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https://www.readbyqxmd.com/read/28724535/priming-in-a-permissive-type-i-c-crispr-cas-system-reveals-distinct-dynamics-of-spacer-acquisition-and-loss
#1
Chitong Rao, Denny Chin, Alexander Wilson Ensminger
CRISPR-Cas is a bacterial and archaeal adaptive immune system that uses short, invader-derived sequences termed spacers to target invasive nucleic acids. Upon recognition of previously encountered invaders, the system can stimulate secondary spacer acquisitions, a process known as primed adaptation. Previous studies of primed adaptation have been complicated by intrinsically high interference efficiency of most systems against bona fide targets. As such, most primed adaptation to date has been studied within the context of imperfect sequence complementarity between spacers and targets...
July 19, 2017: RNA
https://www.readbyqxmd.com/read/28724534/m6aviewer-software-for-the-detection-analysis-and-visualization-of-n6-methyl-adenosine-peaks-from-m6a-seq-me-rip-sequencing-data
#2
Agne Antanaviciute, Belinda Baquero-Perez, Christopher M Watson, Sally M Harrison, Carolina Lascelles, Laura Crinnion, Alexander F Markham, David T Bonthron, Adrian Whitehouse, Ian M Carr
Recent methods for transcriptome-wide N6-methyl-adenosine(m6A) profiling have facilitated investigations into the RNA methylome and established m6A as a dynamic modification that has critical regulatory roles in gene expression and may play a role in human disease. However, bioinformatics resources available for the analysis of m6A sequencing data are still limited. Here, we describe m6aViewer - a cross-platform application for analysis and visualisation of m6A peaks from sequencing data. m6aViewer implements a novel m6A peak-calling algorithm that identifies high-confidence methylated residues with more precision than previously described approaches...
July 19, 2017: RNA
https://www.readbyqxmd.com/read/28701522/yeast-non-coding-rna-interaction-network
#3
Simona Panni, Ananth Prakash, Alex Bateman, Sandra Orchard
This article describes the creation of the first expert manually curated non-coding RNA interaction networks for S. cerevisiae. The RNA-RNA and RNA-protein interaction networks have been carefully extracted from the experimental literature and made available through the IntAct database (www.ebi.ac.uk/intact). We provide an initial network analysis and compare their properties to the much larger protein-protein interaction network. We find that the proteins that bind to ncRNAs in the network contain only a small proportion of classical RNA binding domains...
July 12, 2017: RNA
https://www.readbyqxmd.com/read/28701521/translational-repression-of-the-drosophila-nanos-mrna-involves-the-rna-helicase-belle-and-rna-coating-by-me31b-and-trailer-hitch
#4
Michael Götze, Jérémy Dufourt, Christian Ihling, Christiane Rammelt, Stéphanie Pierson, Nagraj Sambrani, Claudia Temme, Andrea Sinz, Martine Simonelig, Elmar Wahle
Translational repression of maternal mRNAs is an essential regulatory mechanism during early embryonic development. Repression of the Drosophila nanos mRNA, required for the formation of the anterior-posterior body axis, depends on the protein Smaug binding to two Smaug recognition elements (SREs) in the nanos 3' UTR. In a comprehensive mass-spectrometric analysis of the SRE-dependent repressor complex, we identified Smaug, Cup, Me31B, Trailer hitch, eIF4E and PABPC, in agreement with earlier data. As a novel component, the RNA-dependent ATPase Belle (DDX3) was found, and its involvement in deadenylation and repression of nanos was confirmed in vivo...
July 12, 2017: RNA
https://www.readbyqxmd.com/read/28701520/ligand-recognition-and-helical-stacking-formation-are-intimately-linked-in-the-sam-i-riboswitch-regulatory-mechanism
#5
Anne-Marie Dussault, Audrey Dubé, Frédéric Jacques, Jonathan P Grondin, Daniel Lafontaine
Riboswitches are non-coding mRNA elements that control gene expression by altering their structure upon metabolite binding. Although riboswitch crystal structures provide detailed information about RNA-ligand interactions, little knowledge has been gathered to understand how riboswitches modulate gene expression. Here, we study the molecular recognition mechanism of the S-adenosylmethionine SAM-I riboswitch by characterizing the formation of a helical stacking interaction involving the ligand binding process...
