novel miRNAs mir-590 and mir-629 are found to cooperate with well-known onco-miRNAs in the modules to contribute to cancers. We also found that PTEN, which is a well known tumor suppressor that regulates the cell cycle, is a common target of miRNAs in the top-one module and cooperative control of PTEN can be a reason for the miRNAs' cooperation. We believe that analyzing the cooperative mechanism of the miRNAs in modules rather than focusing on only single miRNAs may help us know more about the complicated relationship between miRNAs and cancers and develop more effective treatment strategies for cancers. PMID:26052692

  5. MicroRNA Expression Profiles as Biomarkers of Minor Salivary Gland Inflammation and Dysfunction in Sjögren's Syndrome

    PubMed Central

    Alevizos, Ilias; Alexander, Stefanie; Turner, R. James; Illei, Gabor G.

    2013-01-01

    Objective MicroRNA reflect physiologic and pathologic processes and may be used as biomarkers of concurrent pathophysiologic events in complex settings such as autoimmune diseases. We generated microRNA microarray profiles from the minor salivary glands of control subjects without Sjögren's syndrome (SS) and patients with SS who had low-grade or high-grade inflammation and impaired or normal saliva production, to identify microRNA patterns specific to salivary gland inflammation or dysfunction. Methods MicroRNA expression profiles were generated by Agilent microRNA arrays. We developed a novel method for data normalization by identifying housekeeping microRNA. MicroRNA profiles were compared by unsupervised mathematical methods to test how well they distinguish between control subjects and various subsets of patients with SS. Several bioinformatics methods were used to predict the messenger RNA targets of the differentially expressed microRNA. Results MicroRNA expression patterns accurately distinguished salivary glands from control subjects and patients with SS who had low-degree or high-degree inflammation. Using real-time quantitative polymerase chain reaction, we validated 2 microRNA as markers of inflammation in an independent cohort. Comparing microRNA from patients with preserved or low salivary flow identified a set of differentially expressed microRNA, most of which were up-regulated in the group with decreased salivary gland function, suggesting that the targets of microRNA may have a protective effect on epithelial cells. The predicted biologic targets of microRNA associated with inflammation or salivary gland dysfunction identified both overlapping and distinct biologic pathways and processes. Conclusion Distinct microRNA expression patterns are associated with salivary gland inflammation and dysfunction in patients with SS, and microRNA represent a novel group of potential biomarkers. PMID:21280008

  6. Integrative Analysis of MicroRNA and mRNA Data Reveals an Orchestrated Function of MicroRNAs in Skeletal Myocyte Differentiation in Response to TNF-α or IGF1

    PubMed Central

    Meyer, Swanhild U.; Sass, Steffen; Mueller, Nikola S.; Krebs, Stefan; Bauersachs, Stefan; Kaiser, Sebastian; Blum, Helmut; Thirion, Christian; Krause, Sabine; Theis, Fabian J.; Pfaffl, Michael W.

    2015-01-01

    Introduction Skeletal muscle cell differentiation is impaired by elevated levels of the inflammatory cytokine tumor necrosis factor-α (TNF-α) with pathological significance in chronic diseases or inherited muscle disorders. Insulin like growth factor-1 (IGF1) positively regulates muscle cell differentiation. Both, TNF-α and IGF1 affect gene and microRNA (miRNA) expression in this process. However, computational prediction of miRNA-mRNA relations is challenged by false positives and targets which might be irrelevant in the respective cellular transcriptome context. Thus, this study is focused on functional information about miRNA affected target transcripts by integrating miRNA and mRNA expression profiling data. Methodology/Principal Findings Murine skeletal myocytes PMI28 were differentiated for 24 hours with concomitant TNF-α or IGF1 treatment. Both, mRNA and miRNA expression profiling was performed. The data-driven integration of target prediction and paired mRNA/miRNA expression profiling data revealed that i) the quantity of predicted miRNA-mRNA relations was reduced, ii) miRNA targets with a function in cell cycle and axon guidance were enriched, iii) differential regulation of anti-differentiation miR-155-5p and miR-29b-3p as well as pro-differentiation miR-335-3p, miR-335-5p, miR-322-3p, and miR-322-5p seemed to be of primary importance during skeletal myoblast differentiation compared to the other miRNAs, iv) the abundance of targets and affected biological processes was miRNA specific, and v) subsets of miRNAs may collectively regulate gene expression. Conclusions Joint analysis of mRNA and miRNA profiling data increased the process-specificity and quality of predicted relations by statistically selecting miRNA-target interactions. Moreover, this study revealed miRNA-specific predominant biological implications in skeletal muscle cell differentiation and in response to TNF-α or IGF1 treatment. Furthermore, myoblast differentiation-associated mi

  7. Identification of microRNA-regulated pathways using an integration of microRNA-mRNA microarray and bioinformatics analysis in CD34+ cells of myelodysplastic syndromes.

    PubMed

    Xu, Feng; Zhu, Yang; He, Qi; Wu, Ling-Yun; Zhang, Zheng; Shi, Wen-Hui; Liu, Li; Chang, Chun-Kang; Li, Xiao

    2016-01-01

    The effect of microRNA (miRNA) and targeted mRNA on signal transduction is not fully understood in myelodysplastic syndromes (MDS). Here, we tried to identify the miRNAs-regulated pathways through a combination of miRNA and mRNA microarray in CD34+ cells from MDS patients. We identified 34 differentially expressed miRNAs and 1783 mRNAs in MDS. 25 dysregulated miRNAs and 394 targeted mRNAs were screened by a combination of Pearson's correlation analysis and software prediction. Pathway analysis showed that several pathways such as Notch, PI3K/Akt might be regulated by those miRNA-mRNAs pairs. Through a combination of Pathway and miRNA-Gene or GO-Network analysis, miRNAs-regulated pathways, such as miR-195-5p/DLL1/Notch signaling pathway, were identified. Further qRT-PCR showed that miR-195-5p was up-regulated while DLL1 was down-regulated in patients with low-grade MDS compared with normal controls. Luciferase assay showed that DLL1 was a direct target of miR-195-5p. Overexpression of miR-195-5p led to increased cell apoptosis and reduced cell growth through inhibition of Notch signaling pathway. In conclusion, alteration expression of miRNAs and targeted mRNAs might have an important impact on cancer-related cellular pathways in MDS. Inhibition of Notch signaling pathway by miR-195-5p-DLL1 axis contributes to the excess apoptosis in low-grade MDS. PMID:27571714

  8. Identification of microRNA-regulated pathways using an integration of microRNA-mRNA microarray and bioinformatics analysis in CD34+ cells of myelodysplastic syndromes

    PubMed Central

    Xu, Feng; Zhu, Yang; He, Qi; Wu, Ling-Yun; Zhang, Zheng; Shi, Wen-Hui; Liu, Li; Chang, Chun-Kang; Li, Xiao

    2016-01-01

    The effect of microRNA (miRNA) and targeted mRNA on signal transduction is not fully understood in myelodysplastic syndromes (MDS). Here, we tried to identify the miRNAs-regulated pathways through a combination of miRNA and mRNA microarray in CD34+ cells from MDS patients. We identified 34 differentially expressed miRNAs and 1783 mRNAs in MDS. 25 dysregulated miRNAs and 394 targeted mRNAs were screened by a combination of Pearson’s correlation analysis and software prediction. Pathway analysis showed that several pathways such as Notch, PI3K/Akt might be regulated by those miRNA-mRNAs pairs. Through a combination of Pathway and miRNA-Gene or GO-Network analysis, miRNAs-regulated pathways, such as miR-195-5p/DLL1/Notch signaling pathway, were identified. Further qRT-PCR showed that miR-195-5p was up-regulated while DLL1 was down-regulated in patients with low-grade MDS compared with normal controls. Luciferase assay showed that DLL1 was a direct target of miR-195-5p. Overexpression of miR-195-5p led to increased cell apoptosis and reduced cell growth through inhibition of Notch signaling pathway. In conclusion, alteration expression of miRNAs and targeted mRNAs might have an important impact on cancer-related cellular pathways in MDS. Inhibition of Notch signaling pathway by miR-195-5p-DLL1 axis contributes to the excess apoptosis in low-grade MDS. PMID:27571714

  9. MicroRNA-34a regulation of endothelial senescence

    SciTech Connect

    Ito, Takashi; Yagi, Shusuke; Yamakuchi, Munekazu

    2010-08-06

    Research highlights: {yields} MicroRNA-34a (miR-34a) regulates senescence and cell cycle progression in endothelial cells. {yields} MiR-34a expression increases during endothelial cell senescence and in older mice. {yields} SIRT1 is a miR-34a target gene in endothelial cells. {yields} SIRT1 mediates the effects of miR-34a upon cell senescence in endothelial cells. -- Abstract: Endothelial senescence is thought to play a role in cardiovascular diseases such as atherosclerosis. We hypothesized that endothelial microRNAs (miRNAs) regulate endothelial survival and senescence. We found that miR-34a is highly expressed in primary endothelial cells. We observed that miR-34a expression increases in senescent human umbilical cord vein endothelial cells (HUVEC) and in heart and spleen of older mice. MiR-34a over-expression induces endothelial cell senescence and also suppresses cell proliferation by inhibiting cell cycle progression. Searching for how miR-34a affects senescence, we discovered that SIRT1 is a target of miR-34a. Over-expressing miR-34a inhibits SIRT1 protein expression, and knocking down miR-34a enhances SIRT1 expression. MiR-34a triggers endothelial senescence in part through SIRT1, since forced expression of SIRT1 blocks the ability of miR-34a to induce senescence. Our data suggest that miR-34a contributes to endothelial senescence through suppression of SIRT1.

  10. Circulating microRNA-200 Family as Diagnostic Marker in Hepatocellular Carcinoma

    PubMed Central

    Dhayat, Sameer A.; Hüsing, Anna; Senninger, Norbert; Schmidt, Hartmut H.; Haier, Jörg

    2015-01-01

    Goals In this clinical study, we aimed to evaluate the role of circulating microRNA-200 family as a non-invasive tool to identify patients with cirrhosis-associated hepatocellular carcinoma (HCC). Background Prognosis of HCC remains poor with increasing incidence worldwide, mainly related to liver cirrhosis. So far, no reliable molecular targets exist for early detection of HCC at surgically manageable stages. Recently, we identified members of the microRNA-200 family as potential diagnostic markers of cirrhosis-associated HCC in patient tissue samples. Their value as circulating biomarkers for HCC remained undefined. Methods Blood samples and clinicopathological data of consecutive patients with liver diseases were collected prospectively. Expression of the microRNA-200 family was investigated by qRT-PCR in blood serum samples of 22 HCC patients with and without cirrhosis. Serum samples of patients with non-cancerous chronic liver cirrhosis (n = 22) and of healthy volunteers (n = 15) served as controls. Results MicroRNA-141 and microRNA-200a were significantly downregulated in blood serum of patients with HCC compared to liver cirrhosis (p<0.007) and healthy controls (p<0.002). MicroRNA-141 and microRNA-200a could well discriminate patients with cirrhosis-associated HCC from healthy volunteers with area under the receiver-operating characteristic curve (AUC) values of 0.85 and 0.82, respectively. Additionally, both microRNAs could differentiate between HCC and non-cancerous liver cirrhosis with a fair accuracy. Conclusions Circulating microRNA-200 family members are significantly deregulated in patients with HCC and liver cirrhosis. Further studies are necessary to confirm the diagnostic value of the microRNA-200 family as accurate serum marker for cirrhosis-associated HCC. PMID:26447841

  11. Three-Dimensional Characterization of Cell Clusters Using Synchrotron-Radiation-Based Micro-Computed Tomography

    NASA Astrophysics Data System (ADS)

    Müller, Ert; Riedel, Marco; Thurner, Philipp J.

    2006-04-01

    Micro-computed tomography with the highly intense, monochromatic X rays produced by the synchrotron is a superior method to nondestructively measure the local absorption in three-dimensional space. Because biological tissues and cells consist mainly of water as the surrounding medium, higher absorbing agents have to be incorporated into the structures of interest. Even without X-ray optics such as refractive lens, one can uncover the stain distribution with the spatial resolution of about 1 [mu]m. Incorporating the stain at selected cell compartments, for example, binding to the RNA/DNA, their density distribution becomes quantified. In this communication, we demonstrate that tomograms obtained at the beamlines BW2 and W2 (HASYLAB at DESY, Hamburg, Germany) and 4S (SLS, Villigen, Switzerland) clearly show that the RNA/DNA-stained HEK 293 cell clusters have a core of high density and a peripheral part of lower density, which correlate with results of optical microscopy. The inner part of the clusters is associated with nonvital cells as the result of insufficient oxygen and nutrition supply. This necrotic part is surrounded by (6 ± 1) layers of vital cells.

  12. Similar Squamous Cell Carcinoma Epithelium microRNA Expression in Never Smokers and Ever Smokers

    PubMed Central

    Kolokythas, Antonia; Zhou, Yalu; Schwartz, Joel L.; Adami, Guy R.

    2015-01-01

    The incidence of oral tumors in patients who never used mutagenic agents such as tobacco is increasing. In an effort to better understand these tumors we studied microRNA (miRNA) expression in tumor epithelium of never tobacco users, tumor epithelium of ever tobacco users, and nonpathological control oral epithelium. A comparison of levels among 372 miRNAs in 12 never tobacco users with oral squamous cell carcinoma (OSCC) versus 10 healthy controls was made using the reverse transcription quantitative polymerase chain reaction. A similar analysis was done with 8 ever tobacco users with OSCC. These comparisons revealed miR-10b-5p, miR-196a-5p, and miR-31-5p as enriched in the tumor epithelium in OSCC of both never and ever tobacco users. Examination of The Cancer Genome Atlas (TCGA) project miRNA data on 305 OSCCs and 30 controls revealed 100% of those miRNAs enriched in never smoker OSCCs in this patient group were also enriched in ever smoker OSCCs. Nonsupervised clustering of TCGA OSCCs was suggestive of two or four subgroups of tumors based on miRNA levels with limited evidence for differences in tobacco exposure among the groups. Results from both patient groups together stress the importance of miR196a-5p in OSCC malignancy in both never and ever smokers, and emphasize the overall similarity of miRNA expression in OSCCs in these two risk groups. It implies that there may be great similarity in etiology of OSCC in never and ever smokers and that classifying OSCC based on tobacco exposure may not be helpful in the clinic. PMID:26544609

  13. MicroRNA-mediated target mRNA cleavage and 3′-uridylation in human cells

    PubMed Central

    Xu, Kai; Lin, Jing; Zandi, Roza; Roth, Jack A.; Ji, Lin

    2016-01-01

    MicroRNAs (miRNAs) play an important role in targeted gene silencing by facilitating posttranscriptional and translational repression. However, the precise mechanism of mammalian miRNA-mediated gene silencing remains to be elucidated. Here, we used a stem-loop array reverse-transcription polymerase chain reaction assay to analyse miRNA-induced mRNA recognition, cleavage, posttranscriptional modification, and degradation. We detected endogenous let-7 miRNA-induced and Argonaute-catalysed endonucleolytic cleavage on target mRNAs at various sites within partially paired miRNA:mRNA sequences. Most of the cleaved mRNA 5′-fragments were 3′-oligouridylated by activities of terminal uridylyl transferases (TUTases) in miRNA-induced silencing complexes and temporarily accumulated in the cytosol for 5′-3′ degradation or other molecular fates. Some 3′-5′ decayed mRNA fragments could also be captured by the miRNA-induced silencing complex stationed at the specific miRNA:mRNA target site and oligouridylated by other TUTases at its proximity without involving Argonaute-mediated RNA cleavage. Our findings provide new insights into the molecular mechanics of mammalian miRNA-mediated gene silencing by coordinated target mRNA recognition, cleavage, uridylation and degradation. PMID:27440378

  14. TWIST1-induced microRNA-424 reversibly drives mesenchymal programming while inhibiting tumor initiation

    PubMed Central

    Drasin, David J.; Guarnieri, Anna L.; Neelakantan, Deepika; Kim, Jihye; Cabrera, Joshua H.; Wang, Chu-An; Zaberezhnyy, Vadym; Gasparini, Pierluigi; Cascione, Luciano; Huebner, Kay; Tan, Aik-Choon; Ford, Heide L.

