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NATURE GENETICS SUPPLEMENT | VOLUME 38 | JUNE 2006 P E R S P E C T I V E S : MOLECULAR BIOLOGY Glimpses of a Tiny RNA World. SCIENCE VOL 294 26 OCTOBER 2001 Gary Ruvkun Functions
of microRNAs and related small RNAs in plants.
NATURE GENETICS SUPPLEMENT | VOLUME 38 | JUNE 2006 Allison C Mallory & Hervé Vaucheret MicroRNAs (miRNAs)
and short interfering RNAs (siRNAs), 20- to 27-nt in length, are
essential regulatory molecules that act as sequence-specific
guides in several processes in most eukaryotes (with the notable
exception of the yeast Saccharomyces cerevisiae). These
processes include DNA elimination, heterochromatin assembly, mRNA
cleavage and translational repression. This review focuses
on the regulatory roles of plant miRNAs during development, in the
adaptive response to stresses and in the miRNA pathway itself.
This review also covers the regulatory roles of two classes of
endogenous plant siRNAs, ta-siRNAs and nat-siRNAs, which
participate in post-transcriptional control of gene expression.
Evidence
that microRNAs are associated with translating messenger RNAs in human
cells.
VOLUME 13 NUMBER 12 DECEMBER 2006 NATURE STRUCTURAL & MOLECULAR BIOLOGY Patricia A Maroney, Yang Yu, Jesse Fisher & Timothy W Nilsen MicroRNAs (miRNAs)
regulate gene expression post-transcriptionally by binding the 3¢
untranslated regions of target mRNAs. We examined the
subcellular distribution of three miRNAs in exponentially growing HeLa
cells and found that the vast majority are associated with
mRNAs in polysomes. Several lines of evidence indicate that most of
these mRNAs, including a known miRNA-regulated
target (KRAS mRNA), are actively being translated.
How
microRNAs control cell division, differentiation and death.
Eric A Miska Current Opinion in Genetics & Development 2005, 15: 563–568 After the milestone
discovery of the first microRNA in 1993, the past five years have
seen a phenomenal surge of interest in these short,
regulatory RNAs. Given that 2% of all known human genes encode
microRNAs, one main goal is to uncover microRNA function.
Although it has been more difficult to assign function to
microRNAs in animals than it has been in plants, important
roles are emerging: in invertebrates, microRNAs control
developmental timing, neuronal differentiation,
tissue growth and programmed cell death. Functional studies in
zebrafish and mice point toward important roles for microRNAs
during morphogenesis and organogenesis.
Finally, microRNAs might regulate viral infection and human
cancer.
microRNA
target predictions in animals.
In recent
years, microRNAs (miRNAs) have emerged as a major class of regulatory
genes, present in most metazoans and important for a
diverse range of biological functions. Because experimental
identification of miRNA targets is difficult, there has been an
explosion
of computational target predictions. Although the initial
round of predictions resulted in very diverse results,
subsequent computational and experimental analyses suggested
that at least a certain class of conserved miRNA targets can be
confidently predicted and that this class of targets is
large, covering, for example, at least 30% of all human genes
when considering about 60 conserved vertebrate miRNA gene
families. Most recent approaches have also shown that
there are correlations between domains of miRNA expression
and mRNA levels of their targets. Our understanding of
miRNA function is still extremely limited, but it may be that by
integrating mRNA and miRNA sequence and expression data with
other comparative genomic data, we will be able to gain
global and yet specific insights into the function and evolution of a
broad layer of post-transcriptional control.VOLUME 38 | JUNE 2006 | NATURE GENETICS SUPPLEMENT Nikolaus Rajewsky Human
let-7a miRNA blocks protein production on actively translating
polyribosomes.
