Hyeshik Chang

Ever growing interest in microRNAs has immensely populated the number of resources and research papers devoted to the field and, as a result, it becomes more and more demanding to find miRNA data of interest. To mitigate this problem, we created miRNEST database (http://mirnest.amu.edu.pl), an integrative microRNAs resource. In its updated version, named miRNEST 2.0, the database is complemented with our extensive miRNA predictions from deep sequencing libraries, data from plant degradome analyses, results of pre-miRNA classification with HuntMi and miRNA splice sites information. We also added download and upload options and improved the user interface to make it easier to browse through miRNA records.

The removal of the 5' cap structure by the decapping enzyme DCP2 inhibits translation and generally commits the mRNA to irreversible 5'-to-3' exonucleolytic degradation by XRN1. DCP2 catalytic activity is stimulated by DCP1, and these proteins form the conserved core of the decapping complex. Additional decapping factors orchestrate the recruitment and activity of this complex in vivo. These factors include enhancer of decapping 3 (EDC3), EDC4, like Sm14A (LSm14A), Pat, the LSm1–7 complex, and the RNA helicase DDX6. Decapping factors are often modular and feature folded domains flanked or connected by low-complexity disordered regions. Recent studies have made important advances in understanding how these disordered regions contribute to the assembly of decapping complexes and promote phase transitions that drive RNP granule formation. These studies have also revealed that the decapping network is governed by interactions mediated by short linear motifs (SLiMs) in these disordered regions. Consequently, the network has rapidly evolved, and although decapping factors are conserved, individual interactions between orthologs have been rewired during evolution. The plasticity of the network facilitates the acquisition of additional subunits or domains in pre-existing subunits, enhances opportunities for regulating mRNA degradation, and eventually leads to the emergence of novel functions.

by Igor Ruiz de los Mozos, Marta Vergara-Irigaray, Victor Segura, Maite Villanueva, Nerea Bitarte, Margarida Saramago, Susana Domingues, Cecilia M. Arraiano, Pierre Fechter, Pascale Romby, Jaione Valle, Cristina Solano, Iñigo Lasa, Alejandro Toledo-Arana

The presence of regulatory sequences in the 3′ untranslated region (3′-UTR) of eukaryotic mRNAs controlling RNA stability and translation efficiency is widely recognized. In contrast, the relevance of 3′-UTRs in bacterial mRNA functionality has been disregarded. Here, we report evidences showing that around one-third of the mapped mRNAs of the major human pathogen Staphylococcus aureus carry 3′-UTRs longer than 100-nt and thus, potential regulatory functions. We selected the long 3′-UTR of icaR, which codes for the repressor of the main exopolysaccharidic compound of the S. aureus biofilm matrix, to evaluate the role that 3′-UTRs may play in controlling mRNA expression. We showed that base pairing between the 3′-UTR and the Shine-Dalgarno (SD) region of icaR mRNA interferes with the translation initiation complex and generates a double-stranded substrate for RNase III. Deletion or substitution of the motif (UCCCCUG) within icaR 3′-UTR was sufficient to abolish this interaction and resulted in the accumulation of IcaR repressor and inhibition of biofilm development. Our findings provide a singular example of a new potential post-transcriptional regulatory mechanism to modulate bacterial gene expression through the interaction of a 3′-UTR with the 5′-UTR of the same mRNA.

Background: RNA-seq is now widely used to quantitatively assess gene expression, expression differences and isoform switching, and promises to deliver results for the entire transcriptome. However, whether the transcriptional state of a gene can be captured accurately depends critically on library preparation, read alignment, expression estimation and the tests for differential expression and isoform switching. There are comparisons available for the individual steps but there is not yet a systematic investigation which specific genes are impacted by biases throughout the entire analysis workflow. It is especially unclear whether for a given gene, with current methods and protocols, expression changes and isoform switches can be detected. Results: For the human genes, we report their detectability under various conditions using different approaches. Overall, we find that the input material has the biggest influence and may, depending on the protocol and RNA degradation, exhibit already strong length-dependent over- and underrepresentation of transcripts. The alignment step aligns for 50% of the isoforms up to 99% of the reads correctly; only in the presence of transcript modifications mainly short isoforms will have a low alignment rate. In our dataset, we found that, depending on the aligner and the input material used, the expression estimation of up to 93% of the genes being accurate within a factor of two; with the deviations being due to ambiguous alignments. Detection of differential expression using a negative-binomial count model works reliably for our simulated data but is dependent on the count accuracy. Interestingly, using the fold-change instead of the p-value as a score for differential expression yields the same performance in the situation of three replicates and the true change being two-fold. Isoform switching is harder to detect and for at least 109 genes the isoform differences evade detection independent of the method used. Conclusions: RNA-seq is a reliable tool but the repetitive nature of the human genome makes the origin of the reads ambiguous and limits the detectability for certain genes. RNA-seq does not equally well represent isoforms independent of their size which may range from ~200nt to ~100′000nt. Researchers are advised to verify that their target genes do not have extreme properties with respect to repeated regions, GC content, and isoform length and complexity.