    2015-01-01

    Epithelial-to-mesenchymal transition (EMT) is a dynamic process that relies on cellular plasticity. Recently, the process of an oncogenic EMT, followed by a reverse mesenchymal-to-epithelial transition (MET), has been implicated as critical in the metastatic colonization of carcinomas. Unlike governance of epithelial programming, regulation of mesenchymal programming is not well understood in EMT. Here, we describe and characterize the first microRNA that enhances exclusively mesenchymal programming. We demonstrate that microRNA-424 is upregulated early during a TWIST1 or SNAI1-induced EMT, and that it causes cells to express mesenchymal genes without affecting epithelial genes, resulting in a mixed/intermediate EMT. Furthermore, microRNA-424 increases motility, decreases adhesion and induces a growth arrest, changes associated with a complete EMT, that can be reversed when microRNA-424 expression is lowered, concomitant with an MET-like process. Breast cancer patient microRNA-424 levels positively associate with TWIST1/2 and EMT-like gene signatures, and miR-424 is increased in primary tumors versus matched normal breast. However, microRNA-424 is downregulated in patient metastases versus matched primary tumors. Correspondingly, microRNA-424 decreases tumor initiation and is post-transcriptionally downregulated in macrometastases in mice, suggesting the need for biphasic expression of miR-424 to transit the EMT-MET axis. Next-generation RNA sequencing revealed microRNA-424 regulates numerous EMT and cancer stemness-associated genes, including TGFBR3, whose downregulation promotes mesenchymal phenotypes, but not tumor-initiating phenotypes. Instead, we demonstrate that increased MAPK/ERK signaling is critical for miR-424-mediated decreases in tumor-initiating phenotypes. These findings suggest microRNA-424 plays distinct roles in tumor progression, potentially facilitating earlier, but repressing later, stages of metastasis by regulating an EMT-MET axis. PMID

  15. Circulating microRNA Profiles during the Bovine Oestrous Cycle

    PubMed Central

    Ioannidis, Jason; Donadeu, F. Xavier

    2016-01-01

    Up to 50% of ovulations go undetected in modern dairy herds due to attenuated oestrus behavior and a lack of high-accuracy methods for detection of fertile oestrus. This significantly reduces overall herd productivity and constitutes a high economic burden to the dairy industry. MicroRNAs (miRNAs) are ubiquitous regulators of gene expression during both health and disease and they have been shown to regulate different reproductive processes. Extracellular miRNAs are stable and can provide useful biomarkers of tissue function; changes in circulating miRNA profiles have been reported during menstrual cycles. This study sought to establish the potential of circulating miRNAs as biomarkers of oestrus in cattle. We collected plasma samples from 8 Holstein-Friesian heifers on days Days 0, 8 and 16 of an oestrous cycle and analysed small RNA populations on each Day using two independent high-throughput approaches, namely, Illumina sequencing (n = 24 samples) and Qiagen PCR arrays (n = 9 sample pools, 3–4 samples / pool). Subsequently, we used RT-qPCR (n = 24 samples) to validate the results of high-throughput analyses, as well as to establish the expression profiles of additional miRNAs previously reported to be differentially expressed during reproductive cycles. Overall, we identified four miRNAs (let-7f, miR-125b, miR-145 and miR-99a-5p), the plasma levels of which distinctly increased (up to 2.2-fold, P < 0.05) during oestrus (Day 0) relative to other stages of the cycle (Days 8 and 16). Moreover, we identified several hundred different isomiRs and established their relative abundance in bovine plasma. In summary, our results reveal the dynamic nature of plasma miRNAs during the oestrous cycle and provide evidence of the feasibility of using circulating miRNAs as biomarkers of reproductive function in livestock in the future. PMID:27340826

  16. Characterization of the rainbow trout egg microRNA transcriptome.

    PubMed

    Ma, Hao; Hostuttler, Mark; Wei, Hairong; Rexroad, Caird E; Yao, Jianbo

    2012-01-01

    MicroRNAs (miRNAs) are a class of endogenous small non-coding RNA molecules that regulate post-transcriptional expression of target genes and play important roles in animal development. The objectives of this study were to characterize the egg miRNA transcriptome and identify novel egg-predominant miRNAs in rainbow trout. Small RNAs isolated from mature unfertilized rainbow trout eggs were subjected to deep sequencing using an Illumina Genome Analyzer. The massive sequencing produced 24,621,741 quality reads, among which, 266 known miRNAs were identified and 230 putatively novel miRNAs were predicted. The most abundantly known miRNAs are let-7 and miR-21, accounting for 24.06% and 18.71% of the known miRNAs, respectively. Other known miRNAs which are abundantly present in eggs include miR-24, miR-202, miR-148, miR-30, miR-10, miR-146, miR-25, and miR-143. Real time PCR analysis using cDNAs derived from 10 tissues validated 87 out of 90 selected putative miRNAs and identified three novel miRNAs predominantly expressed in rainbow trout eggs. Each of these novel egg-predominant miRNAs is predicted to target a significant number of genes, most of which are significantly down-regulated in naturally ovulated rainbow trout eggs based on analysis of publicly available microarray data sets. Quantitative real time PCR analysis also demonstrated low expression of a selected number of target genes in eggs relative to liver and muscle tissues. This study represents the first complete survey of miRNAs in fish eggs and provides a starting point for future studies aimed at understanding the roles of miRNAs in controlling egg quality and early embryogenesis in rainbow trout. PMID:22761856

  17. Polymerase II Promoter Strength Determines Efficacy of microRNA Adapted shRNAs

    PubMed Central

    Lebbink, Robert Jan; Lowe, Maggie; Chan, Theresa; Khine, Htet; Wang, Xiaoyin; McManus, Michael T.

    2011-01-01

    Since the discovery of RNAi and microRNAs more than 10 years ago, much research has focused on the development of systems that usurp microRNA pathways to downregulate gene expression in mammalian cells. One of these systems makes use of endogenous microRNA pri-cursors that are expressed from polymerase II promoters where the mature microRNA sequence is replaced by gene specific duplexes that guide RNAi (shRNA-miRs). Although shRNA-miRs are effective in directing target mRNA knockdown and hence reducing protein expression in many cell types, variability of RNAi efficacy in cell lines has been an issue. Here we show that the choice of the polymerase II promoter used to drive shRNA expression is of critical importance to allow effective mRNA target knockdown. We tested the abundance of shRNA-miRs expressed from five different polymerase II promoters in 6 human cell lines and measured their ability to drive target knockdown. We observed a clear positive correlation between promoter strength, siRNA expression levels, and protein target knockdown. Differences in RNAi from the shRNA-miRs expressed from the various promoters were particularly pronounced in immune cells. Our findings have direct implications for the design of shRNA-directed RNAi experiments and the preferred RNAi system to use for each cell type. PMID:22031824

  18. MicroRNA overexpression increases cortical neuronal vulnerability to injury

    PubMed Central

    Truettner, Jessie S.; Motti, Dario; Dietrich, W. Dalton

    2013-01-01

    Previously we reported that several microRNAs (miRNA) are upregulated following experimentally induced traumatic brain injury (TBI) using both in vivo and in vitro approaches. Specific miRNAs were found to be sensitive to therapeutic hypothermia and may therefore be important targets for neuroprotective strategies. In this study we developed plasmid constructs that overexpress temperature sensitive miRNAs: miR-34a, miR-451, and miR-874. These constructs were transfected into cultured cortical neurons that were subjected to stretch injury using a cell injury controller device. Levels of expression of genes associated with stress, inflammation, apoptosis and transcriptional regulation were measured by qRT-PCR. mRNA levels of cytokines interleukin 1-β (IL1-β) and tumor necrosis factor alpha (TNF-α) as well as heat shock protein 70 (HSP70) and Caspase 11 were found to be increased up to 24 fold higher than controls in cells overexpressing these miRNAs. After moderate stretch injury, the expression of IL1-β, TNF-α, HSP70 and Caspase 11 all increased over control levels found in uninjured cells suggesting that overexpression of these miRNAs increases cellular vulnerability. miR-34a directly inhibits Bcl2 and XIAP, both anti-apoptotic proteins. The observed increase in Caspase 11 with over-expression of miR-34a indicates that miR-34a may be inducing apoptosis by reducing the levels of antiapoptotic proteins. miR-34a is predicted to inhibit Jun, which was seen to decrease in cells overexpressing this miRNA along with Fos. Over expression of several miRNAs found to be induced by TBI in vivo (miR-34a, miR-451 and miR-874) leads to increased vulnerability in transfected neurons. Therapeutic hypothermia blunts the expression of these miRNAs in vivo and antisense silencing could be a potential therapeutic approach to targeting the consequences of TBI. PMID:23948100

  19. Skeletal Micro-RNA Responses to Simulated Weightlessness

    NASA Technical Reports Server (NTRS)

    Thomas, Nicholas J.; Choi, Catherine Y.; Alwood, Joshua S.

    2016-01-01

    Astronauts lose bone structure during long-duration spaceflight. These changes are due, in part, to insufficient bone formation by the osteoblast cells. Little is known about the role that small (approximately 22 nucleotides), non-coding micro-RNAs (miRNAs) play in the osteoblast response to microgravity. We hypothesize that osteoblast-lineage cells alter their miRNA status during microgravity exposure, contributing to impaired bone formation during weightlessness. To simulate weightlessness, female mice (C57BL/6, Charles River, 10 weeks of age, n = 7) were hindlimb unloaded up to 12 days. Age-matched and normally ambulating mice served as controls (n=7). To assess the expression of miRNAs in skeletal tissue, the tibia was collected ex vivo and cleaned of soft-tissue and marrow. Total RNA was collected from tibial bone and relative abundance was measured for miRNAs of interest using quantitative real time PCR array looking at 372 unique and well-characterized mature miRNAs using the delta-delta Ct method. Transcripts of interest were normalized to an average of 6 reference RNAs. Preliminary results show that hindlimb unloading decreased the expression of 14 miRNAs to less than 0.5 times that of the control levels and increased the expression of 5 miRNAs relative to the control mice between 1.2-1.5-fold (p less than 0.05, respectively). Using the miRSystem we assessed overlapping target genes predicted to be regulated by multiple members of the 19 differentially expressed miRNAs as well as in silico predicted targets of our individual miRNAs. Our miRsystem results indicated that a number of our differentially expressed miRNAs were regulators of genes related to the Wnt-Beta Catenin pathway-a known regulator of bone health-and, interestingly, the estrogen-mediated cell-cycle regulation pathway, which may indicate that simulated weightlessness modulated systemic hormonal levels or hormonal transduction that additionally contributed to bone loss. We plan to follow up

  20. Arsenic exposure triggers a shift in microRNA expression.

    PubMed

    Sturchio, Elena; Colombo, Teresa; Boccia, Priscilla; Carucci, Nicoletta; Meconi, Claudia; Minoia, Claudio; Macino, Giuseppe

    2014-02-15

    Exposure to inorganic Arsenic (iAs) through drinking water is a major public health problem affecting most countries. iAs has been classified by the International Agency for Research on Cancer as Group 1: "Carcinogenic to humans". Although numerous studies have shown the related adverse effects of iAs, sensitive appropriate biomarkers for studies of environmental epidemiology are still required. The present work aims at investigate the role of microRNAs (miRNAs), powerful negative regulators of gene expression, playing a key role in many physiological and pathological cellular processes, in iAs exposure. To this end, we analyzed miRNA changes in expression profile triggered by iAs exposure in Jurkat cell line. We used microarray technology to profile the expression of miRNAs following 2 μmol/L sodium arsenite treatment at different time points. Moreover, we performed phenotypic analysis of iAs treated cells. Real Time Polymerase Chain Reaction (RT-PCR) was used to validate miRNA microarray data and to assay expression modulation of selected relevant mRNAs. Finally, bioinformatics techniques were applied to reconstruct iAs-relevant molecular pathways and miRNA regulatory networks from the expression data. We report miRNAs modulated after iAs treatment in Jurkat cells. In particular, we highlight 36 miRNAs exhibiting consistent dysregulation and particularly a panel of 8 miRNAs which we also validated by RT-PCR analysis. Computational analysis of lists of putative target genes for these 8 miRNAs points to an involvement in arsenic-response pathways, for a subset of them, that were analyzed by RT-PCR. Furthermore, iAs exposure reveals induction of cell cycle progression and the failure of apoptosis, supporting the idea of iAs carcinogenic activity. Our study provides a list of miRNAs whose expression levels are affected by iAs treatment, corroborating the importance of proceeding with the hunt for specific subset of miRNAs, which can serve as potential biomarkers of

  1. Next-generation sequencing of the porcine skeletal muscle transcriptome for computational prediction of microRNA gene targets

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNA are a class of small RNAs that regulate gene expression by inhibiting translation of protein encoding transcripts. Inhibition is exerted through targeting of a microRNA-protein complex by base-pairing of the microRNA sequence to cognate recognition sequences in the 3’ untranslated region (...

  2. The microRNA feedback regulation of p63 in cancer progression

    PubMed Central

    Lin, Changwei; Li, Xiaorong; Zhang, Yi; Guo, Yihang; Zhou, Jianyu; Gao, Kai; Dai, Jing; Hu, Gui; Lv, Lv; Du, Juan; Zhang, Yi

    2015-01-01

    The transcription factor p63 is a member of the p53 gene family that plays a complex role in cancer due to its involvement in epithelial differentiation, cell cycle arrest and apoptosis. MicroRNAs are a class of small, non-coding RNAs with an important regulatory role in various cellular processes, as well as in the development and progression of cancer. A number of microRNAs have been shown to function as transcriptional targets of p63. Conversely, microRNAs also can modulate the expression and activity of p63. However, the p63–microRNA regulatory circuit has not been addressed in depth so far. Here, computational genomic analysis was performed using miRtarBase, Targetscan, microRNA.ORG, DIANA-MICROT, RNA22-HSA and miRDB to analyze miRNA binding to the 3′UTR of p63. JASPAR (profile score threshold 80%) and TFSEARCH datasets were used to search transcriptional start sites for p53/p63 response elements. Remarkably, these data revealed 63 microRNAs that targeted p63. Furthermore, there were 39 microRNAs targeting p63 that were predicted to be regulated by p63. These analyses suggest a crosstalk between p63 and microRNAs. Here, we discuss the crosstalk between p63 and the microRNA network, and the role of their interactions in cancer. PMID:25726529

  3. Microarray based analysis of gene regulation by microRNA in intervertebral disc degeneration

    PubMed Central

    HU, PENG; FENG, BO; WANG, GUANGLIN; NING, BIN; JIA, TANGHONG

    2015-01-01

    The present study aimed to explore the underlying mechanism of the development of intervertebral disc degeneration (IDD) by bioinformatics based on microarray datasets. GSE 19943 and GSE 34095 datasets downloaded from Gene Expression Omnibus data were used to screen the differentially expressed genes (DEGs) in IDD. The correlation between microRNAs and target genes was investigated using different algorithms. The underlying molecular mechanisms of the target genes were then explored using Kyoto Encyclopedia of Genes and Genomes pathway and Gene Ontology function enrichment analysis. A total of 9 differentially expressed microRNAs, including 3 down- and 6 upregulated microRNAs and 850 DEGs were identified in tissue from patients with IDD. Two regulation networks of the target genes by microRNAs were constructed, including 33 upregulated microRNA-target gene pairs and 4 downregulated microRNA-target gene pairs. Certain target genes had been demonstrated to be involved in IDD progression via various pathways, including in the cell cycle and pathways in cancer. In addition, two important microRNAs (microRNA-222 and microRNA-589) were identified that were pivotal for the development of IDD, and their target genes, CDKNAB and SMAD4. In conclusion, a comprehensive miRNA-target gene regulatory network was constructed, which was found to be important in IDD progression. PMID:26134418

