For the past decade, researchers have been focusing extensively on miRNA profiles in various plants. Here, for the first time in a non-model plant—Abelmoschus esculentus with negligible genome information—we are reporting the global profiling to characterize the miRNAs and their associated pre-miRNAs by applying a next generation sequencing approach. Preliminarily, we performed small RNA sRNA sequencing with five biological replicates of leaf samples to attain , , reads; data analysis using miRPlant revealed known and novel miRNA candidates. With the objective of seizing their associated hairpin precursors, we accomplished pre-miRNA sequencing to attain 83, , reads. The paired end reads are merged and adaptor trimmed, and the resulting 40— nt nucleotide sequences were picked out for analysis by using perl scripts from the miRGrep tool and an in-house built shell script for Minimum Fold Energy Index MFEI calculation. Applying the stringent criteria of the Dicer cleavage pattern and the perfect stem loop structure, precursors for 57 known miRNAs of 15 families and 18 novel miRNAs were revealed. The elimination of the pre-miRNA loop is subsequently followed by the continuous stretch of two- or multi-step cleavage at nt intervals alongside the stem, resulting in the formation miRNA: The precise miRNA recognition from the pri-miRNA relies on the precursor composition comprising processing signals such as 14— bp nt paired in stem proximal; a specific length of nt paired in stem distal to the miRNA: In addition, an alternative biogenesis pathway has been proposed: The mature miRNA, in association with AGO, silences the target transcript either by direct cleavage slicing or by destabilization through slicer-independent turnover mechanisms and translational repression [ 9 ].

Small RNA sorting: matchmaking for Argonautes

Studies also suggest that miRNAs are important in host—virus interactions where the host limits virus infection by differentially expressing miRNAs that target essential viral genes. Here, we identified conserved and new miRNAs from Spodoptera frugiperda cells Sf9 using a combination of deep sequencing and bioinformatics as well as experimental approaches. The predominant miRNAs were found to be conserved among arthropods.

hAgo2 has been shown to be much less susceptible to protease treatment when bound to guide RNA than the unbound protein, and eukaryotic Argonautes retain small RNAs when expressed recombinantly in Escherichia coli or in insect cells 38, Strikingly, guide RNA interacts with all domains of Argonaute including its linkers (Figure 1A).

A family of microRNAs present in plants and animals. An endogenous, systemic RNAi pathway in plants. An RNA-dependent RNA polymerase prevents meristem invasion by potato virus X and is required for the activity but not the production of a systemic silencing signal. Arabidopsis thaliana DNA methylation mutants. Assaying Chimeric Genes in Plants: Comparative transcriptomics of Arabidopsis sperm cells. Criteria for annotation of plant MicroRNAs.

Cytoplasmic connection of sperm cells to the pollen vegetative cell nucleus:

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Organisms employ a variety of strategies to defend themselves by distinguishing self and nonself substances and disabling these invasive nucleic acids. Furthermore, they have developed ways to remember this exposure to invaders and transmit the experience to their descendants. The mechanism underlying this inheritance has remained elusive. Recent research has shed light on the initiation and maintenance of RNA-mediated inherited gene silencing.

Small regulatory RNAs play a variety of crucial roles in organisms, including gene regulation, developmental timing, antiviral defense, and genome integrity, via a process termed as RNA interference RNAi. Recent research has revealed that small RNAs and the RNAi machinery are engaged in establishing and promoting transgenerational gene silencing.

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Your dirty mouth may be partly to blame Enlarge credit: Media for Medical Chronic intestinal disturbances may in part be handed down from above, according to a study published Thursday in Science. Intestinal pathogens can lurk in the mouth and—at just the right moments— interlope in the gut to help trigger severe, recurring bouts of inflammation , researchers found. AP Senate lawmakers on Thursday unveiled their first major legislative effort to increase transparency in online political advertising, called the Honest Ads Act.

Assembly and function of small RNA Argonaute protein complexes

This expansion generates a mutant huntingtin protein that contains an abnormally elongated polyglutamine tract, which, together with mutant transcript, causes cellular dysfunction. Currently, there is no curative treatment available to patients suffering from HD; however, the selective inhibition of the mutant allele expression is a promising therapeutic option. In this study, we developed a new class of CAG repeat-targeting silencing reagents that consist of self-duplexing CUG repeats.

Self-duplex formation was induced through one or several U-base substitutions. A number of self-duplexing guide-strand-only short interfering RNAs have been tested through transfection into cells derived from HD patients, showing distinct activity profiles. The best reagents were highly discriminatory between the normal and mutant HTT alleles allele selectivity and the HTT transcript and other transcripts containing shorter CAG repeats gene selectivity.

Small RNAs directly or indirectly impact nearly every biological process in eukaryotic cells. To perform their myriad roles, not only must precise small RNA species be generated, but they must also be loaded into specific effector complexes called RNA-induced silencing complexes (RISCs).

Advanced Search Abstract Motivation: Recent progress in high-throughput sequencing technologies has largely contributed to reveal a highly complex landscape of small non-coding RNAs sRNAs , including novel non-canonical sRNAs derived from long non-coding RNA, repeated elements, transcription start sites and splicing site regions among others. The published frameworks for sRNA data analysis are focused on miRNA detection and prediction, ignoring further information in the dataset.

As a consequence, tools for the identification and classification of the sRNAs not belonging to miRNA family are currently lacking. This new module deals with sequences mapping onto multiple locations and permits a highly versatile and user-friendly interaction with the data in order to easily classify sRNA sequences with a putative functional importance.