Nature Reviews Genetics. doi:10.1038/nrg3661

Author: Hannah Stower

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Drosha Regulates Gene Expression Independently of RNA Cleavage Function.

Cell Rep. 2013 Dec 18;

Authors: Gromak N, Dienstbier M, Macias S, Plass M, Eyras E, Cáceres JF, Proudfoot NJ

Abstract
Drosha is the main RNase III-like enzyme involved in the process of microRNA (miRNA) biogenesis in the nucleus. Using whole-genome ChIP-on-chip analysis, we demonstrate that, in addition to miRNA sequences, Drosha specifically binds promoter-proximal regions of many human genes in a transcription-dependent manner. This binding is not associated with miRNA production or RNA cleavage. Drosha knockdown in HeLa cells downregulated nascent gene transcription, resulting in a reduction of polyadenylated mRNA produced from these gene regions. Furthermore, we show that this function of Drosha is dependent on its N-terminal protein-interaction domain, which associates with the RNA-binding protein CBP80 and RNA Polymerase II. Consequently, we uncover a previously unsuspected RNA cleavage-independent function of Drosha in the regulation of human gene expression.

PMID: 24360955 [PubMed - as supplied by publisher]

In eukaryotes, transfer RNAs (tRNAs) are transcribed in the nucleus yet function in the cytoplasm; thus, tRNA movement within the cell was believed to be unidirectional—from the nucleus to the cytoplasm. It is now known that mature tRNAs also move in a retrograde direction from the cytoplasm to the nucleus...

By using chromosome conformation capture technology, a recent study has revealed two alternative three-dimensional folding states of the human genome during the cell cycle.

Genome-wide assessment of post-transcriptional control in the fly brain.

Front Mol Neurosci. 2013;6:49

Authors: Mezan S, Ashwal-Fluss R, Shenhav R, Garber M, Kadener S

Abstract
Post-transcriptional control of gene expression has central importance during development and adulthood and in physiology in general. However, little is known about the extent of post-transcriptional control of gene expression in the brain. Most post-transcriptional regulatory effectors (e.g., miRNAs) destabilize target mRNAs by shortening their polyA tails. Hence, the fraction of a given mRNA that it is fully polyadenylated should correlate with its stability and serves as a good measure of post-transcriptional control. Here, we compared RNA-seq datasets from fly brains that were generated either from total (rRNA-depleted) or polyA-selected RNA. By doing this comparison we were able to compute a coefficient that measures the extent of post-transcriptional control for each brain-expressed mRNA. In agreement with current knowledge, we found that mRNAs encoding ribosomal proteins, metabolic enzymes, and housekeeping genes are among the transcripts with least post-transcriptional control, whereas mRNAs that are known to be highly unstable, like circadian mRNAs and mRNAs expressing synaptic proteins and proteins with neuronal functions, are under strong post-transcriptional control. Surprisingly, the latter group included many specific groups of genes relevant to brain function and behavior. In order to determine the importance of miRNAs in this regulation, we profiled miRNAs from fly brains using oligonucleotide microarrays. Surprisingly, we did not find a strong correlation between the expression levels of miRNAs in the brain and the stability of their target mRNAs; however, genes identified as highly regulated post-transcriptionally were strongly enriched for miRNA targets. This demonstrates a central role of miRNAs for modulating the levels and turnover of brain-specific mRNAs in the fly.