July 12, 2017: RNA
https://www.readbyqxmd.com/read/28701519/nineteen-complex-related-factor-prp45-is-required-for-the-early-stages-of-cotranscriptional-spliceosome-assembly
#6
Martina Hálová, Ondřej Gahura, Martin Převorovský, Zdeněk Cit, Marian Novotný, Anna Valentová, Kateřina Abrhámová, František Půta, Petr Folk
Splicing in S. cerevisiae has been shown to proceed cotranscriptionally, but the nature of the coupling remains a subject of debate. Here, we examine the effect of nineteen complex-related splicing factor Prp45 (a homolog of SNW1/SKIP) on cotranscriptional splicing. RNA-sequencing and RT-qPCR showed elevated pre-mRNA levels but only limited reduction of spliced mRNAs in cells expressing C-terminally truncated Prp45, Prp45(1-169). Assays with a series of reporters containing the AMA1 intron with regulatable splicing confirmed decreased splicing efficiency and showed the leakage of unspliced RNAs in prp45(1-169) cells...
July 12, 2017: RNA
https://www.readbyqxmd.com/read/28698239/enzymatic-production-of-single-molecule-fish-and-rna-capture-probes
#7
Imre Gaspar, Frank Wippich, Anne Ephrussi
Arrays of singly-labelled short oligonucleotides that hybridize to a specific target revolutionized RNA biology, enabling quantitative, single molecule microscopy analysis and high efficiency RNA/RNP capture. Here, we describe a simple and efficient method that allows flexible functionalization of inexpensive DNA oligonucleotides by different fluorescent dyes or biotin using terminal deoxynucleotidyl transferase and custom-made functional group conjugated dideoxy-UTP. We show that 1) all steps of the oligonucleotide labelling - including conjugation, enzymatic synthesis and product purification - can be performed in a standard biology laboratory, 2) the process yields >90 %, often >95 % labeled product with minimal carry-over of impurities and 3) the oligonucleotides can be labeled with different dyes or biotin, allowing single molecule FISH or RNA affinity purification, and northern-blot analysis to be performed...
July 11, 2017: RNA
https://www.readbyqxmd.com/read/28694328/the-contribution-of-the-c5-protein-subunit-of-escherichia-coli-ribonuclease-p-to-specificity-for-precursor-trna-is-modulated-by-proximal-5-leader-sequences
#8
Courtney Nicole Niland, David R Anderson, Eckhard Jankowsky, Michael E Harris
Molecular recognition of RNA by RNA processing enzymes and RNA binding proteins often involves the cooperation between multiple subunits. However, the interdependent contributions of RNA and protein subunits to molecular recognition by ribonucleoproteins and protein-protein complexes is relatively unexplored. RNase P is an endonuclease that removes 5' leaders from precursor tRNAs and functions in bacteria as dimer formed by a catalytic RNA subunit (P RNA) and a protein subunit (C5 in E. coli). The P RNA subunit contacts the tRNA body and proximal 5' leader sequences (N(-1) and N(-2)) while C5 binds distal 5' leader sequences (N(-3) to N(-6))...
July 10, 2017: RNA
https://www.readbyqxmd.com/read/28630141/temperature-responsive-mirnas-in-drosophila-orchestrate-adaptation-to-different-ambient-temperatures
#9
Isabel Fast, Charlotte Hewel, Laura Wester, Julia Schumacher, Daniel Gebert, Hans Zischler, Christian Berger, David Rosenkranz
The majority of Drosophila genes are expressed in a temperature-dependent manner, but the way in which small RNAs may contribute to this effect is completely unknown as we currently lack an idea of how small RNA transcriptomes change as a function of temperature. Applying high throughput sequencing techniques complemented by quantitative real-time PCR experiments, we demonstrate that altered ambient temperature induces drastic, but reversible changes in sequence composition and total abundance of both, miRNA- and piRNA populations...
June 19, 2017: RNA
https://www.readbyqxmd.com/read/28630140/three-dimensional-structure-of-the-3-x-tail-of-hepatitis-c-virus-rna-in-monomeric-and-dimeric-states
#10
Angel Cantero-Camacho, Lixin Fan, Yun-Xing Wang, José Gallego
The 3'X domain is a 98-nucleotide region located at the 3' end of hepatitis C virus genomic RNA that plays essential functions in the viral life cycle. It contains an absolutely conserved, 16-base palindromic sequence that promotes viral RNA dimerization, overlapped with a 7-nucleotide tract implicated in a distal contact with a nearby functional sequence. Using small angle X-ray scattering measurements combined with model building guided by NMR spectroscopy, we have studied the stoichiometry, structure and flexibility of domain 3'X and two smaller subdomain sequences as a function of ionic strength, and obtained a three-dimensional view of the full-length domain in its monomeric and dimeric states...