MicroRNAs
(miRNAs) regulate gene expression at a post-transcriptional level
through base-pairing to 3¢ untranslated regions (UTRs) of
messenger
RNAs. The mechanism by which human let-7a miRNA regulates mRNA
translation was examined in HeLa cells expressing
reporter mRNAs containing the Caenorhabditis elegans lin-41 3¢
UTR. let-7a miRNA strongly repressed translation, yet the
majority of control and lin-41–bearing RNAs sedimented with
polyribosomes in sucrose gradients; these polyribosomes,
together with let-7a miRNA and the miRISC protein AGO, were released
from those structures by puromycin. RNA containing the
lin-41 3¢ UTR and an iron response element in the 5¢ UTR
sedimented with polysomes when cells were incubated with iron,
but showed ribosome run-off when the iron was chelated. These data
indicate that let-7a miRNA inhibits actively translating
polyribosomes. Nascent polypeptide coimmunoprecipitation experiments
further suggest that let-7a miRNA interferes with the
accumulation of growing polypeptides.VOLUME 13 NUMBER 12 DECEMBER 2006 NATURE STRUCTURAL & MOLECULAR BIOLOGY Stephanie Nottrott, Martin J Simard & Joel D Richter Computational
identification of microRNA targets.
Developmental Biology 267 (2004) 529– 535 Nikolaus Rajewskya, and Nicholas D. Soccib, Department of Biology, New York University, New York, NY 10003-6688, USA; Department of Pathology, and Seaver Foundation for Bioinformatics, Albert Einstein College of Medicine, Bronx, NY 10461, USA; Computational Biology Center, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA Recent experiments
have shown that the genomes of organisms such as worm, fly, human, and
mouse encode hundreds of microRNA genes. Many of these
microRNAs are thought to regulate the translational expression of other
genes by binding to partially complementary sites in messenger
RNAs. Phenotypic and expression analysis suggests an important role of
microRNAs during development. Therefore, it is of fundamental
importance to identify microRNA targets. However, no experimental or
computational high-throughput method for target site identification in
animals has been published yet. Our main result is a new computational
method that is designed to identify microRNA target sites. This method
recovers with high specificity known microRNA target sites that have
previously been defined experimentally. Based on these results, we
present a simple model for the mechanism of microRNA target site
recognition. Our model incorporates both kinetic and thermodynamic
components of target recognition. When we applied our method to a set
of 74 Drosophila melanogaster microRNAs, searching 3V UTR
sequences of a predefined set of fly mRNAs for target sites which were
evolutionary conserved between D. melanogaster and Drosophila
pseudoobscura, we found that many key developmental body patterning
genes such as hairy and fushi-tarazu are likely to be translationally
regulated by microRNAs.
Incorporating
structure to predict microRNA targets.
PNAS 2005 vol. 102 no. 11, 4006–4009 Harlan Robins*†, Ying Li‡, and Richard W. Padgett‡ *Institute for Advanced Study, Olden Lane, Princeton, NJ 08540; and ‡Department of Molecular Biology and Biochemistry, Waksman Institute, Cancer Institute of New Jersey, Rutgers, The State University of New Jersey, Piscataway, NJ 08854-1020 MicroRNAs (miRNAs) are a recently discovered set of regulatory genes that constitute up to an estimated 1% of the total number of genes in animal genomes, including Caenorhabditis elegans, Drosophila, mouse, and humans [Lagos-Quintana, M., Rauhut, R., Lendeckel, W. & Tuschl, T. (2001) Science 294, 853–858; Lai, E. C., Tomancak, P., Williams, R. W. & Rubin, G.M. (2003) Genome