  4. microRNA Profiling of Amniotic Fluid: Evidence of Synergy of microRNAs in Fetal Development

    PubMed Central

    Li, Tianpeng; Ling, Shucai

    2016-01-01

    Amniotic fluid (AF) continuously exchanges molecules with the fetus, playing critical roles in fetal development especially via its complex components. Among these components, microRNAs are thought to be transferred between cells loaded in microvesicles. However, the functions of AF microRNAs remain unknown. To date, few studies have examined microRNAs in amniotic fluid. In this study, we employed miRCURY Locked Nucleotide Acid arrays to profile the dynamic expression of microRNAs in AF from mice on embryonic days E13, E15, and E17. At these times, 233 microRNAs were differentially expressed (p< 0.01), accounting for 23% of the total Mus musculus microRNAs. These differentially-expressed microRNAs were divided into two distinct groups based on their expression patterns. Gene ontology analysis showed that the intersectional target genes of these differentially-expressed microRNAs were mainly distributed in synapse, synaptosome, cell projection, and cytoskeleton. Pathway analysis revealed that the target genes of the two groups of microRNAs were synergistically enriched in axon guidance, focal adhesion, and MAPK signaling pathways. MicroRNA-mRNA network analysis and gene- mapping showed that these microRNAs synergistically regulated cell motility, cell proliferation and differentiation, and especially the axon guidance process. Cancer pathways associated with growth and proliferation were also enriched in AF. Taken together, the results of this study are the first to show the functions of microRNAs in AF during fetal development, providing novel insights into interpreting the roles of AF microRNAs in fetal development. PMID:27166676

  5. Clustering siRNA conjugates for MMP-responsive therapeutics in chronic wounds of diabetic animals

    NASA Astrophysics Data System (ADS)

    Kim, Hye Sung; Son, Young Ju; Yoo, Hyuk Sang

    2016-07-01

    The MMP-responsive breakdown of siRNA clusters was translated to site-specific gene transfection and enhanced wound healing in diabetic ulcers. MMP-2 siRNA was chemically tethered to the end of multi-armed PEG via MMP-cleavable linkers (4PEG-siRNA) and subsequently clustered into submicron particles complexed with LPEI. 4PEG-siRNA was more tightly complexed with LPEI and the associated cluster showed higher resistance against RNase attack, in comparison to naked siRNA. Because the size of the clusters increased depending on the increase in charge ratio of LPEI to siRNA, cellular uptake of the 4PEG-siRNA/LPEI cluster was significantly attenuated due to the huge size of the cluster. However, upon MMP treatment, the cluster dissociated into smaller particles and was efficiently endocytosed by cells. An in vivo fluorescence resonance energy transfer (FRET) study also revealed that the clusters were effectively dissociated in MMP-rich environments of dorsal wounds in diabetic animals. In addition, diabetic ulcers treated with the clusters showed a faster wound closure rate and the recovered tissue expressed a larger amount of cytokeratin along with a lower expression level of MMP-2 compared to the other groups.The MMP-responsive breakdown of siRNA clusters was translated to site-specific gene transfection and enhanced wound healing in diabetic ulcers. MMP-2 siRNA was chemically tethered to the end of multi-armed PEG via MMP-cleavable linkers (4PEG-siRNA) and subsequently clustered into submicron particles complexed with LPEI. 4PEG-siRNA was more tightly complexed with LPEI and the associated cluster showed higher resistance against RNase attack, in comparison to naked siRNA. Because the size of the clusters increased depending on the increase in charge ratio of LPEI to siRNA, cellular uptake of the 4PEG-siRNA/LPEI cluster was significantly attenuated due to the huge size of the cluster. However, upon MMP treatment, the cluster dissociated into smaller particles and was

  6. Enhanced hepatic delivery of siRNA and microRNA using oleic acid based lipid nanoparticle formulations

    PubMed Central

    Wang, Xinmei; Yu, Bo; Ren, Wei; Mo, Xiaokui; Zhou, Chenguang; He, Hongyan; Jia, HuLiang; Wang, Lu; Jacob, Samson T.; Lee, Robert J.; Ghoshal, Kalpana; Lee, L. James

    2015-01-01

    Many cationic lipids have been developed for lipid-based nanoparticles (LNPs) for delivery of siRNA and microRNA (miRNA). However, less attention has been paid to “helper lipids”. Here, we investigated several “helper lipids” and examined their effects on the physicochemical properties such as particle size and zeta potential, as well as cellular uptake and transfection efficiency. We found that inclusion of oleic acid (OA), an unsaturated fatty acid; into the LNP formulation significantly enhanced the delivery efficacy for siRNA and miRNA. For proof-of-concept, miR-122, a liver-specific microRNA associated with many liver diseases, was used as a model agent to demonstrate the hepatic delivery efficacy both in tumor cells and in animals. Compared to Lipofectamine 2000, a commercial transfection agent, OA containing LNPs delivered microRNA-122 in a more efficient manner with a 1.8-fold increase in mature miR-122 expression and a 20% decrease in Bcl-w, a target of microRNA-122. In comparison with Invivofectamine, a commercial transfection agent specifically designed for hepatic delivery, OA containing LNPs showed comparable liver accumulation and in vivo delivery efficiency. These findings demonstrated the importance of “helper lipid” components of the LNP formulation on the cellular uptake and transfection activity of siRNA and miRNA. OA containing LNPs are a promising nanocarrier system for the delivery of RNA-based therapeutics in liver diseases. PMID:24121065

  7. MicroRNA Targeted Therapeutic Approach for Pancreatic Cancer

    PubMed Central

    Li, Yiwei; Sarkar, Fazlul H.

    2016-01-01

    Pancreatic cancer remains the fourth leading cause of cancer-related death in the US and is expected to be the second leading cause of cancer-related death by 2030. Therefore, it is important to better understand the molecular pathogenesis, phenotypes and features of pancreatic cancer in order to design novel molecularly targeted therapies for achieving better therapeutic outcome of patients with pancreatic cancer. Recently, the roles of microRNAs (miRNAs) in the development and progression of pancreatic cancer became a hot topic in the scientific community of pancreatic cancer research. By conducting miRNA expression profiling, the aberrant expression of miRNAs was revealed in the serum and in cancer tissues from patients with pancreatic cancer. These aberrantly expressed miRNAs are critically correlated with the disease stage, drug resistance, and survival of pancreatic cancer patients. Hence, targeting these tiny molecules, the specific miRNAs, could provide an efficient and optimal approach in the therapy of pancreatic cancer. Indeed, the pre-clinical and in vivo experiments showed that nanoparticle delivery of synthetic oligonucleotides or treatment with natural agents could be useful to modulate the expression of miRNAs and thereby inhibit pancreatic cancer growth and progression, suggesting that targeting miRNAs combined with conventional anti-cancer therapeutics could be a novel therapeutic strategy for increasing drug sensitivity and achieving better therapeutic outcome of patients diagnosed with pancreatic cancer. PMID:26929739

  8. microRNA modulation of circadian clock period and entrainment

    PubMed Central

    Cheng, Hai-Ying M.; Papp, Joseph W.; Varlamova, Olga; Dziema, Heather; Russell, Brandon; Curfman, John P.; Nakazawa, Takanobu; Shimizu, Kimiko; Okamura, Hitoshi; Impey, Soren; Obrietan, Karl

    2007-01-01

    microRNAs (miRNAs) are a class of small, non-coding, RNAs that regulate the stability or translation of mRNA transcripts. Although recent work has implicated miRNAs in development and in disease, the expression and function of miRNAs in the adult mammalian nervous system has not been extensively characterized. Here, we examine the role of two brain-specific miRNAs, miR-219 and miR-132, in modulating the circadian clock located in the suprachiasmatic nucleus. miR-219 is a target of the CLOCK/BMAL1 complex, exhibits robust circadian rhythms of expression and the in vivo knockdown of miR-219 lengthens the circadian period. miR-132 is induced by photic entrainment cues via a MAPK/CREB-dependent mechanism, modulates clock gene expression, and attenuates the entraining effects of light. Collectively, these data reveal miRNAs as clock- and light-regulated genes and provide a mechanistic examination of their roles as effectors of pacemaker activity and entrainment. PMID:17553428

  9. The Lupus Autoantigen La Prevents Mis-channeling of tRNA Fragments into the Human MicroRNA Pathway.

    PubMed

    Hasler, Daniele; Lehmann, Gerhard; Murakawa, Yasuhiro; Klironomos, Filippos; Jakob, Leonhard; Grässer, Friedrich A; Rajewsky, Nikolaus; Landthaler, Markus; Meister, Gunter

    2016-07-01

    The Lupus autoantigen La is an RNA-binding protein that stabilizes RNA polymerase III (Pol III) transcripts and supports RNA folding and has in addition been implicated in the mammalian microRNA (miRNA) pathway. Here, we have analyzed effects of La depletion on Argonaute (Ago)-bound small RNAs in human cells. We find that in the absence of La, distinct tRNA fragments are loaded into Ago proteins. Thus, La functions as gatekeeper ensuring correct tRNA maturation and protecting the miRNA pathway from potentially functional tRNA fragments. However, one specific isoleucin pre-tRNA produces both a functional tRNA and a miRNA even when La is present. We demonstrate that the fully complementary 5' leader and 3' trailer of the pre-tRNA-Ile form a double-stranded RNA molecule that has low affinity to La. Instead, Exportin-5 (Xpo5) recognizes it as miRNA precursor and transports it into the cytoplasm for Dicer processing and Ago loading. PMID:27345152

  10. Smoking alters circulating plasma microvesicle pattern and microRNA signatures.

    PubMed

    Badrnya, S; Baumgartner, R; Assinger, A

    2014-07-01

    Circulating plasma microvesicles (PMVs) and their microRNA content are involved in the development of atherosclerosis and could serve as biomarkers for cardiovascular disease (CVD) progression. However, little is known on how smoking influences the levels of PMVs and microRNA signatures in vivo. Therefore, we aimed to investigate the effects of smoking on circulating PMV levels and CVD-related PMV-derived microRNAs in young, healthy smokers. Twenty young (10 female, 10 male; 25 ± 4 years) healthy smokers (16 ± 6 cigarettes per day for 8 ± 4 years) and age- and sex-matched controls were included in this study. While complete blood count revealed no differences between both groups, smoking significantly enhanced intracellular reactive oxygen species in platelets and leukocytes as well as platelet-leukocyte aggregate formation. Total circulating PMV counts were significantly reduced in smokers, which could be attributed to decreased platelet-derived PMVs. While the number of endothelial PMVs remained unaffected, smoking propagated circulating leukocyte-derived PMVs. Despite reduced total PMVs, PMV-derived microRNA-profiling of six smoker/control pairs revealed a decrease of only a single microRNA, the major platelet-derived microRNA miR-223. Conversely, miR-29b, a microRNA associated with aortic aneurysm and fibrosis, and RNU6-2, a commonly used reference-RNA, were significantly up-regulated. Smoking leads to alterations in the circulating PMV profile and changes in the PMV-derived microRNA signature already in young, healthy adults. These changes may contribute to the development of smoking-related cardiovascular pathologies. Moreover, these smoking-related changes have to be considered when microRNA or PMV profiles are used as disease-specific biomarkers. PMID:24573468

  11. MicroRNA Expression Profiling of Oligodendrocyte Differentiation from Human Embryonic Stem Cells

    PubMed Central

    Letzen, Brian S.; Liu, Cyndi; Thakor, Nitish V.; Gearhart, John D.; All, Angelo H.; Kerr, Candace L.

    2010-01-01

    Background Cells of the oligodendrocyte (OL) lineage play a vital role in the production and maintenance of myelin, a multilamellar membrane which allows for saltatory conduction along axons. These cells may provide immense therapeutic potential for lost sensory and motor function in demyelinating conditions, such as spinal cord injury, multiple sclerosis, and transverse myelitis. However, the molecular mechanisms controlling OL differentiation are largely unknown. MicroRNAs (miRNAs) are considered the “micromanagers” of gene expression with suggestive roles in cellular differentiation and maintenance. Although unique patterns of miRNA expression in various cell lineages have been characterized, this is the first report documenting their expression during oligodendrocyte maturation from human embryonic stem (hES) cells. Here, we performed a global miRNA analysis to reveal and identify characteristic patterns in the multiple stages leading to OL maturation from hES cells including those targeting factors involved in myelin production. Methodology/Principal Findings We isolated cells from 8 stages of OL differentiation. Total RNA was subjected to miRNA profiling and validations preformed using real-time qRT-PCR. A comparison of miRNAs from our cultured OLs and OL progenitors showed significant similarities with published results from equivalent cells found in the rat and mouse central nervous system. Principal component analysis revealed four main clusters of miRNA expression corresponding to early, mid, and late progenitors, and mature OLs. These results were supported by correlation analyses between adjacent stages. Interestingly, the highest differentially-expressed miRNAs demonstrated a similar pattern of expression throughout all stages of differentiation, suggesting that they potentially regulate a common target or set of targets in this process. The predicted targets of these miRNAs include those with known or suspected roles in oligodendrocyte development

  12. EpCAM knockdown alters microRNA expression in retinoblastoma--functional implication of EpCAM regulated miRNA in tumor progression.