June 19, 2017: RNA
https://www.readbyqxmd.com/read/28625967/rnase-ii-regulates-rnase-ph-and-is-essential-for-cell-survival-during-starvation-and-stationary-phase
#11
Shaheen Sulthana, Ernesto Quesada, Murray P Deutscher
RNase II is the most active exoribonuclease in Escherichia coli cell extracts. Yet, its removal appears to have no deleterious effect on growing cells. Here, we show that RNase II is required for cell survival during prolonged stationary phase and upon starvation. The absence of RNase II leads to greatly increased rRNA degradation and to the accumulation of rRNA fragments, both of which lead to a decline in cell survival. The deleterious effects of RNase II removal can be completely reversed by the simultaneous absence of a second exoribonuclease, RNase PH, an enzyme known to be required to initiate ribosome degradation in starving cells...
June 16, 2017: RNA
https://www.readbyqxmd.com/read/28611253/n6-methyladenosine-is-required-for-the-hypoxic-stabilization-of-specific-mrnas
#12
Nate J Fry, Brittany A Law, Olga R Ilkayeva, Christopher L Holley, Kyle D Mansfield
Post-transcriptional regulation of mRNA during oxygen deprivation, or hypoxia, can affect the survivability of cells. Hypoxia has been shown to increase stability of a subset of ischemia-related mRNAs, including VEGF. RNA binding proteins and miRNAs have been identified as important for post-transcriptional regulation of individual mRNAs, but corresponding mechanisms that regulate global stability are not well understood. Recently, mRNA modification by N6-methyladenosine (m6A) has been shown to be involved in post-transcriptional regulation processes including mRNA stability and promotion of translation, but the role of m6A in the hypoxia response is unknown...
June 13, 2017: RNA
https://www.readbyqxmd.com/read/28606943/role-of-terminator-hairpin-in-biogenesis-of-functional-hfq-binding-srnas
#13
Teppei Morita, Ryo Nishino, Hiroji Aiba
Many Rho-independent transcription terminators of the genes encoding bacterial Hfq-binding sRNAs possess a set of 7 or more T residues at the 3' end (Otaka et al. 2011; Ishikawa et al. 2012). Here, we have studied the role of terminator hairpin in biogenesis of sRNAs focusing on SgrS and RyhB in Esherichia coli. We constructed variant sRNA genes in which the GC-rich inverted repeat sequences are extended to stabilize the terminator hairpins. We demonstrate that the extension of the hairpin stem leads to generation of heterogeneous transcripts in which the polyU tail is shortened...
June 12, 2017: RNA
https://www.readbyqxmd.com/read/28600356/structural-basis-for-ligand-binding-to-the-guanidine-ii-riboswitch
#14
Caroline W Reiss, Scott A Strobel
The guanidine-II riboswitch, also known as mini-ykkC, is a conserved mRNA element with more than 800 examples in bacteria. It consists of two stemloops capped by identical, conserved tetraloops that are separated by a linker region of variable length and sequence. Like the guanidine-I riboswitch, it controls the expression of guanidine carboxylases and SugE-like genes. The guanidine-II riboswitch specifically binds free guanidinium cation and functions as a translationally-controlled on-switch. Here we report the structure of a P2 stemloop from the Pseudomonas aeruginosa guanidine-II riboswitch aptamer bound to guanidine at 1...
June 9, 2017: RNA
https://www.readbyqxmd.com/read/28592461/rbm24-promotes-u1-snrnp-recognition-of-the-mutated-5-splice-site-in-the-ikbkap-gene-of-familial-dysautonomia
#15
Kenji Ohe, Mayumi Yoshida, Akiko Nakano-Kobayashi, Motoyasu Hosokawa, Yukiya Sako, Maki Sakuma, Yukiko Okuno, Tomomi Usui, Kensuke Ninomiya, Takayuki Nojima, Naoyuki Kataoka, Masatoshi Hagiwara
The 5' splice site mutation (IVS20+6T>C) of the inhibitor of kappa light polypeptide gene enhancer in B cells, kinase complex-associated protein (IKBKAP) gene in familial dysautonomia (FD) is at the sixth intronic nucleotide of the 5' splice site. It is known to weaken U1 snRNP recognition and result in aberrantly spliced mRNA product in neuronal tissue, but normally spliced mRNA in other tissues. Aberrantly spliced IKBKAP mRNA abrogates IKK complex-associated protein (IKAP)/elongator protein 1 (ELP1) expression and results in a defect of neuronal cell development in FD...