Biol. 4, R42; Lau, N. C., Lim, L. P., Weinstein, E. G. & Bartel, D. P. (2001) Science 294, 858–862; Lee, R. C. & Ambros, V. (2001) Science 294, 862-8644; and Lee, R. C., Feinbaum, R. L. & Ambros, V. (1993) Cell 115, 787–798]. In animals, miRNAs regulate genes by attenuating protein translation through imperfect base pair binding to 3 UTR sequences of target genes. A major challenge in understanding the regulatory role of miRNAs is to accurately predict regulated targets. We have developed an algorithm for predicting targets that does not rely on evolutionary conservation. As one of the features of this algorithm, we incorporate the folded structure of mRNA. By using Drosophila miRNAs as a test case, we have validated our predictions in 10 of 15 genes tested. One of these validated genes is mad as a target for bantam. Furthermore, our computational and experimental data suggest that miRNAs have fewer targets than previously reported. Nature Letters Vol 448:5 2007 J. Graham Ruby1,2*, Calvin H. Jan1,2* & David P. Bartel1,2 MicroRNAs (miRNAs)
are 22-nucleotide endogenous RNAs that often repress
the expression of complementary messenger RNAs1. In animals,
miRNAs derive from characteristic hairpins in primary
transcripts through two sequential RNase III-mediated cleavages; Drosha
cleaves near the base of the stem to liberate a 60-nucleotide
pre-miRNA hairpin, then Dicer cleaves near the loop to generate a
miRNA:miRNA* duplex2,3. From that duplex, the mature miRNA is
incorporated into the silencing complex. Here we identify an
alternative pathway for miRNA biogenesis, in which certain
debranched introns mimic the structural features of pre-miRNAs to
enter the miRNA-processing pathway without Drosha-mediated
cleavage. We call these pre-miRNAs/introns ‘mirtrons’, and have
identified 14 mirtrons in Drosophila melanogaster and another four in
Caenorhabditis elegans (including the reclassification of
mir-62). Some of these have been selectively maintained during
evolution with patterns of sequence conservation suggesting important
regulatory functions in the animal. The abundance of
introns comparable in size to pre-miRNAs appears to have
created a context favourable for the emergence of mirtrons in flies and
nematodes. This suggests that other lineages with many similarly
sized introns probably also have mirtrons, and that the mirtron
pathway could have provided an early avenue for the emergence of
miRNAs before the advent of Drosha.
MicroInspector: a web tool for detection of miRNA binding sites in an RNA sequence. Nucleic Acids Research, 2005, Vol. 33, Web Server issue W696–W700 Ventsislav Rusinov1,2, Vesselin Baev1,2, Ivan Nikiforov Minkov2 and Martin Tabler1,* 1Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology—Hellas, PO Box 1527, GR -71110 Heraklion /Crete, Greece and 2 Depar tment of Pla nt Phys iology and Mole cular Biology, Universit y of Plo vdiv 24, Tsar Assen St, 4000 Plovdiv, Bulgaria Regulation
ofpost-transcriptionalgeneexpressionby microRNAs (miRNA) has
so far been validated for only a few mRNA targets.
Based on the large number of miRNA genes and the
possibility that one miRNA might influence gene
expression of several targets simultaneously, the
quantity of ribo-regulated genes is expected to be
much higher. Here, we describe the web tool
MicroInspector that will analyse a userdefined RNA sequence, which
is typically an mRNA or a part of an mRNA,
for the occurrence of binding sites for known and
registered miRNAs. The program allows variation of
temperature, the setting of energy values as well as the
selection of different miRNA databases to identify
miRNA-binding sites of different strength.