    PubMed

    Beta, Madhu; Khetan, Vikas; Chatterjee, Nivedita; Suganeswari, Ganesan; Rishi, Pukhraj; Biswas, Jyotirmay; Krishnakumar, Subramanian

    2014-01-01

    The co-ordinated regulation of oncogenes along with miRNAs play crucial role in carcinogenesis. In retinoblastoma (RB), several miRNAs are known to be differentially expressed. Epithelial cell adhesion molecule (EpCAM) gene is involved in many epithelial cancers including, retinoblastoma (RB) tumorigenesis. EpCAM silencing effectively reduces the oncogenic miR-17-92 cluster. In order to investigate whether EpCAM has wider effect as an inducer or silencer of miRNAs, we performed a global microRNA expression profile in EpCAM siRNA knockdown Y79 cells. MicroRNA profiling in EpCAM silenced Y79 cells showed seventy-three significantly up regulated and thirty-six down regulated miRNAs. A subset of these miRNAs was also validated in tumors. Functional studies on Y79 and WERI-Rb-1 cells transfected with antagomirs against two miRNAs of miR-181c and miR-130b showed striking changes in tumor cell properties in RB cells. Treatment with anti-miR-181c and miR-130b showed significant decrease in cell viability and cell invasion. Increase in caspase-3 level was noticed in antagomir transfected cell lines indicating the induction of apoptosis. Possible genes altered by EpCAM influenced microRNAs were predicted by bioinformatic tools. Many of these belong to pathways implicated in cancer. The study shows significant influence of EpCAM on global microRNA expression. EpCAM regulated miR-181c and miR-130b may play significant roles in RB progression. EpCAM based targeted therapies may reduce carcinogenesis through several miRNAs and target genes. PMID:25502397

  13. EpCAM Knockdown Alters MicroRNA Expression in Retinoblastoma- Functional Implication of EpCAM Regulated MiRNA in Tumor Progression

    PubMed Central

    Beta, Madhu; Khetan, Vikas; Chatterjee, Nivedita; Suganeswari, Ganesan; Rishi, Pukhraj; Biswas, Jyotirmay; Krishnakumar, Subramanian

    2014-01-01

    The co-ordinated regulation of oncogenes along with miRNAs play crucial role in carcinogenesis. In retinoblastoma (RB), several miRNAs are known to be differentially expressed. Epithelial cell adhesion molecule (EpCAM) gene is involved in many epithelial cancers including, retinoblastoma (RB) tumorigenesis. EpCAM silencing effectively reduces the oncogenic miR-17-92 cluster. In order to investigate whether EpCAM has wider effect as an inducer or silencer of miRNAs, we performed a global microRNA expression profile in EpCAM siRNA knockdown Y79 cells. MicroRNA profiling in EpCAM silenced Y79 cells showed seventy-three significantly up regulated and thirty-six down regulated miRNAs. A subset of these miRNAs was also validated in tumors. Functional studies on Y79 and WERI-Rb-1 cells transfected with antagomirs against two miRNAs of miR-181c and miR-130b showed striking changes in tumor cell properties in RB cells. Treatment with anti-miR-181c and miR-130b showed significant decrease in cell viability and cell invasion. Increase in caspase-3 level was noticed in antagomir transfected cell lines indicating the induction of apoptosis. Possible genes altered by EpCAM influenced microRNAs were predicted by bioinformatic tools. Many of these belong to pathways implicated in cancer. The study shows significant influence of EpCAM on global microRNA expression. EpCAM regulated miR-181c and miR-130b may play significant roles in RB progression. EpCAM based targeted therapies may reduce carcinogenesis through several miRNAs and target genes. PMID:25502397

  14. Diagnostic value of a plasma microRNA signature in gastric cancer: a microRNA expression analysis

    PubMed Central

    Zhou, Xin; Zhu, Wei; Li, Hai; Wen, Wei; Cheng, Wenfang; Wang, Fang; Wu, Yinxia; Qi, Lianwen; Fan, Yong; Chen, Yan; Ding, Yin; Xu, Jing; Qian, Jiaqi; Huang, Zebo; Wang, Tongshan; Zhu, Danxia; Shu, Yongqian; Liu, Ping

    2015-01-01

    The differential expression of microRNAs (miRNAs) in plasma of gastric cancer (GC) patients may serve as a diagnostic biomarker. A total of 33 miRNAs were identified through the initial screening phase (3 GC pools vs. 1 normal control (NC) pool) using quantitative reverse transcription polymerase chain reaction (qRT-PCR) based Exiqon panel (miRCURY-Ready-to-Use-PCR-Human-panel-I + II-V1.M). By qRT-PCR, these miRNAs were further assessed in training (30 GC VS. 30 NCs) and testing stages (71 GC VS. 61 NCs). We discovered a plasma miRNA signature including five up-regulated miRNAs (miR-185, miR-20a, miR-210, miR-25 and miR-92b), and this signature was evaluated to be a potential diagnostic marker of GC. The areas under the receiver operating characteristic curve of the signature were 0.86, 0.74 and 0.87 for the training, testing and the external validation stages (32 GC VS. 18 NCs), respectively. The five miRNAs were consistently dysregulated in GC tissues (n = 30). Moreover, miR-185 was decreased while miR-20a, miR-210 and miR-92b were increased in arterial plasma (n = 38). However, none of the miRNAs in the exosomes showed different expression between 10 GC patients and 10 NCs. In conclusion, we identified a five-miRNA signature in the peripheral plasma which could serve as a non-invasive biomarker in detection of GC. PMID:26059512

  15. Therapeutic Potential of Modulating MicroRNA in Peripheral Artery Disease

    PubMed Central

    Hamburg, Naomi M.; Leeper, Nicholas J.

    2015-01-01

    Peripheral artery disease (PAD) produces significant disability attributable to lower extremity ischemia. Limited treatment modalities exist to ameliorate clinical symptoms in patients with PAD. Growing evidence links microRNAs to key processes that govern disease expression in PAD including angiogenesis, endothelial function, inflammation, vascular regeneration, vascular smooth muscle cell function, restenosis, and mitochondrial function. MicroRNAs have been identified in circulation and may serve as novel biomarkers in PAD. This article reviews the potential contribution of microRNA to key pathways of disease development in PAD that may lead to microRNA-based diagnostic and therapeutic approaches. PMID:23713861

  16. ARGONAUTE 1 homeostasis invokes the coordinate action of the microRNA and siRNA pathways.

    PubMed

    Mallory, Allison C; Vaucheret, Hervé

    2009-05-01

    ARGONAUTE 1 (AGO1) slices endogenous messenger RNAs (mRNAs) during both microRNA (miRNA)- and short interfering RNA (siRNA)-guided post-transcriptional silencing. We have previously reported that AGO1 homeostasis is maintained through the repressive action of miR168 on AGO1 mRNA and the stabilizing effect of AGO1 protein on miR168, but siRNA-mediated AGO1 regulation has not been reported. Here, we show that AGO1-derived siRNAs trigger RNA DEPENDENT RNA POLYMERASE 6 (RDR6)-, SUPPRESSOR OF GENE SILENCING 3 (SGS3)- and SILENCING DEFECTIVE 5 (SDE5)-dependent AGO1 silencing, which also requires DICER-LIKE 2 (DCL2) and DCL4. By varying the efficacy of miR168-guided AGO1 mRNA cleavage, we show that siRNA-mediated AGO1 silencing depends on correct miRNA targeting, pointing to coordinated regulatory actions of the miRNA and siRNA pathways during the maintenance of AGO1 homeostasis. Finally, our results reveal that dcl2, dcl3 and dcl4 mutations similarly affect post-transcriptional gene silencing (PTGS) mediated by a sense transgene and PTGS mediated by inverted repeats, validating the branched pathway model proposed previously. PMID:19343050

  17. Combined RT-qPCR of mRNA and microRNA Targets within One Fluidigm Integrated Fluidic Circuit

    PubMed Central

    Baldwin, Don A.; Horan, Annamarie D.; Hesketh, Patrick J.

    2016-01-01

    The ability to profile expression levels of a large number of mRNAs and microRNAs (miRNAs) within the same sample, using a single assay method, would facilitate investigations of miRNA effects on mRNA abundance and streamline biomarker screening across multiple RNA classes. A protocol is described for reverse transcription of long RNA and miRNA targets, followed by preassay amplification of the pooled cDNAs and quantitative PCR (qPCR) detection for a mixed panel of candidate RNA biomarkers. The method provides flexibility for designing custom target panels, is robust over a range of input RNA amounts, and demonstrated a high assay success rate. PMID:26977138

  18. Combined RT-qPCR of mRNA and microRNA Targets within One Fluidigm Integrated Fluidic Circuit.

    PubMed

    Baldwin, Don A; Horan, Annamarie D; Hesketh, Patrick J; Mehta, Samir

    2016-07-01

    The ability to profile expression levels of a large number of mRNAs and microRNAs (miRNAs) within the same sample, using a single assay method, would facilitate investigations of miRNA effects on mRNA abundance and streamline biomarker screening across multiple RNA classes. A protocol is described for reverse transcription of long RNA and miRNA targets, followed by preassay amplification of the pooled cDNAs and quantitative PCR (qPCR) detection for a mixed panel of candidate RNA biomarkers. The method provides flexibility for designing custom target panels, is robust over a range of input RNA amounts, and demonstrated a high assay success rate. PMID:26977138

  19. microRNA-103a functions as a mechanosensitive microRNA to inhibit bone formation through targeting Runx2.

    PubMed

    Zuo, Bin; Zhu, JunFeng; Li, Jiao; Wang, ChuanDong; Zhao, XiaoYing; Cai, GuiQuan; Li, Zheng; Peng, Jianping; Wang, Peng; Shen, Chao; Huang, Yan; Xu, Jiake; Zhang, XiaoLing; Chen, XiaoDong

    2015-02-01

    Emerging evidence indicates that microRNAs (miRNAs) play essential roles in regulating osteoblastogenesis and bone formation. However, the role of miRNA in osteoblast mechanotransduction remains to be defined. In this study, we aimed to investigate whether miRNAs regulate mechanical stimulation-triggered osteoblast differentiation and bone formation through modulation of Runx2, the master transcription factor for osteogenesis. We first investigated the role of mechanical loading both in a mouse model and in an osteoblast culture system and the outcomes clearly demonstrated that mechanical stimuli can regulate osteogenesis and bone formation both in vivo and in vitro. Using bioinformatic analyses and subsequent confirmation by quantitative real-time PCR (qRT-PCR), we found that multiple miRNAs that potentially target Runx2 were responding to in vitro mechanical stimulation, among which miR-103a was fully characterized. miR-103a and its host gene PANK3 were both downregulated during cyclic mechanical stretch (CMS)-induced osteoblast differentiation, whereas Runx2 protein expression was upregulated. Overexpression of miR-103a significantly decreased and inhibition of miR-103a increased Runx2 protein level, suggesting that miR-103a acts as an endogenous attenuator of Runx2 in osteoblasts. Mutation of putative miR-103a binding sites in Runx2 mRNA abolishes miR-103a-mediated repression of the Runx2 3'-untranslated region (3'UTR) luciferase reporter activity, suggesting that miR-103a binds to Runx2 3'UTR. Osteoblast marker gene profiling and osteogenic phenotype assays demonstrated that miR-103a negatively correlates with CMS-induced osteogenesis. Further, the perturbation of miR-103a also has a significant effect on osteoblast activity and matrix mineralization. More importantly, we found an inhibitory role of miR-103a in regulating bone formation in hindlimb unloading mice, and pretreatment with antagomir-103a partly rescued the osteoporosis caused by mechanical

  20. Uridylation of RNA Hairpins by Tailor Confines the Emergence of MicroRNAs in Drosophila

    PubMed Central

    Reimão-Pinto, Madalena M.; Ignatova, Valentina; Burkard, Thomas R.; Hung, Jui-Hung; Manzenreither, Raphael A.; Sowemimo, Ivica; Herzog, Veronika A.; Reichholf, Brian; Fariña-Lopez, Sara; Ameres, Stefan L.

    2015-01-01

    Summary Uridylation of RNA species represents an emerging theme in post-transcriptional gene regulation. In the microRNA pathway, such modifications regulate small RNA biogenesis and stability in plants, worms, and mammals. Here, we report Tailor, an uridylyltransferase that is required for the majority of 3′ end modifications of microRNAs in Drosophila and predominantly targets precursor hairpins. Uridylation modulates the characteristic two-nucleotide 3′ overhang of microRNA hairpins, which regulates processing by Dicer-1 and destabilizes RNA hairpins. Tailor preferentially uridylates mirtron hairpins, thereby impeding the production of non-canonical microRNAs. Mirtron selectivity is explained by primary sequence specificity of Tailor, selecting substrates ending with a 3′ guanosine. In contrast to mirtrons, conserved Drosophila precursor microRNAs are significantly depleted in 3′ guanosine, thereby escaping regulatory uridylation. Our data support the hypothesis that evolutionary adaptation to Tailor-directed uridylation shapes the nucleotide composition of precursor microRNA 3′ ends. Hence, hairpin uridylation may serve as a barrier for the de novo creation of microRNAs in Drosophila. PMID:26145176

  1. Describing a Transcription Factor Dependent Regulation of the MicroRNA Transcriptome.

    PubMed

    Rozovski, Uri; Hazan-Halevy, Inbal; Calin, George; Harris, David; Li, Ping; Liu, Zhiming; Keating, Michael J; Estrov, Zeev

    2016-01-01

    While the transcription regulation of protein coding genes was extensively studied, little is known on how transcription factors are involved in transcription of non-coding RNAs, specifically of microRNAs. Here, we propose a strategy to study the potential role of transcription factor in regulating transcription of microRNAs using publically available data, computational resources and high throughput data. We use the H3K4me3 epigenetic signature to identify microRNA promoters and chromatin immunoprecipitation (ChIP)-sequencing data from the ENCODE project to identify microRNA promoters that are enriched with transcription factor binding sites. By transfecting cells of interest with shRNA targeting a transcription factor of interest and subjecting the cells to microRNA array, we study the effect of this transcription factor on the microRNA transcriptome. As an illustrative example we use our study on the effect of STAT3 on the microRNA transcriptome of chronic lymphocytic leukemia (CLL) cells. PMID:27341356

  2. Retinoic Acid Induces Embryonic Stem Cell Differentiation by Altering Both Encoding RNA and microRNA Expression

    PubMed Central

    Yu, Mengying; Wu, Haibo; Ai, Zhiying; Wu, Yongyan; Liu, Hongliang; Du, Juan; Guo, Zekun; Zhang, Yong

    2015-01-01

    Retinoic acid (RA) is a vitamin A metabolite that is essential for early embryonic development and promotes stem cell neural lineage specification; however, little is known regarding the impact of RA on mRNA transcription and microRNA levels on embryonic stem cell differentiation. Here, we present mRNA microarray and microRNA high-output sequencing to clarify how RA regulates gene expression. Using mRNA microarray analysis, we showed that RA repressed pluripotency-associated genes while activating ectoderm markers in mouse embryonic stem cells (mESCs). Moreover, RA modulated the DNA methylation of mESCs by altering the expression of epigenetic-associated genes such as Dnmt3b and Dnmt3l. Furthermore, H3K4me2, a pluripotent histone modification, was repressed by RA stimulation. From microRNA sequence data, we identified two downregulated microRNAs, namely, miR-200b and miR-200c, which regulated the pluripotency of stem cells. We found that miR-200b or miR-200c deficiency suppressed the expression of pluripotent genes, including Oct4 and Nanog, and activated the expression of the ectodermal marker gene Nestin. These results demonstrate that retinoid induces mESCs to differentiate by regulating miR-200b/200c. Our findings provide the landscapes of mRNA and microRNA gene networks and indicate the crucial role of miR-200b/200c in the RA-induced differentiation of mESCs. PMID:26162091

  3. MicroRNA modulation combined with sunitinib as a novel therapeutic strategy for pancreatic cancer

    PubMed Central

    Passadouro, Marta; Pedroso de Lima, Maria C; Faneca, Henrique

    2014-01-01

    Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive and mortal cancer, characterized by a set of known mutations, invasive features, and aberrant microRNA expression that have been associated with hallmark malignant properties of PDAC. The lack of effective PDAC treatment options prompted us to investigate whether microRNAs would constitute promising therapeutic targets toward the generation of a gene therapy approach with clinical significance for this disease. In this work, we show that the developed human serum albumin–1-palmitoyl-2-oleoyl-sn-glycero-3-ethylphosphocholine:cholesterol/anti-microRNA oligonucleotides (+/−) (4/1) nanosystem exhibits the ability to efficiently deliver anti-microRNA oligonucleotides targeting the overexpressed microRNAs miR-21, miR-221, miR-222, and miR-10 in PDCA cells, promoting an almost complete abolishment of microRNA expression. Silencing of these microRNAs resulted in a significant increase in the levels of their targets. Moreover, the combination of microRNA silencing, namely miR-21, with low amounts of the chemotherapeutic drug sunitinib resulted in a strong and synergistic antitumor effect, showing that this combined strategy could be of great importance for therapeutic application in PDAC. PMID:25061297

  4. The functional consequences of age-related changes in microRNA expression in skeletal muscle.

    PubMed

    Soriano-Arroquia, Ana; House, Louise; Tregilgas, Luke; Canty-Laird, Elizabeth; Goljanek-Whysall, Katarzyna

    2016-06-01

    A common characteristic of ageing is disrupted homeostasis between growth and atrophy of skeletal muscle resulting in loss of muscle mass and function, which is associated with sarcopenia. Sarcopenia is related to impaired balance, increased falls and decline in quality of life of older people. Ageing-related transcriptome and proteome changes in skeletal muscle have been characterised, however the molecular mechanisms underlying sarcopenia are still not fully understood. microRNAs are novel regulators of gene expression known to modulate skeletal muscle development and homeostasis. Expression of numerous microRNAs is disrupted in skeletal muscle with age however, the functional consequences of this are not yet understood. Given that a single microRNA can simultaneously affect multiple signalling pathways, microRNAs are potent modulators of pathophysiological changes occurring during ageing. Here we use microRNA and transcript expression profiling together with microRNA functional assays to show that disrupted microRNA:target interactions play an important role in maintaining muscle homeostasis. We identified miR-181a as a regulator of the sirtuin1 (Sirt1) gene expression in skeletal muscle and show that the expression of miR-181a and its target gene is disrupted in skeletal muscle from old mice. Moreover, we show that miR-181a:Sirt1 interactions regulate myotube size. Our results demonstrate that disrupted microRNA:target interactions are likely related to the pathophysiological changes occurring in skeletal muscle during ageing. PMID:26922183