June 7, 2017: RNA
https://www.readbyqxmd.com/read/28588079/focal-accumulation-of-preribosomes-outside-the-nucleolus-during-metaphase-anaphase-in-budding-yeast
#16
Giulia Moriggi, Sonia G Gaspar, Blanca Nieto, Xosé R Bustelo, Mercedes Dosil
Saccharomyces cerevisiae contains one nucleolus that remains intact in the mother-cell side of the nucleus throughout most of mitosis. Based on this, it is assumed that the bulk of ribosome production during cell division occurs in the mother cell. Here, we show that the ribosome synthesis machinery localizes not only in the nucleolus but also at a center that is present in the bud side of the nucleus after the initiation of mitosis. This center can be visualized by live microscopy as a punctate body located in close proximity to the nuclear envelope and opposite to the nucleolus...
June 6, 2017: RNA
https://www.readbyqxmd.com/read/28576826/archaeal-fibrillarin-nop5-heterodimer-2-o-methylates-rna-independently-of-the-c-d-guide-rnp-particle
#17
Miglė Tomkuvienė, Janina Ličytė, Ingrida Olendraitė, Zita Liutkevičiūtė, Béatrice Clouet-d'Orval, Saulius Klimašauskas
Archaeal fibrillarin (aFib) is a well-characterized S-Adenosyl methionine (SAM)-dependent RNA 2'-O-methyltransferase that is known to act in a large C/D ribonucleoprotein (RNP) complex together with Nop5 and L7Ae proteins and a box C/D guide RNA. In the reaction, the guide RNA is critical to direct the methylation reaction to a specific site in tRNA or rRNA by sequence complementarity. Here we show that Pyrococcus abyssi aFib-Nop5 heterodimer can alone perform a SAM-dependent 2'-O-methylation of 16S and 23S ribosomal RNAs in vitro independently of L7Ae and C/D guide RNAs...
June 2, 2017: RNA
https://www.readbyqxmd.com/read/28576825/identification-of-an-optimized-2-o-methylated-tri-nucleotide-rna-motif-inhibiting-toll-like-receptor-7-and-8
#18
Felix C F Schmitt, Isabel Freund, Markus A Weigand, Mark Helm, Alexander H Dalpke, Tatjana Eigenbrod
Bacterial RNA serves an important function as activator of the innate immune system. In humans bacterial RNA is sensed by the endosomal receptors TLR7 and TLR8. Differences in the posttranscriptional modification profile of prokaryotic as compared to eukaryotic RNA allow innate immune cells to discriminate between 'host' and 'foreign' RNA. Ribose 2'-O-methylation is of particular importance and has been reported to antagonize TLR7/8 activation. Yet, the exact sequence context in which 2'-O-methylation has to occur to mediate its inhibitory activity remains largely undefined...
June 2, 2017: RNA
https://www.readbyqxmd.com/read/28559491/antagonistic-actions-of-two-human-pan3-isoforms-on-global-mrna-turnover
#19
Chyi-Ying A Chen, Yueqiang Zhang, Yu Xiang, Leng Han, Ann-Bin Shyu
Deadenylation is a fundamental process that regulates eukaryotic gene expression. Mammalian deadenylation exhibits biphasic kinetics, with the Pan2-Pan3 and Ccr4-Caf1 deadenylase complexes mediating the first and second phase, respectively; however, the significance of the biphasic nature of deadenylation in mRNA turnover remains unclear. In this study, we discovered that two distinct isoforms of human Pan3 display opposing properties necessary for coordinating the two phases of deadenylation. The shorter isoform (Pan3S) interacts more strongly with PABP than the longer isoform (Pan3L) does...
May 30, 2017: RNA
https://www.readbyqxmd.com/read/28559490/adar-rna-editing-below-the-backbone
#20
Liam P Keegan, Anzer Khan, Dragana Vukic, Mary A O'Connell
ADAR RNA editing enzymes (Adenosine Deaminases acting on RNA), that convert adenosine bases to inosines were first identified biochemically thirty years ago. Since then studies on ADARs in genetic model organisms, and evolutionary comparisons between them, continue to reveal a surprising range of pleiotropic biological effects of ADARs. This review focuses on Drosophila melanogaster, which has a single Adar gene encoding a homolog of vertebrate ADAR2 that site-specifically edits hundreds of transcripts to change individual codons in ion channel subunits and membrane and cytoskeletal proteins...
May 30, 2017: RNA
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