PMID: 24367289 [PubMed]

Molecular biology: Microbes meddle with microRNA

Nature 504, 7480 (2013). doi:10.1038/504334d

Microbes in mouse guts can interfere with infection responses by changing the expression of small RNAs that regulate some genes.Cristel Archambaud and Pascale Cossart at the Pasteur Institute in Paris and their colleagues looked at the expression of microRNA molecules in the small intestines

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Heon-Ki Kim, Ryu-Ryun Kim, Jang-Hyun Oh, Hanna Cho, Alexander Varshavsky, Cheol-Sang Hwang. The Arg/N-end rule pathway targets for degradation proteins that bear specific unacetylated N-terminal residues while the Ac/N-end rule pathway targets proteins through their N^α-termina....

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Article

Cells secrete micro-RNAs by packaging them into exosomes; however, the mechanisms by which this packaging occurs are unclear. Here, the authors identify a sequence motif that confers exosomal targeting to micro-RNAs and identify a ribonucleoprotein complex that plays a role in this process.

Nature Communications doi: 10.1038/ncomms3980

Authors: Carolina Villarroya-Beltri, Cristina Gutiérrez-Vázquez, Fátima Sánchez-Cabo, Daniel Pérez-Hernández, Jesús Vázquez, Noa Martin-Cofreces, Dannys Jorge Martinez-Herrera, Alberto Pascual-Montano, María Mittelbrunn, Francisco Sánchez-Madrid

Nature advance online publication 22 December 2013. doi:10.1038/nature12890

Authors: Basil J. Greber, Daniel Boehringer, Alexander Leitner, Philipp Bieri, Felix Voigts-Hoffmann, Jan P. Erzberger, Marc Leibundgut, Ruedi Aebersold & Nenad Ban

Nature Reviews Cancer 14, 70 (2014). doi:10.1038/nrc3477-c1

Authors: Jae Il Shin & Guy G. Brusselle

In a recent article (Mechanistic links between COPD and lung cancer. Nature Rev. Cancer13, 233–245 (2013)), McGarry Houghton reviewed the association between the presence of chronic obstructive pulmonary disease (COPD) and the development of lung cancer

Nature Reviews Cancer 14, 9 (2014). doi:10.1038/nrc3665

Author: Sarah Seton-Rogers

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Nature Reviews Molecular Cell Biology 15, 1 (2014). doi:10.1038/nrm3718

Author: Andrea Du Toit

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Nature Reviews Molecular Cell Biology 15, 4 (2014). doi:10.1038/nrm3715

Author: Katharine H. Wrighton

Lin28a enhances tissue repair by increasing oxidative metabolism.

Nature Reviews Molecular Cell Biology 15, 4 (2014). doi:10.1038/nrm3732

Author: Andrea du Toit

Typically, translation terminates when ribososmes encounter a stop codon. Stop codon readthrough results in carboxy-terminally extended nascent peptides, but its biological roles in eukaryotes have remained elusive. Here, Dunn et al. use a modified ribosome profiling assay to analyse genome-wide translation in Drosophila melanogaster

High-resolution Xist binding maps reveal two-step spreading during X-chromosome inactivation

Nature 504, 7480 (2013). doi:10.1038/nature12719

Authors: Matthew D. Simon, Stefan F. Pinter, Rui Fang, Kavitha Sarma, Michael Rutenberg-Schoenberg, Sarah K. Bowman, Barry A. Kesner, Verena K. Maier, Robert E. Kingston & Jeannie T. Lee

The Xist long noncoding RNA (lncRNA) is essential for X-chromosome inactivation (XCI), the process by which mammals compensate for unequal numbers of sex chromosomes. During XCI, Xist coats the future inactive X chromosome (Xi) and recruits Polycomb repressive complex 2 (PRC2) to the X-inactivation centre (Xic). How Xist spreads silencing on a 150-megabases scale is unclear. Here we generate high-resolution maps of Xist binding on the X chromosome across a developmental time course using CHART-seq. In female cells undergoing XCI de novo, Xist follows a two-step mechanism, initially targeting gene-rich islands before spreading to intervening gene-poor domains. Xist is depleted from genes that escape XCI but may concentrate near escapee boundaries. Xist binding is linearly proportional to PRC2 density and H3 lysine 27 trimethylation (H3K27me3), indicating co-migration of Xist and PRC2. Interestingly, when Xist is acutely stripped off from the Xi in post-XCI cells, Xist recovers quickly within both gene-rich and gene-poor domains on a timescale of hours instead of days, indicating a previously primed Xi chromatin state. We conclude that Xist spreading takes distinct stage-specific forms. During initial establishment, Xist follows a two-step mechanism, but during maintenance, Xist spreads rapidly to both gene-rich and gene-poor regions.