  5. RNACluster: An integrated tool for RNA secondary structure comparison and clustering.

    PubMed

    Liu, Qi; Olman, V; Liu, Huiqing; Ye, Xiuzi; Qiu, Shilun; Xu, Ying

    2008-07-15

    RNA structure comparison is a fundamental problem in structural biology, structural chemistry, and bioinformatics. It can be used for analysis of RNA energy landscapes, conformational switches, and facilitating RNA structure prediction. The purpose of our integrated tool RNACluster is twofold: to provide a platform for computing and comparison of different distances between RNA secondary structures, and to perform cluster identification to derive useful information of RNA structure ensembles, using a minimum spanning tree (MST) based clustering algorithm. RNACluster employs a cluster identification approach based on a MST representation of the RNA ensemble data and currently supports six distance measures between RNA secondary structures. RNACluster provides a user-friendly graphical interface to allow a user to compare different structural distances, analyze the structure ensembles, and visualize predicted structural clusters. PMID:18271070

  6. MicroRNA-203 represses selection and expansion of oncogenic Hras transformed tumor initiating cells

    PubMed Central

    Riemondy, Kent; Wang, Xiao-jing; Torchia, Enrique C; Roop, Dennis R; Yi, Rui

    2015-01-01

    In many mouse models of skin cancer, only a few tumors typically form even though many cells competent for tumorigenesis receive the same oncogenic stimuli. These observations suggest an active selection process for tumor-initiating cells. Here, we use quantitative mRNA- and miR-Seq to determine the impact of HrasG12V on the transcriptome of keratinocytes. We discover that microRNA-203 is downregulated by HrasG12V. Using a knockout mouse model, we demonstrate that loss of microRNA-203 promotes selection and expansion of tumor-initiating cells. Conversely, restoration of microRNA-203 using an inducible model potently inhibits proliferation of these cells. We comprehensively identify microRNA-203 targets required for Hras-initiated tumorigenesis. These targets include critical regulators of the Ras pathway and essential genes required for cell division. This study establishes a role for the loss of microRNA-203 in promoting selection and expansion of Hras mutated cells and identifies a mechanism through which microRNA-203 antagonizes Hras-mediated tumorigenesis. DOI: http://dx.doi.org/10.7554/eLife.07004.001 PMID:26203562

  7. MicroRNA-mediated regulation of target genes in several brain regions is correlated to both microRNA-targeting-specific promoter methylation and differential microRNA expression

    PubMed Central

    2013-01-01

    Background Public domain databases nowadays provide multiple layers of genome-wide data e.g., promoter methylation, mRNA expression, and miRNA expression and should enable integrative modeling of the mechanisms of regulation of gene expression. However, researches along this line were not frequently executed. Results Here, the public domain dataset of mRNA expression, microRNA (miRNA) expression and promoter methylation patterns in four regions, the frontal cortex, temporal cortex, pons and cerebellum, of human brain were sourced from the National Center for Biotechnology Informations gene expression omnibus, and reanalyzed computationally. A large number of miRNA-mediated regulation of target genes and miRNA-targeting-specific promoter methylation were identified in the six pairwise comparisons among the four brain regions. The miRNA-mediated regulation of target genes was found to be highly correlated with one or both of miRNA-targeting-specific promoter methylation and differential miRNA expression. Genes enriched for Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways that were related to brain function and/or development were found among the target genes of miRNAs whose differential expression patterns were highly correlated with the miRNA-mediated regulation of their target genes. Conclusions The combinatorial analysis of miRNA-mediated regulation of target genes, miRNA-targeting-specific promoter methylation and differential miRNA expression can help reveal the brain region-specific contributions of miRNAs to brain function and development. PMID:23725297

  8. Down-regulation of a host microRNA by a viral noncoding RNA.

    PubMed

    Cazalla, D; Steitz, J A

    2010-01-01

    Primate herpesviruses express more noncoding RNAs (ncRNAs) than any other class of mammalian viruses during either latency or the lytic phase of the viral life cycle. T cells transformed by the monkey virus Herpesvirus saimiri (HVS) express seven viral U-rich ncRNAs called HSURs. Conserved sequences in HSURs1 and 2 exhibit complementarity to three host-cell microRNAs (miRNAs). The predicted interactions of HSURs1 and 2 with these miRNAs were confirmed by coimmuno-precipitation experiments performed on extracts of marmoset T cells transformed by a wild-type or a mutant HVS lacking these two HSURs. Mutational analyses demonstrated that the binding of miR-27 to HSUR1 and that of miR-16 to HSUR2 involves base pairing. One of these miRNAs, miR-27, is dramatically lowered in abundance in HVS-transformed cells, with consequent effects on the expression of miR-27 target genes. Transient knockdown and ectopic expression of HSUR1 demonstrated that degradation of mature miR-27 occurs in a sequence-specific and binding-dependent manner but does not occur by AU-rich element (ARE)-mediated decay, which controls the intracellular level of HSUR1 itself. This viral strategy exemplifies the use of an ncRNA to control host-cell gene expression via the miRNA pathway and has potential applications both experimentally and therapeutically. PMID:21139068

  9. MicroRNA profile in very young women with breast cancer

    PubMed Central

    2014-01-01

    Background Breast cancer is rarely diagnosed in very young women (35years old or younger), and it often presents with distinct clinical-pathological features related to a more aggressive phenotype and worse prognosis when diagnosed at this early age. A pending question is whether breast cancer in very young women arises from the deregulation of different underlying mechanisms, something that will make this disease an entity differentiated from breast cancer diagnosed in older patients. Methods We performed a comprehensive study of miRNA expression using miRNA Affymetrix2.0 array on paraffin-embedded tumour tissue of 42 breast cancer patients 35 years old or younger, 17 patients between 45 and 65 years old and 29 older than 65 years. Data were statistically analyzed by t-test and a hierarchical clustering via average linkage method was conducted. Results were validated by qRT-PCR. Putative targeted pathways were obtained using DIANA miRPath online software. Results The results show a differential and unique miRNA expression profile of 121 miRNAs (p-value <0.05), 96 of those with a FDR-value <0.05. Hierarchical clustering grouped the samples according to their age, but not by subtype nor by tumour characteristics. We were able to validate by qRT-PCR differences in the expression of 6 miRNAs: miR-1228*, miR-3196, miR-1275, miR-92b, miR-139 and miR-1207. Moreover, all of the miRNAs maintained the expression trend. The validated miRNAs pointed out pathways related to cell motility, invasion and proliferation. Conclusions The study suggests that breast cancer in very young women appears as a distinct molecular signature. To our knowledge, this is the first time that a validated microRNA profile, distinctive to breast cancer in very young women, has been presented. The miRNA signature may be relevant to open an important field of research in order to elucidate the underlying mechanism in this particular disease, which in a more clinical setting, could potentially help to

  10. MicroRNA-21 suppression impedes medulloblastoma cell migration.

    PubMed

    Grunder, Eveline; D'Ambrosio, Rocco; Fiaschetti, Giulio; Abela, Lucia; Arcaro, Alexandre; Zuzak, Tycho; Ohgaki, Hiroko; Lv, Sheng-Qing; Shalaby, Tarek; Grotzer, Michael

    2011-11-01

    Medulloblastoma (MB), the most common malignant brain tumour in children, is characterised by a high risk of leptomeningeal dissemination. But little is known about the molecular mechanisms that promote cancer cell migration in MB. Aberrant expression of miR-21 is recognised to be causatively linked to metastasis in a variety of human neoplasms including brain tumours; however its function in MB is still unknown. In this study we investigated the expression level and the role of miR-21 in MB cell migration. miR-21 was found to be up-regulated, compared to normal cerebellum, in 29/29 MB primary samples and 6/6 MB-derived cell lines. Inverse correlation was observed between miR-21 expression and the metastasis suppressor PDCD4, while miR-21 repression increased the release of PDCD4 protein, suggesting negative regulation of PDCD4 by miR-21 in MB cells. Anti-miR-21 decreased protein expression of the tumour cell invasion mediators MAP4K1 and JNK, which are also known to be negatively regulated by PDCD4, and down-regulated integrin protein that is essential for MB leptomeningeal dissemination. Moreover miR-21 knockdown in MB cells increased the expression of two eminent negative modulators of cancer cell migration, E-Cadherin and TIMP2 proteins that are known to be positively regulated by PDCD4. Finally and importantly, suppression of miR-21 decreased the motility of MB cells and reduced their migration across basement membranes in vitro. Together, these compelling data propose miR-21 pathway as a novel mechanism impacting MB cell dissemination and raises the possibility that curability of selected MB may be improved by pharmaceutical strategies directed towards microRNA-21. PMID:21775132

  11. MicroRNA-214 Antagonism Protects against Renal Fibrosis

    PubMed Central

    Ramdas, Vasudev; Lu, Ruifang; Conway, Bryan R.; Grant, Jennifer S.; Dickinson, Brent; Aurora, Arin B.; McClure, John D.; Kipgen, David; Delles, Christian; van Rooij, Eva

    2014-01-01

    Renal tubulointerstitial fibrosis is the common end point of progressive renal disease. MicroRNA (miR)-214 and miR-21 are upregulated in models of renal injury, but the function of miR-214 in this setting and the effect of its manipulation remain unknown. We assessed the effect of inhibiting miR-214 in an animal model of renal fibrosis. In mice, genetic deletion of miR-214 significantly attenuated interstitial fibrosis induced by unilateral ureteral obstruction (UUO). Treatment of wild-type mice with an anti-miR directed against miR-214 (anti-miR-214) before UUO resulted in similar antifibrotic effects, and in vivo biodistribution studies demonstrated that anti–miR-214 accumulated at the highest levels in the kidney. Notably, in vivo inhibition of canonical TGF-β signaling did not alter the regulation of endogenous miR-214 or miR-21. Whereas miR-21 antagonism blocked Smad 2/3 activation, miR-214 antagonism did not, suggesting that miR-214 induces antifibrotic effects independent of Smad 2/3. Furthermore, TGF-β blockade combined with miR-214 deletion afforded additional renal protection. These phenotypic effects of miR-214 depletion were mediated through broad regulation of the transcriptional response to injury, as evidenced by microarray analysis. In human kidney tissue, miR-214 was detected in cells of the glomerulus and tubules as well as in infiltrating immune cells in diseased tissue. These studies demonstrate that miR-214 functions to promote fibrosis in renal injury independent of TGF-β signaling in vivo and that antagonism of miR-214 may represent a novel antifibrotic treatment in the kidney. PMID:24158985

  12. MicroRNA Dysregulation in Diabetic Ischemic Heart Failure Patients

    PubMed Central

    Greco, Simona; Fasanaro, Pasquale; Castelvecchio, Serenella; D’Alessandra, Yuri; Arcelli, Diego; Di Donato, Marisa; Malavazos, Alexis; Capogrossi, Maurizio C.; Menicanti, Lorenzo; Martelli, Fabio

    2012-01-01

    Increased morbidity and mortality associated with ischemic heart failure (HF) in type 2 diabetic patients requires a deeper understanding of the underpinning pathogenetic mechanisms. Given the implication of microRNAs (miRNAs) in HF, we investigated their regulation and potential role. miRNA expression profiles were measured in left ventricle biopsies from 10 diabetic HF (D-HF) and 19 nondiabetic HF (ND-HF) patients affected by non–end stage dilated ischemic cardiomyopathy. The HF groups were compared with each other and with 16 matched nondiabetic, non-HF control subjects. A total of 17 miRNAs were modulated in D-HF and/or ND-HF patients when compared with control subjects. miR-216a, strongly increased in both D-HF and ND-HF patients, negatively correlated with left ventricular ejection fraction. Six miRNAs were differently expressed when comparing D-HF and ND-HF patients: miR-34b, miR-34c, miR-199b, miR-210, miR-650, and miR-223. Bioinformatic analysis of their modulated targets showed the enrichment of cardiac dysfunctions and HF categories. Moreover, the hypoxia-inducible factor pathway was activated in the noninfarcted, vital myocardium of D-HF compared with ND-HF patients, indicating a dysregulation of the hypoxia response mechanisms. Accordingly, miR-199a, miR-199b, and miR-210 were modulated by hypoxia and high glucose in cardiomyocytes and endothelial cells cultured in vitro. In conclusion, these findings show a dysregulation of miRNAs in HF, shedding light on the specific disease mechanisms differentiating diabetic patients. PMID:22427379

  13. MicroRNA loss enhances learning and memory in mice.

    PubMed

    Konopka, Witold; Kiryk, Anna; Novak, Martin; Herwerth, Marina; Parkitna, Jan Rodriguez; Wawrzyniak, Marcin; Kowarsch, Andreas; Michaluk, Piotr; Dzwonek, Joanna; Arnsperger, Tabea; Wilczynski, Grzegorz; Merkenschlager, Matthias; Theis, Fabian J; Köhr, Georg; Kaczmarek, Leszek; Schütz, Günther

    2010-11-01

    Dicer-dependent noncoding RNAs, including microRNAs (miRNAs), play an important role in a modulation of translation of mRNA transcripts necessary for differentiation in many cell types. In vivo experiments using cell type-specific Dicer1 gene inactivation in neurons showed its essential role for neuronal development and survival. However, little is known about the consequences of a loss of miRNAs in adult, fully differentiated neurons. To address this question, we used an inducible variant of the Cre recombinase (tamoxifen-inducible CreERT2) under control of Camk2a gene regulatory elements. After induction of Dicer1 gene deletion in adult mouse forebrain, we observed a progressive loss of a whole set of brain-specific miRNAs. Animals were tested in a battery of both aversively and appetitively motivated cognitive tasks, such as Morris water maze, IntelliCage system, or trace fear conditioning. Compatible with rather long half-life of miRNAs in hippocampal neurons, we observed an enhancement of memory strength of mutant mice 12 weeks after the Dicer1 gene mutation, before the onset of neurodegenerative process. In acute brain slices, immediately after high-frequency stimulation of the Schaffer collaterals, the efficacy at CA3-to-CA1 synapses was higher in mutant than in control mice, whereas long-term potentiation was comparable between genotypes. This phenotype was reflected at the subcellular and molecular level by the elongated filopodia-like shaped dendritic spines and an increased translation of synaptic plasticity-related proteins, such as BDNF and MMP-9 in mutant animals. The presented work shows miRNAs as key players in the learning and memory process of mammals. PMID:21048142

  14. miReg: a resource for microRNA regulation.