MicroInspector could spot the correct sites for miRNA-interaction
in known target mRNAs. Using other mRNAs, for
which such an interaction has not yet been described,
we discovered frequently potential miRNA binding sites
of similar quality, which can now be analysed
experimentally. The MicroInspector program is easy to
use and does not require specific computer skills. The
service can be accessed via the MicroInspector
web server at http://www.imbb.forth.gr/microinspector
Target
labelling for
the detection and profiling of microRNAs expressed in CNS tissue using
microarrays.BMC Biotechnology 2006, 6:47 Reuben Saba1,2 and Stephanie A Booth*1,2 1Division of Host Genetics & Prion Diseases, National Microbiology Laboratory, 1015 Arlington Street, Public Health Agency of Canada, Winnipeg, MB, R3E 3R2, Canada and 2Department of Medical Microbiology and Infectious Diseases, Faculty of Medicine, University of Manitoba, Winnipeg, MB, R3B 1Y6, Canada Background: MicroRNAs
(miRNA) are a novel class of small, non-coding, gene regulatory RNA
molecules that have diverse roles in a variety of eukaryotic biological
processes. High-throughput detection and differential expression
analysis of these molecules, by microarray technology, may contribute
to a greater understanding of the many biological events regulated by
these molecules. In this investigation we compared two different
methodologies for the preparation of labelled miRNAs from mouse CNS
tissue for microarray analysis. Labelled miRNAs were prepared either by
a procedure involving linear amplification of miRNAs (labelled-aRNA) or
using a direct labelling strategy (labelled-cDNA) and analysed using a
custom miRNA microarray platform. Our aim was to develop a rapid,
sensitive methodology to profile miRNAs that could be adapted for use
on limited amounts of tissue. Results: We demonstrate the detection of
an equivalent set of miRNAs from mouse CNS tissues using both amplified
and non-amplified labelled miRNAs. Validation of the expression of
these miRNAs in the CNS by multiplex real-time PCR confirmed the
reliability of our microarray platform. We found that although the
amplification step increased the sensitivity of detection of miRNAs, we
observed a concomitant decrease in specificity for closely related
probes, as well as increased variation introduced by dye bias.
Conclusion: The data presented in this investigation identifies several
important sources of systematic bias that must be considered upon
linear amplification of miRNA for microarray analysis in comparison to
directly labelled miRNA.
A brain-specific microRNA regulates dendritic spine development. Gerhard M. Schratt1,2,3, Fabian Tuebing4, Elizabeth A. Nigh1,2,3, Christina G. Kane1,2,3, Mary E. Sabatini3, Michael Kiebler4 & Michael E. Greenberg1,2,3 Nature Vol 439:19 January 2006 MicroRNAs are small, non-coding RNAs that control the translation of target messenger RNAs, thereby regulating critical aspects of plant and animal development. In the mammalian nervous system, the spatiotemporal control of mRNA translation has an important role in synaptic development and plasticity. Although a number of microRNAs have been isolated from the mammalian brain, neither the specific microRNAs that regulate synapse function nor their target mRNAs have been identified. Here we show that a brain-specific microRNA, miR-134, is localized to the synaptodendritic compartment of rat hippocampal neurons and negatively regulates the size of dendritic spines-postsynaptic sites of excitatory synaptic transmission. This effect is mediated by miR-134 inhibition of the translation of an mRNA encoding a protein kinase, Limk1, that controls spine development. Exposure of neurons to extracellular stimuli such as brain-derived neurotrophic factor relieves miR-134 inhibition of Limk1 translation and in this way may contribute to synaptic development, maturation and/or plasticity. Oncogene (2006), 1–5 M-L Si, S Zhu, H Wu, Z Lu, F Wu and Y-Y Mo Department of Medical Microbiology, Immunology and Cell Biology, Southern Illinois University School of Medicine, Springfield, IL, USA MicroRNAs (miRNAs)
are B22 nucleotide non-coding RNA molecules that
regulate gene expression post-transcriptionally. Although aberrant
expression of miRNAs in various human cancers
suggests a role for miRNAs in tumorigenesis, it
remains largely unclear as to whether knockdown of a
specific miRNA affects tumor growth. In this study, we
profiled miRNA expression in matched normal breast tissue and
breast tumor tissues by TaqMan real-time polymerase chain
reaction miRNA array methods. Consistent with previous
findings, we found that miR-21 was highly overexpressed in
breast tumors compared to the matched normal breast tissues
among 157 human miRNAs analysed. To better evaluate
the role of miR-21 in tumorigenesis, we
transfected breast cancer MCF-7 cells with anti-miR-21 oligonucleotides and found that anti-miR-21 suppressed both cell growth in vitro and tumor growth in the xenograft mouse model. Furthermore, this anti-miR-21-mediated cell growth inhibition was associated with increased apoptosis and decreased cell proliferation, which could be in part owing to downregulation of the antiapoptotic Bcl-2 in anti-miR-21- treated tumor cells. Together, these results suggest that miR-21 functions as an oncogene and modulates tumorigenesis through regulation of genes such as bcl-2 and thus, it may serve as a novel therapeutic target. Discovering
microRNAs from deep sequencing data using miRDeep
The capacity of highly parallel sequencing technologies to
detect small RNAs at unprecedented depth suggests their value in
systematically identifying microRNAs (miRNAs). However, the
identification of miRNAs from the large pool of sequenced transcripts
from a single deep sequencing run remains a major challenge. Here, we
present an algorithm, miRDeep, which uses a probabilistic model of
miRNA biogenesis to score compatibility of the position and frequency
of sequenced RNA with the secondary structure of the miRNA precursor.
