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https://www.readbyqxmd.com/read/28163933/analysis-of-argonaute-complex-bound-mrnas-in-du145-prostate-carcinoma-cells-reveals-new-mirna-target-genes
#1
Jaroslaw Szczyrba, Volker Jung, Michaela Beitzinger, Elke Nolte, Sven Wach, Martin Hart, Sandra Sapich, Marc Wiesehöfer, Helge Taubert, Gunther Wennemuth, Norbert Eichner, Thomas Stempfl, Bernd Wullich, Gunter Meister, Friedrich A Grässer
Posttranscriptional gene regulation by microRNAs (miRNAs) contributes to the induction and maintenance of prostate carcinoma (PCa). To identify mRNAs enriched or removed from Ago2-containing RISC complexes, these complexes were immunoprecipitated from normal prostate fibroblasts (PNFs) and the PCa line DU145 and the bound mRNAs were quantified by microarray. The analysis of Ago complexes derived from PNFs or DU145 confirmed the enrichment or depletion of a variety of mRNAs already known from the literature to be deregulated...
2017: Prostate Cancer
https://www.readbyqxmd.com/read/28148824/biochemical-and-genetic-functional-dissection-of-the-p38-viral-suppressor-of-rna-silencing
#2
Taichiro Iki, Marie-Aude Tschopp, Olivier Voinnet
Phytoviruses encode viral suppressors of RNA-silencing (VSRs) to counteract the plant antiviral silencing response, which relies on virus-derived small interfering (si)RNAs processed by Dicer RNaseIII enzymes and subsequently loaded into ARGONAUTE (AGO) effector proteins. Here, a tobacco cell-free system was engineered to recapitulate the key steps of antiviral RNA-silencing and, in particular, the most upstream double-stranded (ds)RNA processing reaction, not kinetically investigated thus far in the context of plant VSR studies...
February 1, 2017: RNA
https://www.readbyqxmd.com/read/28127848/mir-30c-protects-diabetic-nephropathy-by-suppressing-epithelial-to-mesenchymal-transition-in-db-db-mice
#3
Yanru Zhao, Zhongwei Yin, Huaping Li, Jiahui Fan, Shenglan Yang, Chen Chen, Dao Wen Wang
Epithelial-to-mesenchymal transition (EMT) plays a significant role in tubulointerstitial fibrosis, which is a hallmark of diabetic nephropathy. Thus, identifying the mechanisms of EMT activation could be meaningful. In this study, loss of miR-30c accompanied with increased EMT was observed in renal tubules of db/db mice and cultured HK2 cells exposed to high glucose. To further explore the roles of miR-30c in EMT and tubulointerstitial fibrosis, recombinant adeno-associated viral vector was applied to manipulate the expression of miR-30c...
January 27, 2017: Aging Cell
https://www.readbyqxmd.com/read/28125794/development-of-novel-small-hairpin-rnas-that-do-not-require-processing-by-dicer-or-ago2
#4
Shin-Ichiro Ohno, Karen Itano, Yuichirou Harada, Koutaro Asada, Keiki Oikawa, Mikie Kashiwazako, Hikaru Okuyama, Katsuyoshi Kumagai, Masakatsu Takanashi, Katsuko Sudo, Norihiko Ikeda, Masahiko Kuroda
The innate cytokine response to nucleic acid is the most challenging problem confronting the practical use of nucleic acid medicine. The degree of stimulation of the innate cytokine response strongly depends on the length of the nucleic acid. In this study, we developed a 30-nucleotide single-strand RNA, termed "guide hairpin RNA (ghRNA, ghR)", that has a physiological function similar to that of miRNA and siRNA. The ghR caused no innate cytokine response either in vitro or in vivo. In addition, its structure does not contain a passenger strand seed sequence, reducing the unwanted gene repression relative to existing short RNA reagents...
July 2016: Molecular Therapy: the Journal of the American Society of Gene Therapy
https://www.readbyqxmd.com/read/28119422/non-canonical-targets-destabilize-micrornas-in-human-argonautes
#5
June Hyun Park, Sang-Yoon Shin, Chanseok Shin
Although much is known about microRNA (miRNA) biogenesis and the mechanism by which miRNAs regulate their targets, little is known about the regulation of miRNA stability. Mature miRNAs are stabilized by binding to Argonaute (Ago) proteins, the core components of the RNA-induced silencing complex (RISC). Recent studies suggest that interactions between miRNAs and their highly complementary target RNAs promote release of miRNAs from Ago proteins, and this in turn can lead to destabilization of miRNAs. However, the physiological triggers of miRNA destabilization with molecular mechanisms remain largely unknown...
January 24, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28118842/microglial-derived-mirna-let-7-and-hmgb1-contribute-to-ethanol-induced-neurotoxicity-via-tlr7
#6
Leon G Coleman, Jian Zou, Fulton T Crews
BACKGROUND: Toll-like receptor (TLR) signaling is emerging as an important component of neurodegeneration. TLR7 senses viral RNA and certain endogenous miRNAs to initiate innate immune responses leading to neurodegeneration. Alcoholism is associated with hippocampal degeneration, with preclinical studies linking ethanol-induced neurodegeneration with central innate immune induction and TLR activation. The endogenous miRNA let-7b binds TLR7 to cause neurodegeneration. METHODS: TLR7 and other immune markers were assessed in postmortem human hippocampal tissue that was obtained from the New South Wales Tissue Bank...