    PubMed

    Barh, Debmalya; Bhat, Dattatraya; Viero, Cedric

    2010-01-01

    MicroRNAs (miRNAs/miRs) are important cellular components that regulate gene expression at posttranscriptional level. Various upstream components regulate miR expression and any deregulation causes disease conditions. Therefore, understanding of miR regulatory network both at upstream and downstream level is crucial and a resource on this aspect will be helpful. Currently available miR databases are mostly related to downstream targets, sequences, or diseases. But as of now, no database is available that provides a complete picture of miR regulation in a specific condition. Our miR regulation web resource (miReg) is a manually curated one that represents validated upstream regulators (transcription factor, drug, physical, and chemical) along with downstream targets, associated biological process, experimental condition or disease state, up or down regulation of the miR in that condition, and corresponding PubMed references in a graphical and user friendly manner, browseable through 5 browsing options. We have presented exact facts that have been described in the corresponding literature in relation to a given miR, whether it's a feed-back/feed-forward loop or inhibition/activation. Moreover we have given various links to integrate data and to get a complete picture on any miR listed. Current version (Version 1.0) of miReg contains 47 important human miRs with 295 relations using 190 absolute references. We have also provided an example on usefulness of miReg to establish signalling pathways involved in cardiomyopathy. We believe that miReg will be an essential miRNA knowledge base to research community, with its continuous upgrade and data enrichment. This HTML based miReg can be accessed from: www.iioab-mireg.webs.com or www.iioab.webs.com/mireg.htm. PMID:20693604

  15. MicroRNA-421 Gene Polymorphism in Gastric Carcinoma.

    PubMed

    Jin, Xin; Yu, Nong

    2016-01-01

    BACKGROUND As a common malignant tumor, gastric carcinoma requires early diagnosis to improve treatment efficacy. MicroRNA (miR) molecules have highly conserved nucleotide sequences and can negatively regulate target gene expression at the translational level. miR-421 has been suggested to be related with gastric cancer occurrence. The gene polymorphism of miR-421, however, has not been reported. This study thus investigated the G/C polymorphism of miR-421 and its role in progression and prognosis of gastric cancer. MATERIAL AND METHODS A total of 96 gastric cancer patients were recruited in this study and tumor samples were collected from surgical resection. Single-nucleotide polymorphism (SNP) of miR-421 was determined by DNA sequencing for analyzing the correlation between lymph node metastasis and miR-421 genotypes. Logistic regression analysis was used to determine the relationship between genotype and risk factors of gastric cancer. Kaplan-Meier survival analysis was also performed to compare GG and GC carriers. RESULTS Differential expression patterns existed between gastric cancer tissues and normal gastric mucosa. Logistic regression analysis showed GC and GG genotypes were risk factors for gastric cancer. Patients with lymph node metastasis had higher GG genotype frequency compared to those without metastasis. In survival analysis, GG carriers had shorter survival time than GC carriers. Furthermore, GG genotype was correlated with tumor prognosis (p<0.05). CONCLUSIONS G allele of miR-421 is a risk factor for gastric cancer. GG genotype is correlated with lymph node metastasis and prognosis, indicating it is a risk factor for gastric cancer. PMID:27133200

  16. Small RNA and RNA-IP Sequencing Identifies and Validates Novel MicroRNAs in Human Mesenchymal Stem Cells.

    PubMed

    Tsai, Chin-Han; Liao, Ko-Hsun; Shih, Chuan-Chi; Chan, Chia-Hao; Hsieh, Jui-Yu; Tsai, Cheng-Fong; Wang, Hsei-Wei; Chang, Shing-Jyh

    2016-03-01

    Organ regeneration therapies using multipotent mesenchymal stem cells (MSCs) are currently being investigated for a variety of common complex diseases. Understanding the molecular regulation of MSC biology will benefit regenerative medicine. MicroRNAs (miRNAs) act as regulators in MSC stemness. There are approximately 2500 currently known human miRNAs that have been recorded in the miRBase v21 database. In the present study, we identified novel microRNAs involved in MSC stemness and differentiation by obtaining the global microRNA expression profiles (miRNomes) of MSCs from two anatomical locations bone marrow (BM-MSCs) and umbilical cord Wharton's jelly (WJ-MSCs) and from osteogenically and adipogenically differentiated progenies of BM-MSCs. Small RNA sequencing (smRNA-seq) and bioinformatics analyses predicted that 49 uncharacterized miRNA candidates had high cellular expression values in MSCs. Another independent batch of Ago1/2-based RNA immunoprecipitation (RNA-IP) sequencing datasets validated the existence of 40 unreported miRNAs in cells and their associations with the RNA-induced silencing complex (RISC). Nine of these 40 new miRNAs were universally overexpressed in both MSC types; nine others were overexpressed in differentiated cells. A novel miRNA (UNI-118-3p) was specifically expressed in BM-MSCs, as verified using RT-qPCR. Taken together, this report offers comprehensive miRNome profiles for two MSC types, as well as cells differentiated from BM-MSCs. MSC transplantation has the potential to ameliorate degenerative disorders and repair damaged tissues. Interventions involving the above 40 new microRNA members in transplanted MSCs may potentially guide future clinical applications. PMID:26910904

  17. Assessment of mRNA and microRNA Stabilization in Peripheral Human Blood for Multicenter Studies and Biobanks

    PubMed Central

    Weber, Daniel Gilbert; Casjens, Swaantje; Rozynek, Peter; Lehnert, Martin; Zilch-Schöneweis, Sandra; Bryk, Oleksandr; Taeger, Dirk; Gomolka, Maria; Kreuzer, Michaela; Otten, Heinz; Pesch, Beate; Johnen, Georg; Brüning, Thomas

    2010-01-01

    In this study we evaluate the suitability of two methods of RNA conservation in blood samples, PAXgene and RNAlater, in combination with variable shipping conditions for their application in multicenter studies and biobanking. RNA yield, integrity, and purity as well as levels of selected mRNA and microRNA species were analyzed in peripheral human blood samples stabilized by PAXgene or RNAlater and shipped on dry ice or at ambient temperatures from the study centers to the central analysis laboratory. Both examined systems were clearly appropriate for RNA stabilization in human blood independently of the shipping conditions. The isolated RNA is characterized by good quantity and quality and well suited for downstream applications like quantitative RT-PCR analysis of mRNA and microRNA. Superior yield and integrity values were received using RNAlater. It would be reasonable to consider the production and approval of blood collection tubes prefilled with RNAlater to facilitate the use of this excellent RNA stabilization system in large studies. PMID:20981139

  18. MicroRNA-17~92 inhibits colorectal cancer progression by targeting angiogenesis.

    PubMed

    Ma, Huabin; Pan, Jin-Shui; Jin, Li-Xin; Wu, Jianfeng; Ren, Yan-Dan; Chen, Pengda; Xiao, Changchun; Han, Jiahuai

    2016-07-01

    The miR-17~92 microRNA (miRNA) cluster host gene is upregulated in a broad spectrum of human cancers including colorectal cancer (CRC). Previous studies have shown that miR-17~92 promotes tumorigenesis and cancer angiogenesis in some tumor models. However, its role in the initiation and progression of CRC remains unknown. In this study, we found that transgenic mice overexpressing miR-17~92 specifically in epithelial cells of the small and large intestines exhibited decreased tumor size and tumor angiogenesis in azoxymethane and dextran sulfate sodium salt (AOM-DSS)-induced CRC model as compared to their littermates control. Further study showed that miR-17~92 inhibited the progression of CRC via suppressing tumor angiogenesis through targeting multiple tumor angiogenesis-inducing genes, TGFBR2, HIF1α, and VEGFA in vivo and in vitro. Collectively, we demonstrated that miR-17~92 suppressed tumor progression by inhibiting tumor angiogenesis in a genetically engineered mouse model, indicating the presence of cellular context-dependent pro- and anti-cancer effects of miR-17~92. PMID:27080303

  19. Prefrontal microRNA-221 Mediates Environmental Enrichment-Induced Increase of Locomotor Sensitivity to Nicotine

    PubMed Central

    Gomez, Adrian M.; Altomare, Diego; Sun, Wei-Lun; Midde, Narasimha M.; Ji, Hao; Shtutman, Michael; Turner, Jill R.; Creek, Kim E.

    2016-01-01

    Background: Environmental enrichment alters susceptibility in developing drug addiction. We have demonstrated that rats raised in an enriched condition are more sensitive than rats raised in an impoverished condition to nicotine-induced locomotor activity, and this is associated with alterations of phosphorylated extracellular signal-regulated kinase 1/2 within the prefrontal cortex. This study determined the impact of microRNA-221 in the prefrontal cortex on phosphorylated extracellular signal-regulated kinase 1/2 and the enriched environment-dependent behavioral changes in response to nicotine. Methods: A microRNA array was conducted to profile microRNA expression in the prefrontal cortex of enriched condition and impoverished condition rats in response to repeated nicotine (0.35mg/kg, s.c.) administration. microRNA-221 in the prefrontal cortex, nucleus accumbens, and striatum was further verified by quantitative real-time PCR. Lentiviral-mediated overexpression of microRNA-221 in PC12 cells and the medial prefrontal cortex was performed to determine the effects of microRNA-221 on nicotine-mediated phosphorylated extracellular signal-regulated kinase 1/2, phosphorylated cAMP-response element-binding protein, and locomotor activity. Results: microRNA-221 was profoundly upregulated in the prefrontal cortex but not in nucleus accumbens and striatum of enriched condition rats relative to impoverished condition rats following repeated administration of nicotine. Overexpression of lentiviral-microRNA-221 attenuated nicotine-induced increase in phosphorylated extracellular signal-regulated kinase 1/2 in PC12 cells. Lentiviral-microRNA-221 overexpression in the medial prefrontal cortex further increased locomotor activity in impoverished condition but not in enriched condition rats in response to repeated nicotine administration. Accordingly, lentiviral-microRNA-221 attenuated nicotine-induced increases in phosphorylated extracellular signal-regulated kinase 1/2 and

  20. Clustering siRNA conjugates for MMP-responsive therapeutics in chronic wounds of diabetic animals.

    PubMed

    Kim, Hye Sung; Son, Young Ju; Yoo, Hyuk Sang

    2016-07-21

    The MMP-responsive breakdown of siRNA clusters was translated to site-specific gene transfection and enhanced wound healing in diabetic ulcers. MMP-2 siRNA was chemically tethered to the end of multi-armed PEG via MMP-cleavable linkers (4PEG-siRNA) and subsequently clustered into submicron particles complexed with LPEI. 4PEG-siRNA was more tightly complexed with LPEI and the associated cluster showed higher resistance against RNase attack, in comparison to naked siRNA. Because the size of the clusters increased depending on the increase in charge ratio of LPEI to siRNA, cellular uptake of the 4PEG-siRNA/LPEI cluster was significantly attenuated due to the huge size of the cluster. However, upon MMP treatment, the cluster dissociated into smaller particles and was efficiently endocytosed by cells. An in vivo fluorescence resonance energy transfer (FRET) study also revealed that the clusters were effectively dissociated in MMP-rich environments of dorsal wounds in diabetic animals. In addition, diabetic ulcers treated with the clusters showed a faster wound closure rate and the recovered tissue expressed a larger amount of cytokeratin along with a lower expression level of MMP-2 compared to the other groups. PMID:27251781

  1. mRNA Targeting to Endoplasmic Reticulum Precedes Ago Protein Interaction and MicroRNA (miRNA)-mediated Translation Repression in Mammalian Cells*

    PubMed Central

    Barman, Bahnisikha; Bhattacharyya, Suvendra N.

    2015-01-01

    MicroRNA (miRNA) binds to the 3′-UTR of its target mRNAs to repress protein synthesis. Extensive research was done to understand the mechanism of miRNA-mediated repression in animal cells. Considering the progress in understanding the mechanism, information about the subcellular sites of miRNA-mediated repression is surprisingly limited. In this study, using an inducible expression system for an miRNA target message, we have delineated how a target mRNA passes through polysome association and Ago2 interaction steps on rough endoplasmic reticulum (ER) before the miRNA-mediated repression sets in. From this study, de novo formed target mRNA localization to the ER-bound polysomes manifested as the earliest event, which is followed by Ago2 micro-ribonucleoprotein binding, and translation repression of target message. Compartmentalization of this process to rough ER membrane ensures enrichment of miRNA-targeted messages and micro-ribonucleoprotein components on ER upon reaching a steady state. PMID:26304123

  2. MicroRNA dysregulation in spinal cord injury: causes, consequences and therapeutics

    PubMed Central

    Nieto-Diaz, Manuel; Esteban, Francisco J.; Reigada, David; Muñoz-Galdeano, Teresa; Yunta, Mónica; Caballero-López, Marcos; Navarro-Ruiz, Rosa; del Águila, Ángela; Maza, Rodrigo M.

    2014-01-01

    Trauma to the spinal cord causes permanent disability to more than 180,000 people every year worldwide. The initial mechanical damage triggers a complex set of secondary events involving the neural, vascular, and immune systems that largely determine the functional outcome of the spinal cord injury (SCI). Cellular and biochemical mechanisms responsible for this secondary injury largely depend on activation and inactivation of specific gene programs. Recent studies indicate that microRNAs function as gene expression switches in key processes of the SCI. Microarray data from rodent contusion models reveal that SCI induces changes in the global microRNA expression patterns. Variations in microRNA abundance largely result from alterations in the expression of the cells at the damaged spinal cord. However, microRNA expression levels after SCI are also influenced by the infiltration of immune cells to the injury site and the death and migration of specific neural cells after injury. Evidences on the role of microRNAs in the SCI pathophysiology have come from different sources. Bioinformatic analysis of microarray data has been used to identify specific variations in microRNA expression underlying transcriptional changes in target genes, which are involved in key processes in the SCI. Direct evidences on the role of microRNAs in SCI are scarcer, although recent studies have identified several microRNAs (miR-21, miR-486, miR-20) involved in key mechanisms of the SCI such as cell death or astrogliosis, among others. From a clinical perspective, different evidences make clear that microRNAs can be potent therapeutic tools to manipulate cell state and molecular processes in order to enhance functional recovery. The present article reviews the actual knowledge on how injury affects microRNA expression and the meaning of these changes in the SCI pathophysiology, to finally explore the clinical potential of microRNAs in the SCI. PMID:24701199

  3. The Bull Sperm MicroRNAome and the Effect of Fescue Toxicosis on Sperm MicroRNA Expression

    PubMed Central

    Stowe, Heather M.; Calcatera, Samantha M.; Dimmick, Marcy A.; Andrae, John G.; Duckett, Susan K.; Pratt, Scott L.