We demonstrate its accuracy and robustness using published
Caenorhabditis elegans data and data we generated by deep sequencing
human and dog RNAs. miRDeep reports altogether approximately 230
previously unannotated miRNAs, of which four novel C. elegans miRNAs
are validated by northern blot analysis.Friedländer MR, Chen W, Adamidi C, Maaskola J, Einspanier R, Knespel S, Rajewsky N. Nat Biotechnol. 2008 Apr;26(4):407-15. doi: 10.1038/nbt1394. Max Delbrück Centrum für Molekulare Medizin, Robert-Rössle-Strasse 10, D-13125 Berlin-Buch, Germany miRNA profiling for
biomarker discovery in multiple sclerosis: from microarray to deep
sequencing
Guerau-de-Arellano M, Alder H, Ozer HG, Lovett-Racke A, Racke MK. Neurology Department, The Ohio State University Medical Center, 460 W 12th Ave Room 0705, Columbus, OH 43210, USA J Neuroimmunol. 2012 Jul 15;248(1-2): 32-39 MicroRNAs (miRNAs) are
small non-coding RNAs that regulate gene expression at the
post-transcriptional level. miRNAs are highly expressed in cells of the
immune and nervous system, attesting to their importance in
Neuroimmunology. Besides their involvement in modulation of
physiological and pathological processes, miRNAs hold high promise as
disease biomarkers, therapeutic agents and/or drug targets. Several
studies have recently explored the involvement of miRNAs in Multiple
Sclerosis (MS) using a variety of miRNA profiling techniques. In this
review, we discuss basic miRNA biology and nomenclature, the techniques
available for miRNA profiling research and recent miRNA profiling
studies in Multiple Sclerosis.
MicroRNAs accurately
identify cancer tissue origin
Rosenfeld N, Aharonov R, Meiri E, Rosenwald S, Spector Y, Zepeniuk M, Benjamin H, Shabes N, Tabak S, Levy A, Lebanony D, Goren Y, Silberschein E, Targan N, Ben-Ari A, Gilad S, Sion-Vardy N, Tobar A, Feinmesser M, Kharenko O, Nativ O, Nass D, Perelman M, Yosepovich A, Shalmon B, Polak-Charcon S, Fridman E, Avniel A, Bentwich I, Bentwich Z, Cohen D, Chajut A, Barshack I. Rosetta Genomics Ltd., Rehovot 76706, Israel. Nat Biotechnol. 2008 Apr;26(4):462-9. doi: 10.1038/nbt1392 MicroRNAs (miRNAs)
belong to a class of noncoding, regulatory RNAs that is involved in
oncogenesis and shows remarkable tissue specificity. Their potential
for tumor classification suggests they may be used in identifying the
tissue in which cancers of unknown primary origin arose, a major
clinical problem. We measured miRNA expression levels in 400
paraffin-embedded and fresh-frozen samples from 22 different tumor
tissues and metastases. We used miRNA microarray data of 253 samples to
construct a transparent classifier based on 48 miRNAs. Two-thirds of
samples were classified with high confidence, with accuracy >90%. In
an independent blinded test-set of 83 samples, overall high-confidence
accuracy reached 89%. Classification accuracy reached 100% for most
tissue classes, including 131 metastatic samples. We further validated
the utility of the miRNA biomarkers by quantitative RT-PCR using 65
additional blinded test samples. Our findings demonstrate the
effectiveness of miRNAs as biomarkers for tracing the tissue of origin
of cancers of unknown primary origin.