Supplementary data are available at Bioinformatics online. Recent progress in high-throughput sequencing technologies has largely contributed to elucidate the remarkable landscape of sRNAs, revealing new species of sRNAs with unknown functions. These novel sRNAs are classified according to their position in the genome and putative functions. Nowadays, five major groups of sRNAs have been detected:

CiteULike: Tag small_rna [83 articles]

Recent studies of lncRNAs across different species have revealed a diverse population of RNA molecules of differing size and function. RNA sequencing studies suggest transcription throughout the genome, so there is a need to understand how sequence relates to functional and structural relationships amongst RNA molecules. Our synthesis of recent studies suggests that neither size, presence of a poly-A tail, splicing, direction of transcription, nor strand specificity are of importance to lncRNA function.

Rather, relative genomic position in relation to a target is fundamentally important. In this review, we describe issues of key importance in functional assessment of lncRNA and how this might apply to lncRNAs important in neurodevelopment.

A specific small interfering RNA sequence targeting the CRMP‑4 gene (DPYSL3) was constructed and transfected into an SW cell line using a lentivirus vector .

Download as PowerPoint Slide Figure 7. Requirement for exogenous Ago2 in pre-miR processing. A Schematic representation of the evolutionarily conserved short hairpin structure of vertebrate pre-miR , indicating the mature miRNA bold line and the initial Ago2 cleavage site at nucleotide B 32P-labeled pre-miR RNA was injected into oocytes left or one- to two-cell embryos right that were untreated control or programmed to express the indicated hAgo2 proteins, and pre-miR processing was monitored by denaturing PAGE.

The identity of the processing products was confirmed by RNase T1 fingerprinting not shown. S7B,C; data not shown. In this regard, we note that expression of exogenous, catalytically active Ago2 did not perturb embryonic development Supplemental Fig. S3; data not shown. Previous Section Next Section Discussion We found that, during Xenopus oocyte maturation and early embryogenesis, expression of Ago proteins, unlike Exp5 and Dicer, is developmentally controlled, with the levels of Ago proteins being low in both immature oocytes and cleavage stage embryos, but rising after MBT.

Eukaryotic Argonautes come into focus

China Published online on: This is an open access article distributed under the terms of Creative Commons Attribution License. Introduction Colorectal cancer CRC is the third most common malignant tumor in the world, and has a relatively poor prognosis 1. Although chemoprevention or chemotherapy drugs may be a promising approach to reduce the incidence and improve the prognosis of CRC, the clinical application is greatly limited by the development of chemotherapy resistance and the toxic side effects 4 , 5.

The development of CRC involves various genetic and molecular changes in cell proliferation, survival and differentiation, resistance to apoptosis, metastasis and tumor angiogenesis 6. Understanding the etiology of CRC is critical for the treatment of the disease.

Small RNA sorting: matchmaking for Argonautes. Nat. Rev. Genet. ; Crossref; PubMed; Scopus () Google Scholar]. Originally thought to derive exclusively from exogenous sources, siRNAs from endogenous intermolecular and intramolecular mRNA duplexes are now documented, revealing another source of RNA fragments.

The authors have declared that no competing interests exist. Conceived and designed the experiments: Received Nov 12; Accepted Jan Abstract Traditionally, researchers match a microRNA guide strand to mRNA sequences using sequence comparisons to predict its potential target genes. However, many of the predictions can be false positives due to limitations in sequence comparison alone.

In this work, we consider the association of two related RNA structures that share a common guide strand: We have analyzed thousands of such structure pairs and found many of them share high structural similarity. Therefore, we conclude that when predicting microRNA target genes, considering just the microRNA guide strand matches to gene sequences may not be sufficient — The microRNA duplex structure formed by the guide strand and its companion passenger strand must also be considered.

Drosha as an interferon

Received Nov 6; Accepted Feb Abstract Argonaute proteins Ago1—4 are essential components of the microRNA-induced silencing complex and play important roles in both microRNA biogenesis and function. Although Ago2 is the only one with the slicer activity, it is not clear whether the slicer activity is a universally critical determinant for Ago2’s function in mammals.

Distinct AGO1 and AGO2 associated miRNA profiles in human cells and blood plasma. Small RNA sorting: matchmaking for Argonautes. Distinct AGO1 and AGO2 associated miRNA profiles in human cells and blood plasma. All authors. Andrey Turchinovich & Barbara Burwinkel.

This would lead to the fluctuation of global gene expression not only by repression of exo-miRNA target gene expression, but also by the increase of the endo-miRNA target gene expression. In the present study, we quantified the changes in the expression levels of target genes of exo-miRNA and endo-miRNA in the cells transfected with fifteen different exo-miRNAs by microarray experiments. Different exo-miRNAs increased ratios of expression levels of target genes of a given endo-miRNA to different extents, suggesting that the replacement efficiencies might differ according to the exo-miRNA types.

These interfering effects are shown to be caused by competition for RNA silencing components. Evolutionary pressure has selected one particular strand of the duplex as the main regulator, which is preferentially loaded onto RISC, with the opposite strand being less functional [ 22 , 23 ]. The major determinants of RISC loading have been shown to be the thermodynamic properties: In this study, to investigate the mechanism to perturb the endo-miRNA function caused by the introduction of exo-miRNA, we performed microarray profiling to quantify changes in the expression levels of endo-miRNA target genes following the transfection of fifteen different synthetic exo-miRNA duplexes.

Such competition among cellular miRNAs is considered to have a role in normal biological and disease-related cellular processes. The exo-miRNAs, in addition to silencing their own target genes, clearly increased the expression of endo-miRNA target genes. The increased levels of endo-miRNA target gene expression were varied according to the types of the transfected exo-miRNA duplexes, suggesting each exo-miRNA duplex is presumed to have distinct characteristics, such as structures and sequences, which affect its incorporation into the RISC.

However, the increased levels were mostly equivalent according to the types of endo-miRNAs after transfection of any particular exo-miRNA duplex.


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