    2014-01-01

    Tall fescue [Schedonorus phoenix (Scop.) Holub] accounts for nearly 16 million hectares of pasture in the Southeastern and Mid-Atlantic U.S. due to its heat, drought, and pest resistance, conferred to the plant by its symbiotic relationship with the endophyte Neotyphodium coenophialum. The endophyte produces ergot alkaloids that have negative effects on the growth and reproduction of animals, resulting in the syndrome known as fescue toxicosis. The objectives of our study were to identify microRNA (miRNA) present in bovine sperm and to evaluate the effects of fescue toxicosis on sperm miRNA expression. Angus bulls were assigned to treatments of either toxic or non-toxic fescue seed diets. Semen was collected and subjected to RNA isolation. Three samples from each treatment group were chosen and pooled for deep sequencing. To compare miRNA expression between treatment groups, a microarray was designed and conducted. For each of the top ten expressed miRNA, target prediction analysis was conducted using TargetScan. Gene ontology enrichment was assessed using the Database for Annotation, Visualization and Integrated Discovery. Sequencing results elucidated the presence of 1,582 unique small RNA present in sperm. Of those sequences, 382 were known Bos taurus miRNA, 22 were known but novel to Bos taurus, and 816 were predicted candidate miRNA that did not map to any currently reported miRNA. Of the sequences chosen for microarray, twenty-two showed significant differential expression between treatment groups. Gene pathways of interest included: regulation of transcription, embryonic development (including blastocyst formation), Wnt and Hedgehog signaling, oocyte meiosis, and kinase and phosphatase activity. MicroRNA present in mature sperm appears to not only be left over from spermatogenic processes, but may actually serve important regulatory roles in fertilization and early developmental processes. Further, our results indicate the possibility that environmental

  4. MicroRNA-Sequence Profiling Reveals Novel Osmoregulatory MicroRNA Expression Patterns in Catadromous Eel Anguilla marmorata

    PubMed Central

    Li, Peng; Yin, Shaowu; Wang, Li; Jia, Yihe; Shu, Xinhua

    2015-01-01

    MicroRNAs (miRNAs) are a class of endogenous small non-coding RNAs that regulate gene expression by post-transcriptional repression of mRNAs. Recently, several miRNAs have been confirmed to execute directly or indirectly osmoregulatory functions in fish via translational control. In order to clarify whether miRNAs play relevant roles in the osmoregulation of Anguilla marmorata, three sRNA libraries of A. marmorata during adjusting to three various salinities were sequenced by Illumina sRNA deep sequencing methods. Totally 11,339,168, 11,958,406 and 12,568,964 clear reads were obtained from 3 different libraries, respectively. Meanwhile, 34 conserved miRNAs and 613 novel miRNAs were identified using the sequence data. MiR-10b-5p, miR-181a, miR-26a-5p, miR-30d and miR-99a-5p were dominantly expressed in eels at three salinities. Totally 29 mature miRNAs were significantly up-regulated, while 72 mature miRNAs were significantly down-regulated in brackish water (10‰ salinity) compared with fresh water (0‰ salinity); 24 mature miRNAs were significantly up-regulated, while 54 mature miRNAs were significantly down-regulated in sea water (25‰ salinity) compared with fresh water. Similarly, 24 mature miRNAs were significantly up-regulated, while 45 mature miRNAs were significantly down-regulated in sea water compared with brackish water. The expression patterns of 12 dominantly expressed miRNAs were analyzed at different time points when the eels transferred from fresh water to brackish water or to sea water. These miRNAs showed differential expression patterns in eels at distinct salinities. Interestingly, miR-122, miR-140-3p and miR-10b-5p demonstrated osmoregulatory effects in certain salinities. In addition, the identification and characterization of differentially expressed miRNAs at different salinities can clarify the osmoregulatory roles of miRNAs, which will shed lights for future studies on osmoregulation in fish. PMID:26301415

  5. MicroRNA gene expression during retinoic acid-induced differentiation of human acute promyelocytic leukemia.

    PubMed

    Garzon, R; Pichiorri, F; Palumbo, T; Visentini, M; Aqeilan, R; Cimmino, A; Wang, H; Sun, H; Volinia, S; Alder, H; Calin, G A; Liu, C-G; Andreeff, M; Croce, C M

    2007-06-14

    MicroRNAs (miRNAs) are small non-coding RNAs of 19-25 nucleotides that are involved in the regulation of critical cell processes such as apoptosis, cell proliferation and differentiation. However, little is known about the role of miRNAs in granulopoiesis. Here, we report the expression of miRNAs in acute promyelocytic leukemia patients and cell lines during all-trans-retinoic acid (ATRA) treatment by using a miRNA microarrays platform and quantitative real time-polymerase chain reaction (qRT-PCR). We found upregulation of miR-15a, miR-15b, miR-16-1, let-7a-3, let-7c, let-7d, miR-223, miR-342 and miR-107, whereas miR-181b was downregulated. Among the upregulated miRNAs, miR-107 is predicted to target NFI-A, a gene that has been involved in a regulatory loop involving miR-223 and C/EBPa during granulocytic differentiation. Indeed, we have confirmed that miR-107 targets NF1-A. To get insights about ATRA regulation of miRNAs, we searched for ATRA-modulated transcription factors binding sites in the upstream genomic region of the let-7a-3/let-7b cluster and identified several putative nuclear factor-kappa B (NF-kappaB) consensus elements. The use of reporter gene assays, chromatin immunoprecipitation and site-directed mutagenesis revealed that one proximal NF-kappaB binding site is essential for the transactivation of the let-7a-3/let-7b cluster. Finally, we show that ATRA downregulation of RAS and Bcl2 correlate with the activation of known miRNA regulators of those proteins, let-7a and miR-15a/miR-16-1, respectively. PMID:17260024

  6. Role of micro-RNA in colorectal cancer screening.

    PubMed

    Rodríguez-Montes, José Antonio; Menéndez Sánchez, Pablo

    2014-12-01

    MicroRNAs are involved in carcinogenesis through postranscriptional gene regulatory activity. These molecules are involved in various physiological and pathological functions, such as apoptosis, cell proliferation and differentiation, which indicates their functionality in carcinogenesis as tumour suppressor genes or oncogenes. Several studies have determined the presence of microRNAs in different neoplastic diseases such as colon, prostate, breast, stomach, pancreas, and lung cancer. There are promising data on the usefulness of quantifying microRNAs in different organic fluids and tissues. We have conducted a review of the determinations of microRNAs in the diagnosis of colorectal cancer. PMID:25088411

  7. MicroRNA-dependent Genetic Networks During Neural Development

    PubMed Central

    Abernathy, Daniel G.; Yoo, Andrew S.

    2014-01-01

    The development of the structurally and functionally diverse mammalian nervous system requires the integration of numerous levels of gene regulation. Accumulating evidence suggests that microRNAs are key mediators of genetic networks during neural development. Importantly, microRNAs are found to regulate both feedback and feedforward loops during neural development leading to large changes in gene expression. These repressive interactions provide an additional mechanism that facilitates the establishment of complexity within the nervous system. Here, we review studies that have enabled the identification of brain-enriched microRNAs and discuss how genetic networks in neural development depend on microRNAs. PMID:24865244

  8. microRNA-17–92 Regulates IL-10 Production by Regulatory T Cells and Control of Experimental Autoimmune Encephalomyelitis

    PubMed Central

    de Kouchkovsky, Dimitri; Esensten, Jonathan H.; Rosenthal, Wendy L.; Morar, Malika M.; Bluestone, Jeffrey A.; Jeker, Lukas T.

    2014-01-01

    microRNAs (miRNA) are essential for regulatory T cell (Treg) function but little is known about the functional relevance of individual miRNA loci. We identified the miR-17–92 cluster as CD28 costimulation dependent, suggesting that it may be key for Treg development and function. Although overall immune homeostasis was maintained in mice with miR-17–92–deficient Tregs, expression of the miR-17–92 miRNA cluster was critical for Treg accumulation and function during an acute organ-specific autoimmune disease in vivo. Treg-specific loss of miR-17–92 expression resulted in exacerbated experimental autoimmune encephalitis and failure to establish clinical remission. Using peptide-MHC tetramers, we demonstrate that the miR-17–92 cluster was specifically required for the accumulation of activated Ag-specific Treg and for differentiation into IL-10–producing effector Treg. PMID:23858035

  9. Establishment of cells to monitor Microprocessor through fusion genes of microRNA and GFP

    SciTech Connect

    Tsutsui, Motomu; Hasegawa, Hitoki; Adachi, Koichi; Miyata, Maiko; Huang, Peng; Ishiguro, Naoki; Hamaguchi, Michinari; Iwamoto, Takashi

    2008-08-08

    Microprocessor, the complex of Drosha and DGCR8, promotes the processing of primary microRNA to precursor microRNA, which is a crucial step for microRNA maturation. So far, no convenient assay systems have been developed for observing this step in vivo. Here we report the establishment of highly sensitive cellular systems where we can visually monitor the function of Microprocessor. During a series of screening of transfectants with fusion genes of the EGFP cDNA and primary microRNA genes, we have obtained certain cell lines where introduction of siRNA against DGCR8 or Drosha strikingly augments GFP signals. In contrast, these cells have not responded to Dicer siRNA; thus they have a unique character that GFP signals should be negatively and specifically correlated to the action of Microprocessor among biogenesis of microRNA. These cell lines can be useful tools for real-time analysis of Microprocessor action in vivo and identifying its novel modulators.

  10. MicroRNA expression is altered in lateral septum across reproductive stages.

    PubMed

    Saul, M C; Zhao, C; Driessen, T M; Eisinger, B E; Gammie, S C

    2016-01-15

    MicroRNAs (miRNAs) inhibit RNA targets and may contribute to postpartum central nervous system (CNS) gene expression changes, although this has never been tested. In the present study, we directly evaluated miRNA levels using RNA sequencing during reproduction in female mice in the lateral septum (LS). We found the reliable and robust changes of miRNAs away from the virgin stage at the three other stages, namely pregnant, day 1 postpartum, and day 8 postpartum. For a given miRNA that was significantly different from the virgin condition in more than one group, the direction of change was always the same. Overall, we identified 32 upregulated miRNAs and 25 downregulated miRNAs that were consistently different from the virgin state. 'Arm switching' occurs for miR-433-3 and miR-7b. Unexpectedly, a third of upregulated miRNAs (relative to virgin) were highly localized within the 12qF1 region of chromosome 12 that includes the Dlk1-Dio3 gene cluster implicated in stem cell and neuronal differentiation. Over 1500 genes were targeted by multiple upregulated miRNAs with about 100 genes targeted by five or more miRNAs. Over 1000 genes were targeted by multiple downregulated miRNAs with about 50 genes targeted by five or more miRNAs. Half of the target genes were regulated by up and downregulated miRNAs, indicating homeostatic regulation. Transcriptional regulation was the most enriched pathway for genes linked to up or down regulated miRNAs. Other enriched pathways included protein kinase activity (e.g., MAP kinase), CNS development, axon guidance, neurotrophin signaling, neuron development/differentiation, and neurogenesis. Previously published postpartum LS gene expression changes were enrichment for LS miRNA targets, as expected. Surprisingly, postpartum gene expression changes from other regions were also enriched against LS miRNA targets, suggesting a core group of miRNAs may act across the CNS during reproduction. Together, we directly examine miRNAs and find

  11. miRepress: modelling gene expression regulation by microRNA with non-conventional binding sites

    PubMed Central

    Ghosal, Suman; Saha, Shekhar; Das, Shaoli; Sen, Rituparno; Goswami, Swagata; Jana, Siddhartha S.; Chakrabarti, Jayprokas

    2016-01-01

    Some earlier studies have reported an alternative mode of microRNA-target interaction. We detected target regions within mRNA transcripts from AGO PAR-CLIP that did not contain any conventional microRNA seed pairing but only had non-conventional binding sites with microRNA 3′ end. Our study from 7 set of data that measured global protein fold change after microRNA transfection pointed towards the association of target protein fold change with 6-mer and 7-mer target sites involving microRNA 3′ end. We developed a model to predict the degree of microRNA target regulation in terms of protein fold changes from the number of different conventional and non-conventional target sites present in the target, and found significant correlation of its output with protein expression changes. We validated the effect of non-conventional interactions with target by modulating the abundance of microRNA in a human breast cancer cell line MCF-7. The validation was done using luciferase assay and immunoblot analysis for our predicted non-conventional microRNA-target pair WNT1 (3′ UTR) and miR-367-5p and immunoblot analysis for another predicted non-conventional microRNA-target pair MYH10 (coding region) and miR-181a-5p. Both experiments showed inhibition of targets by transfection of microRNA mimics that were predicted to have only non-conventional sites. PMID:26923536

  12. Predicting Large RNA-Like Topologies by a Knowledge-Based Clustering Approach.

    PubMed

    Baba, Naoto; Elmetwaly, Shereef; Kim, Namhee; Schlick, Tamar

    2016-02-27

    An analysis and expansion of our resource for classifying, predicting, and designing RNA structures, RAG (RNA-As-Graphs), is presented, with the goal of understanding features of RNA-like and non-RNA-like motifs and exploiting this information for RNA design. RAG was first reported in 2004 for cataloging RNA secondary structure motifs using graph representations. In 2011, the RAG resource was updated with the increased availability of RNA structures and was improved by utilities for analyzing RNA structures, including substructuring and search tools. We also classified RNA structures as graphs up to 10 vertices (~200 nucleotides) into three classes: existing, RNA-like, and non-RNA-like using clustering approaches. Here, we focus on the tree graphs and evaluate the newly founded RNAs since 2011, which also support our refined predictions of RNA-like motifs. We expand the RAG resource for large tree graphs up to 13 vertices (~260 nucleotides), thereby cataloging more than 10 times as many secondary structures. We apply clustering algorithms based on features of RNA secondary structures translated from known tertiary structures to suggest which hypothetical large RNA motifs can be considered "RNA-like". The results by the PAM (Partitioning Around Medoids) approach, in particular, reveal good accuracy, with small error for the largest cases. The RAG update here up to 13 vertices offers a useful graph-based tool for exploring RNA motifs and suggesting large RNA motifs for design. PMID:26478223

  13. Functional interplay between RNA-binding protein HuR and microRNAs

    PubMed Central

    Srikantan, Subramanya; Tominaga, Kumiko; Gorospe, Myriam

    2012-01-01

    The mammalian RNA-binding protein (RBP) HuR associates with numerous mRNAs encoding proteins with roles in cell division, cell survival, immune response, and differentiation. HuR was known to stabilize many of these mRNAs and/or modulated their translation, but the molecular processes by which HuR affected the fate of target mRNAs was largely unknown. Evidence accumulated over the past five years has revealed that the influence of HuR on many bound transcripts depends on HuR's interplay with microRNAs which associate with the same mRNAs. Here, we review the interactions of HuR and microRNAs – both competitive and cooperative – that govern expression of shared target mRNAs. Competition between HuR and microRNAs typically results in enhanced gene expression if the HuR-mRNA interaction prevails, and in repression if the microRNA remains associated. Cooperation between HuR and microRNAs leads to lower expression of the shared mRNA. We also describe the regulation of HuR levels by microRNAs as well as the regulation of microRNA levels by HuR. Finally, we discuss transcriptome-wide analyses of HuR-bound mRNAs with neighboring microRNA sites, and review the emerging mechanisms whereby microRNAs confer versatility and strength to the post-transcriptional outcomes of HuR targets. PMID:22708488

  14. MicroRNAfold: pre-microRNA secondary structure prediction based on modified NCM model with thermodynamics-based scoring strategy.