Pre-microRNA and
mature microRNA in human mitochondria
METHODOLOGY/PRINCIPAL FINDINGS: To investigate the
potential origin of mitochondrial miRNA, we in-silico searched for
microRNA candidates in the mtDNA. Twenty five human pre-miRNA and 33
miRNA aligments (E-value<0.1) were found in the reference
mitochondrial sequence and some of the best candidates were chosen for
a co-localization test. In situ hybridization of pre-mir-302a,
pre-let-7b and mir-365, using specific labelled locked nucleic acids
and confocal microscopy, demonstrated that these miRNA were localized
in mitochondria of human myoblasts. Total RNA was extracted from
enriched mitochondria isolated by an immunomagnetic method from a
culture of human myotubes. The detection of 742 human miRNA (miRBase)
were monitored by RT-qPCR at three increasing mtRNA inputs. Forty six
miRNA were significantly expressed (2(nd) derivative method Cp>35)
for the smallest RNA input concentration and 204 miRNA for the maximum
RNA input concentration. In silico analysis predicted 80 putative miRNA
target sites in the mitochondrial genome (E-value<0.05).Barrey E, Saint-Auret G, Bonnamy B, Damas D, Boyer O, Gidrol X. Unité de Biologie Intégrative des Adaptations à l'Exercice-INSERM U902, Genopole Evry, France PLoS One. 2011;6(5):e20220. doi: 10.1371/journal.pone.0020220 BACKGROUND: Because of
the central functions of the mitochondria in providing metabolic energy
and initiating apoptosis on one hand and the role that microRNA (miRNA)
play in gene expression, we hypothesized that some miRNA could be
present in the mitochondria for post-transcriptomic regulation by RNA
interference. We intend to identify miRNA localized in the mitochondria
isolated from human skeletal primary muscular cells.
CONCLUSIONS/SIGNIFICANCE: The present study experimentally demonstrated for the first time the presence of pre-miRNA and miRNA in the human mitochondria isolated from skeletal muscular cells. A set of miRNA were significantly detected in mitochondria fraction. The origin of these pre-miRNA and miRNA should be further investigate to determine if they are imported from the cytosol and/or if they are partially processed in the mitochondria. Toward the
blood-borne miRNome of human diseases
In a multicenter study, we determined the expression
profiles of 863 microRNAs by array analysis of 454 blood samples from
human individuals with different cancers or noncancer diseases, and
validated this 'miRNome' by quantitative real-time PCR. We detected
consistently deregulated profiles for all tested diseases; pathway
analysis confirmed disease association of the respective microRNAs. We
observed significant correlations (P = 0.004) between the genomic
location of disease-associated genetic variants and deregulated
microRNAs.Keller A, Leidinger P, Bauer A, Elsharawy A, Haas J, Backes C, Wendschlag A, Giese N, Tjaden C, Ott K, Werner J, Hackert T, Ruprecht K, Huwer H, Huebers J, Jacobs G, Rosenstiel P, Dommisch H, Schaefer A, Müller-Quernheim J, Wullich B, Keck B, Graf N, Reichrath J, Vogel B, Nebel A, Jager SU, Staehler P, Amarantos I, Boisguerin V, Staehler C, Beier M, Scheffler M, Büchler MW, Wischhusen J, Haeusler SF, Dietl J, Hofmann S, Lenhof HP, Schreiber S, Katus HA, Rottbauer W, Meder B, Hoheisel JD, Franke A, Meese E. Biomarker Discovery Center, Heidelberg, Germany Nat Methods. 2011 Sep 4;8(10): 841-843 Evaluation of two
commercial global miRNA expression profiling platforms for detection of
less abundant miRNAs
BACKGROUND: microRNAs (miRNA) are short, endogenous
transcripts that negatively regulate the expression of specific mRNA
targets. miRNAs are found both in tissues and body fluids such as
plasma. A major perspective for the use of miRNAs in the clinical
setting is as diagnostic plasma markers for neoplasia. While miRNAs are
abundant in tissues, they are often scarce in plasma. For
quantification of miRNA in plasma it is therefore of importance to use
a platform with high sensitivity and linear performance in the low
concentration range. This motivated us to evaluate the performance of
three commonly used commercial miRNA quantification platforms: GeneChip
miRNA 2.0 Array, miRCURY Ready-to-Use PCR, Human panel I+II V1.M, and
TaqMan Human MicroRNA Array v3.0.Jensen SG, Lamy P, Rasmussen MH, Ostenfeld MS, Dyrskjøt L, Orntoft TF, Andersen CL. Department of Molecular Medicine (MOMA), Aarhus University Hospital-Skejby, DK-8200 Aarhus N, Denmark. BMC Genomics. 2011 Aug 26;12: 435 RESULTS: Using synthetic miRNA samples and plasma RNA samples spiked with different ratios of 174 synthetic miRNAs we assessed the performance characteristics reproducibility, recovery, specificity, sensitivity and linearity. It was found that while the qRT-PCR based platforms were sufficiently sensitive to reproducibly detect miRNAs at the abundance levels found in human plasma, the array based platform was not. At high miRNA levels both qRT-PCR based platforms performed well in terms of specificity, reproducibility and recovery. At low miRNA levels, as in plasma, the miRCURY platform showed better sensitivity and linearity than the TaqMan platform. CONCLUSION: For profiling clinical samples with low miRNA abundance, such as plasma samples, the miRCURY platform with its better sensitivity and linearity would probably be superior. An alternative mode
of microRNA target recognition
MicroRNAs (miRNAs) regulate mRNA targets through perfect
pairing with their seed region (positions 2-7). Recently, a precise
genome-wide map of miRNA interaction sites in mouse brain was generated
by high-throughput sequencing and analysis of clusters of
~50-nucleotide mRNA tags cross-linked to 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
now identified an alternative binding mode used by miRNAs.
Specifically, G-bulge sites (positions 5-6) are often bound and
regulated by miR-124 in brain. More generally, bulged sites comprise
≥15% of all Ago-miRNA interactions in mouse brain and are
evolutionarily conserved. We call 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.Chi SW, Hannon GJ, Darnell RB. Laboratory of Neuro-Oncology, The Rockefeller University, Howard Hughes Medical Institute, New York, New York, USA Nat Struct Mol Biol. 2012 19(3): 321-327 Small
non-coding RNAs in animal development
Giovanni Stefani & Frank J. Slack Nature Reviews Molecular Cell Biology 9, 219-230 (2008) The modulation of gene
expression by small non-coding RNAs is a
recently discovered level of gene regulation in animals and plants. In
particular, microRNAs (miRNAs) and Piwi-interacting RNAs (piRNAs) have
been implicated in various aspects of animal development, such as
neuronal, muscle and germline development. During the past year, an
improved understanding of the biological functions of small non-coding
RNAs has been fostered by the analysis of genetic deletions of
individual miRNAs in mammals. These studies show that miRNAs are key
regulators of animal development and are potential human disease loci.
MicroRNAs in
metabolism and metabolic disorders
MicroRNAs (miRNAs) have recently emerged as key regulators
of metabolism. For example, miR-33a and miR-33b have a crucial role in
controlling cholesterol and lipid metabolism in concert with their host
genes, the sterol-regulatory element-binding protein (SREBP)
transcription factors. Other metabolic miRNAs, such as miR-103 and
miR-107, regulate insulin and glucose homeostasis, whereas miRNAs such
as miR-34a are emerging as key regulators of hepatic lipid homeostasis.