    PubMed

    Han, Dianwei; Zhang, Jun; Tang, Guiliang

    2012-01-01

    An accurate prediction of the pre-microRNA secondary structure is important in miRNA informatics. Based on a recently proposed model, nucleotide cyclic motifs (NCM), to predict RNA secondary structure, we propose and implement a Modified NCM (MNCM) model with a physics-based scoring strategy to tackle the problem of pre-microRNA folding. Our microRNAfold is implemented using a global optimal algorithm based on the bottom-up local optimal solutions. Our experimental results show that microRNAfold outperforms the current leading prediction tools in terms of True Negative rate, False Negative rate, Specificity, and Matthews coefficient ratio. PMID:23155762

  15. Roles of microRNA-99 family in human glioma

    PubMed Central

    Zhang, Mingyu; Guo, Yong; Wu, Jun; Chen, Fenghua; Dai, Zhijie; Fan, Shuangshi; Li, Pengcheng; Song, Tao

    2016-01-01

    Objective Deregulation of microRNA (miR)-99 family members (miR-99a, miR-99b, and miR-100) has been reported to play a crucial role in many cancer types. However, their roles in human gliomas have not been fully elucidated. This study aimed to investigate the expression patterns of miR-99a, miR-99b, and miR-100 in glioma tissues and to evaluate their expression profiles with respect to tumor progression. Methods Quantitative real-time polymerase chain reaction was performed to detect the expression levels of miR-99a, miR-99b, and miR-100 in glioma and matched non-neoplastic brain tissues. Then, the associations of their expression with various clinicopathological features of glioma patients were statistically analyzed. Moreover, the roles of miR-99a, miR-99b, and miR-100 in regulating glioma cell migration and invasion were determined via transwell assay in vitro. Results Compared with non-neoplastic brain tissues, miR-99a, miR-99b, and miR-100 expression levels were all significantly decreased in glioma tissues (all P<0.001). miR-99a-low, miR-99b-low, and miR-100-low expression more frequently occurred in glioma patients with low Karnofsky performance score (<90) and high World Health Organization grade (III–IV). Further functional experiments revealed that the enforced expression of miR-99a, miR-99b, and miR-100 resulted in the inhibition of cellular migration and invasion in glioma cells. Conclusion Our results strongly suggest that the aberrant expression of miR-99a, miR-99b, and miR-100 may be a common feature in human gliomas with aggressive clinicopathological features and may participate in malignant phenotypes of the tumors. These findings highlight the potential of the three miR-99 family members as novel therapeutic targets for human gliomas. PMID:27382299

  16. MicroRNA in late lung development and bronchopulmonary dysplasia: the need to demonstrate causality.

    PubMed

    Nardiello, Claudio; Morty, Rory E

    2016-12-01

    MicroRNA are emerging as powerful regulators of cell differentiation and tissue and organ development. Several microRNA have been described to play a role in branching morphogenesis, a key step in early lung development. However, considerably less attention has been paid to microRNA as regulators of the process of secondary septation, which drives lung alveolarization during late lung development. Secondary septation is severely perturbed in bronchopulmonary dysplasia (BPD), a common complication of preterm birth characterized by blunted alveolarization. A number of studies to date have reported microRNA microarray screens in animal models of BPD; however, only two studies have attempted to demonstrate causality. Although the expression of miR-150 was altered in experimental BPD, a miR-150(-/-) knockout mouse did not exhibit appreciable protection in a BPD animal model. Similarly, while the expression of miR-489 in the lung was reduced in clinical and experimental BPD, antagomiR and over-expression approaches could not validate a role for miR-489 in the impaired alveolarization associated with experimental BPD. This mini-review aims to highlight microRNA that have been revealed by multiple microarray studies to be potential causal players in normal and pathological alveolarization. Additionally, the challenges faced in attempting to demonstrate a causal role for microRNA in lung alveolarization are discussed. These include the tremendous variability in the animal models employed, and the limitations and advantages offered by the available tools, including antagomiRs and approaches for the validation of a specific microRNA-mRNA interaction during lung alveolarization. PMID:27216745

  17. Topological patterns in microRNA-gene regulatory network: studies in colorectal and breast cancer.

    PubMed

    Sengupta, Debarka; Bandyopadhyay, Sanghamitra

    2013-06-01

    It is now widely accepted that microRNAs (miRNAs or miRs) along with transcription factors (TFs) weave a complex inter-regulatory network within the cell that is responsible for the combinatorial regulation of gene expression. Recently we have shown that miRNAs and TFs that form network clusters are also associated with a number of common diseases. However, the quest persists to find out topological structures that facilitate disease progression. In the current work we choose colorectal and breast cancers for our analysis. For this, the human genome wide TF-miRNA-gene network (TMG-net) is first built by combining experimentally validated and confidently predicted miRNA→gene (including TF genes), TF→gene and TF→miRNA interactions. Subnetworks active in colorectal and breast cancers are extracted from the TMG-net and then analyzed. Disease specific subnetworks are found to be significantly dense, having a pyramid shaped hierarchical backbone of interactions. Interestingly, most of the top level molecules (e.g., hsa-mir-210, hsa-mir-378) are found to be already established as oncomirs. TFs that are dysregulated in a particular cancer, are found to be well-linked via miRNAs and other TFs, with miRNAs being highly predominant. Analogous to density, a new measure called Inductive Converge (InCov) is proposed and used to analyze the natural association of molecules in the disease specific networks. Finally a web application called DisTMGneT (Disease Specific TF-miRNA-gene Network) is developed for disease specific subnetworks from the TMG-net, based on user supplied sets of dysregulated miRNAs, TFs and non TF genes. DisTMGneT is available at http://www.isical.ac.in/bioinfo_miu/dscsgen.php. PMID:23475160

  18. Subtype-specific micro-RNA expression signatures in breast cancer progression.

    PubMed

    Haakensen, Vilde D; Nygaard, Vegard; Greger, Liliana; Aure, Miriam R; Fromm, Bastian; Bukholm, Ida R K; Lüders, Torben; Chin, Suet-Feung; Git, Anna; Caldas, Carlos; Kristensen, Vessela N; Brazma, Alvis; Børresen-Dale, Anne-Lise; Hovig, Eivind; Helland, Åslaug

    2016-09-01

    Robust markers of invasiveness may help reduce the overtreatment of in situ carcinomas. Breast cancer is a heterogeneous disease and biological mechanisms for carcinogenesis vary between subtypes. Stratification by subtype is therefore necessary to identify relevant and robust signatures of invasive disease. We have identified microRNA (miRNA) alterations during breast cancer progression in two separate datasets and used stratification and external validation to strengthen the findings. We analyzed two separate datasets (METABRIC and AHUS) consisting of a total of 186 normal breast tissue samples, 18 ductal carcinoma in situ (DCIS) and 1,338 invasive breast carcinomas. Validation in a separate dataset and stratification by molecular subtypes based on immunohistochemistry, PAM50 and integrated cluster classifications were performed. We propose subtype-specific miRNA signatures of invasive carcinoma and a validated signature of DCIS. miRNAs included in the invasive signatures include downregulation of miR-139-5p in aggressive subtypes and upregulation of miR-29c-5p expression in the luminal subtypes. No miRNAs were differentially expressed in the transition from DCIS to invasive carcinomas on the whole, indicating the need for subtype stratification. A total of 27 miRNAs were included in our proposed DCIS signature. Significant alterations of expression included upregulation of miR-21-5p and the miR-200 family and downregulation of let-7 family members in DCIS samples. The signatures proposed here can form the basis for studies exploring DCIS samples with increased invasive potential and serum biomarkers for in situ and invasive breast cancer. PMID:27082076

  19. MicroRNA Expression In Lymphohematopoietic Malignancies And Following Formaldehyde Exposure [Poster 2015

    EPA Science Inventory

    Altered microRNA (miRNA) expression is an emerging area that promises future identification of epigenetic biomarkers of disease and exposure to environmental agents. In addition to other carcinogenic mechanisms, such as genotoxicity, miRNAs have been shown to play an important r...

  20. MicroRNA Expression In Lymphohematopoietic Malignancies And Following Formaldehyde Exposure

    EPA Science Inventory

    Altered microRNA (miRNA) expression is an emerging area promising future identification of epigenetic biomarkers of disease and exposure to environmental agents. In addition to other carcinogenic mechanisms, such as genotoxicity, miRNAs have been shown to play an important role ...

  1. MicroRNA: mechanism of gene regulation and application to livestock

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNA (miR) are a class of small RNAs that regulate gene expression by inhibiting translation of protein encoding transcripts through activation of a specific cellular pathway. The small RNA classified as miR are short sequences of 18-26 nucleotide long, encoded by nuclear genes with distinctive...

  2. microRNA (miRNA)-Mediated Pathogenetic Signaling in Alzheimer's Disease (AD).

    PubMed

    Hill, James M; Lukiw, Walter J

    2016-02-01

    Alzheimer's disease (AD) is an expanding health and socioeconomic concern in industrialized societies, and the leading cause of intellectual impairment in our aging population. The cause of AD remains unknown, and there are currently no effective treatments to stop or reverse the progression of this uniquely human and age-related neurological disorder. Elucidation of the AD mechanism and factors that contribute to the initiation, progression, and spreading of this chronic and fatal neurodegeneration will ultimately result in improved and effective diagnostics and therapeutic strategies.microRNAs (miRNAs) comprise a relatively recently discovered category of 20-24 nucleotide non-coding RNAs that function post-transcriptionally in shaping the transcriptome of the cell, and in doing so, contribute to the molecular-genetics and phenotype of human CNS health and disease. To date about 2550 unique mature human miRNAs have been characterized, however only highly selected miRNA populations appear to be enriched in different anatomical compartments within the CNS.This general commentary for the 'Special Issue: 40th Year of Neurochemical Research' will bring into perspective (i) some very recent findings on the extraordinary biophysics and signaling properties of CNS miRNA in AD and aging human brain; (ii) how specific intrinsic biophysical attributes of miRNAs may play defining roles in the establishment, proliferation and spreading of the AD phenotype; and (iii) how miRNAs can serve as prospective therapeutic targets and biomarkers potentially useful in the clinical management of this terminal neurological disease whose incidence in our rapidly aging population is reaching epidemic proportions. PMID:26441222

  3. Sequence analysis of a cluster of twenty-one tRNA genes in Bacillus subtilis.

    PubMed Central

    Green, C J; Vold, B S

    1983-01-01

    The DNA sequence of a cluster of twenty-one tRNA genes distal to a rRNA gene set in B. subtilis was determined. None of the tRNA genes are repeated in the sequence. The only classes of tRNAs that are not represented are those for cysteine, glutamine, tryptophan, and tyrosine. Three of the tRNA genes in this cluster do not have the 3'-CCA sequence encoded in the gene. There is no RNA polymerase terminator sequence in the region between the 5S gene and the first tRNA gene or within the tRNA gene cluster. A terminator sequence was found directly after the last tRNA gene. This rRNA and tRNA gene cluster probably represents one transcriptional unit. However, there may be an RNA polymerase promoter site within this sequence, which raises some interesting questions concerning the regulation of transcription for these tRNA genes. PMID:6310512

  4. MicroRNA-34b promoter hypermethylation induces CREB overexpression and contributes to myeloid transformation.

    PubMed

    Pigazzi, Martina; Manara, Elena; Bresolin, Silvia; Tregnago, Claudia; Beghin, Alessandra; Baron, Emma; Giarin, Emanuela; Cho, Er-Chieh; Masetti, Riccardo; Rao, Dinesh S; Sakamoto, Kathleen M; Basso, Giuseppe

    2013-04-01

    MicroRNA-34b down-regulation in acute myeloid leukemia was previously shown to induce CREB overexpression, thereby causing leukemia proliferation in vitro and in vivo. The role of microRNA-34b and CREB in patients with myeloid malignancies has never been evaluated. We examined microRNA-34b expression and the methylation status of its promoter in cells from patients diagnosed with myeloid malignancies. We used gene expression profiling to identify signatures of myeloid transformation. We established that microRNA-34b has suppressor ability and that CREB has oncogenic potential in primary bone marrow cell cultures and in vivo. MicroRNA-34b was found to be up-regulated in pediatric patients with juvenile myelomonocytic leukemia (n=17) and myelodysplastic syndromes (n=28), but was down-regulated in acute myeloid leukemia patients at diagnosis (n=112). Our results showed that hypermethylation of the microRNA-34b promoter occurred in 66% of cases of acute myeloid leukemia explaining the low microRNA-34b levels and CREB overexpression, whereas preleukemic myelodysplastic syndromes and juvenile myelomonocytic leukemia were not associated with hypermethylation or CREB overexpression. In paired samples taken from the same patients when they had myelodysplastic syndrome and again during the subsequent acute myeloid leukemia, we confirmed microRNA-34b promoter hypermethylation at leukemia onset, with 103 CREB target genes differentially expressed between the two disease stages. This subset of CREB targets was confirmed to associate with high-risk myelodysplastic syndromes in a separate cohort of patients (n=20). Seventy-eight of these 103 CREB targets were also differentially expressed between healthy samples (n=11) and de novo acute myeloid leukemia (n=72). Further, low microRNA-34b and high CREB expression levels induced aberrant myelopoiesis through CREB-dependent pathways in vitro and in vivo. In conclusion, we suggest that microRNA-34b controls CREB expression and

  5. Altered RNA editing in 3′ UTR perturbs microRNA-mediated regulation of oncogenes and tumor-suppressors

    PubMed Central

    Zhang, Liye; Yang, Chih-Sheng; Varelas, Xaralabos; Monti, Stefano

    2016-01-01

    RNA editing is a molecular event that alters specific nucleotides in RNA post-transcriptionally. RNA editing has the potential to impact a variety of cellular processes and is implicated in diseases such as cancer. Yet, the precise mechanisms by which RNA editing controls cellular processes are poorly understood. Here, we characterize sequences altered by RNA editing in patient samples from lymphoma, neuroblastoma and head and neck cancers. We show that A-to-I RNA editing sites are highly conserved across samples of the same tissue type and that most editing sites identified in tumors are also detectable in normal tissues. Next, we identify the significant changes in editing levels of known sites between tumor and paired “normal” tissues across 14 cancer types (627 pairs) from The Cancer Genome Atlas project and show that the complexity of RNA editing regulation cannot be captured by the activity of ADAR family genes alone. Our pan-cancer analysis confirms previous results on individual tumor types and suggests that changes of RNA editing levels in coding and 3′UTR regions could be a general mechanism to promote tumor growth. We also propose a model explaining how altered RNA editing levels affect microRNA-mediated post-transcriptional regulation of oncogenes and tumor-suppressors. PMID:26980570

  6. Nucleolin mediates microRNA-directed CSF-1 mRNA deadenylation but increases translation of CSF-1 mRNA.

    PubMed

    Woo, Ho-Hyung; Baker, Terri; Laszlo, Csaba; Chambers, Setsuko K

    2013-06-01

    CSF-1 mRNA 3'UTR contains multiple unique motifs, including a common microRNA (miRNA) target in close proximity to a noncanonical G-quadruplex and AU-rich elements (AREs). Using a luciferase reporter system fused to CSF-1 mRNA 3'UTR, disruption of the miRNA target region, G-quadruplex, and AREs together dramatically increased reporter RNA levels, suggesting important roles for these cis-acting regulatory elements in the down-regulation of CSF-1 mRNA. We find that nucleolin, which binds both G-quadruplex and AREs, enhances deadenylation of CSF-1 mRNA, promoting CSF-1 mRNA decay, while having the capacity to increase translation of CSF-1 mRNA. Through interaction with the CSF-1 3'UTR miRNA common target, we find that miR-130a and miR-301a inhibit CSF-1 expression by enhancing mRNA decay. Silencing of nucleolin prevents the miRNA-directed mRNA decay, indicating a requirement for nucleolin in miRNA activity on CSF-1 mRNA. Downstream effects followed by miR-130a and miR-301a inhibition of directed cellular motility of ovarian cancer cells were found to be dependent on nucleolin. The paradoxical effects of nucleolin on miRNA-directed CSF-1 mRNA deadenylation and on translational activation were explored further. The nucleolin protein contains four acidic stretches, four RNA recognition motifs (RRMs), and nine RGG repeats. All three domains in nucleolin regulate CSF-1 mRNA and protein levels. RRMs increase CSF-1 mRNA, whereas the acidic and RGG domains decrease CSF-1 protein levels. This suggests that nucleolin has the capacity to differentially regulate both CSF-1 RNA and protein levels. Our finding that nucleolin interacts with Ago2 indirectly via RNA and with poly(A)-binding protein C (PABPC) directly suggests a nucleolin-Ago2-PABPC complex formation on mRNA. This complex is in keeping with our suggestion that nucleolin may work with PABPC as a double-edged sword on both mRNA deadenylation and translational activation. Our findings underscore the complexity of

  7. A Critical Evaluation of microRNA Biomarkers in Non-Neoplastic Disease

    PubMed Central

    Haider, Baqer A.; Baras, Alexander S.; McCall, Matthew N.; Hertel, Joshua A.; Cornish, Toby C.; Halushka, Marc K.

    2014-01-01

    Background MicroRNAs (miRNAs) are small (∼22-nt), stable RNAs that critically modulate post-transcriptional gene regulation. MicroRNAs can be found in the blood as components of serum, plasma and peripheral blood mononuclear cells (PBMCs). Many microRNAs have been reported to be specific biomarkers in a variety of non-neoplastic diseases. To date, no one has globally evaluated these proposed clinical biomarkers for general quality or disease specificity. We hypothesized that the cellular source of circulating microRNAs should correlate with cells involved in specific non-neoplastic disease processes. Appropriate cell expression data would inform on the quality and usefulness of each microRNA as a biomarker for specific diseases. We further hypothesized a useful clinical microRNA biomarker would have specificity to a single disease. Methods and Findings We identified 416 microRNA biomarkers, of which 192 were unique, in 104 publications covering 57 diseases. One hundred and thirty-nine microRNAs (33%) represented biologically plausible biomarkers, corresponding to non-ubiquitous microRNAs expressed in disease-appropriate cell types. However, at a global level, many of these