The discovery of circulating miRNAs has highlighted their potential as
both endocrine signalling molecules and disease markers. Dysregulation
of miRNAs may contribute to metabolic abnormalities, suggesting that
miRNAs may potentially serve as therapeutic targets for ameliorating
cardiometabolic disorders.Rottiers V, Näär AM. Massachusetts General Hospital Cancer Center, Charlestown, Massachusetts 02129, USA. Nat Rev Mol Cell Biol. 2012 13(4): 239-250 Virus-encoded
microRNAs: an overview and a look to the future
MicroRNAs (miRNAs) are small RNAs that play important
roles in the regulation of gene expression. First described as
posttranscriptional gene regulators in eukaryotic hosts, virus-encoded
miRNAs were later uncovered. It is now apparent that diverse virus
families, most with DNA genomes, but at least some with RNA genomes,
encode miRNAs. While deciphering the functions of viral miRNAs has
lagged behind their discovery, recent functional studies are bringing
into focus these roles. Some of the best characterized viral miRNA
functions include subtle roles in prolonging the longevity of infected
cells, evading the immune response, and regulating the switch to lytic
infection. Notably, all of these functions are particularly important
during persistent infections. Furthermore, an emerging view of viral
miRNAs suggests two distinct groups exist. In the first group, viral
miRNAs mimic host miRNAs and take advantage of conserved networks of
host miRNA target sites. In the larger second group, viral miRNAs do
not share common target sites conserved for host miRNAs, and it remains
unclear what fraction of these targeted transcripts are beneficial to
the virus. Recent insights from multiple virus families have revealed
new ways of interacting with the host miRNA machinery including
noncanonical miRNA biogenesis and new mechanisms of posttranscriptional
cis gene regulation. Exciting challenges await the field, including
determining the most relevant miRNA targets and parlaying our current
understanding of viral miRNAs into new therapeutic strategies. To
accomplish these goals and to better grasp miRNA function, new in vivo
models that recapitulate persistent infections associated with viral
pathogens are required.Kincaid RP, Sullivan CS. The University of Texas at Austin, Molecular Genetics & Microbiology, Austin, Texas, United States of America. PLoS Pathog. 2012 8(12): e1003018 A versatile method
to design stem-loop primer-based quantitative PCR assays for detecting
small regulatory RNA molecules
Short regulatory RNA-s have been identified as key
regulators of gene expression in eukaryotes. They have been involved in
the regulation of both physiological and pathological processes such as
embryonal development, immunoregulation and cancer. One of their
relevant characteristics is their high stability, which makes them
excellent candidates for use as biomarkers. Their number is constantly
increasing as next generation sequencing methods reveal more and more
details of their synthesis. These novel findings aim for new detection
methods for the individual short regulatory RNA-s in order to be able
to confirm the primary data and characterize newly identified subtypes
in different biological conditions. We have developed a flexible method
to design RT-qPCR assays that are very sensitive and robust. The newly
designed assays were tested extensively in samples from plant, mouse
and even human formalin fixed paraffin embedded tissues. Moreover, we
have shown that these assays are able to quantify endogenously
generated shRNA molecules. The assay design method is freely available
for anyone who wishes to use a robust and flexible system for the
quantitative analysis of matured regulatory RNAs.Czimmerer Z, Hulvely J, Simandi Z, Varallyay E, Havelda Z, Szabo E, Varga A, Dezso B, Balogh M, Horvath A, Domokos B, Torok Z, Nagy L, Balint BL. Department of Biochemistry and Molecular Biology, Research Center for Molecular Medicine, University of Debrecen Medical and Health Science Center, Debrecen, Hungary. PLoS One. 2013;8(1): e55168 |