January 25, 2017: Journal of Neuroinflammation
https://www.readbyqxmd.com/read/28114302/an-argonaute-phosphorylation-cycle-promotes-microrna-mediated-silencing
#7
Ryan J Golden, Beibei Chen, Tuo Li, Juliane Braun, Hema Manjunath, Xiang Chen, Jiaxi Wu, Vanessa Schmid, Tsung-Cheng Chang, Florian Kopp, Andres Ramirez-Martinez, Vincent S Tagliabracci, Zhijian J Chen, Yang Xie, Joshua T Mendell
MicroRNAs (miRNAs) perform critical functions in normal physiology and disease by associating with Argonaute proteins and downregulating partially complementary messenger RNAs (mRNAs). Here we use clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 9 (Cas9) genome-wide loss-of-function screening coupled with a fluorescent reporter of miRNA activity in human cells to identify new regulators of the miRNA pathway. By using iterative rounds of screening, we reveal a novel mechanism whereby target engagement by Argonaute 2 (AGO2) triggers its hierarchical, multi-site phosphorylation by CSNK1A1 on a set of highly conserved residues (S824-S834), followed by rapid dephosphorylation by the ANKRD52-PPP6C phosphatase complex...
February 9, 2017: Nature
https://www.readbyqxmd.com/read/28108660/from-benchmarking-hits-clip-peak-detection-programs-to-a-new-method-for-identification-of-mirna-binding-sites-from-ago2-clip-data
#8
Silvia Bottini, Nedra Hamouda-Tekaya, Bogdan Tanasa, Laure-Emmanuelle Zaragosi, Valerie Grandjean, Emanuela Repetto, Michele Trabucchi
Experimental evidence indicates that about 60% of miRNA-binding activity does not follow the canonical rule about the seed matching between miRNA and target mRNAs, but rather a non-canonical miRNA targeting activity outside the seed or with a seed-like motifs. Here, we propose a new unbiased method to identify canonical and non-canonical miRNA-binding sites from peaks identified by Ago2 Cross-Linked ImmunoPrecipitation associated to high-throughput sequencing (CLIP-seq). Since the quality of peaks is of pivotal importance for the final output of the proposed method, we provide a comprehensive benchmarking of four peak detection programs, namely CIMS, PIPE-CLIP, Piranha and Pyicoclip, on four publicly available Ago2-HITS-CLIP datasets and one unpublished in-house Ago2-dataset in stem cells...
January 20, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28088781/mir-30c-5p-ameliorates-hepatic-steatosis-in-leptin-receptor-deficient-db-db-mice-via-down-regulating-fasn
#9
Jiahui Fan, Huaping Li, Xiang Nie, Zhongwei Yin, Yanru Zhao, Chen Chen, Dao Wen Wang
Approximately 15-40% of the general adult population suffers from non-alcoholic fatty liver disease (NAFLD) worldwide. However, no drug is currently licensed for its treatment. In this study, we observed a significant reduction of miR-30c-5p in the liver of leptin receptor-deficient (db/db) mice. Remarkably, recombinant adeno-associated virus (rAAV)-mediated delivery of miR-30c-5p was sufficient to attenuate triglyceride accumulation and hepatic steatosis in db/db mice. Through computational prediction, KEGG analysis and Ago2 co-immunoprecipitation, we identified that miR-30c-5p directly targeted fatty acid synthase, a key enzyme in fatty acid biosynthesis...
January 9, 2017: Oncotarget
https://www.readbyqxmd.com/read/28087276/drosophila-argonaute2-turnover-is-regulated-by-the-ubiquitin-proteasome-pathway
#10
Madoka Chinen, Elissa P Lei
Argonaute (AGO) proteins play a central role in the RNA interference (RNAi) pathway, which is a cytoplasmic mechanism important for post-transcriptional regulation of gene expression. In Drosophila, AGO2 also functions in the nucleus to regulate chromatin insulator activity and transcription. Although there are a number of studies focused on AGO2 function, the regulation of AGO2 turnover is not well understood. We found that mutation of T1149 or R1158 in the conserved PIWI domain causes AGO2 protein instability, but only T1149 affects RNAi activity...
February 12, 2017: Biochemical and Biophysical Research Communications
https://www.readbyqxmd.com/read/28075055/argonaute-2-immunoprecipitation-revealed-large-tumor-suppressor-kinase-1-as-a-novel-proapoptotic-target-of-mir-21-in-t-cells
#11
Nato Teteloshvili, Katarzyna Smigielska-Czepiel, Ye Yuan, Annika Seitz, Debora de Jong, Bea Rutgers, Pytrick Jellema, Roelof Jan van der Lei, Izabella Slezak-Prochazka, Elisabeth Brouwer, Annemieke M H Boots, Bart-Jan Kroesen, Anke van den Berg, Joost Kluiver
MicroRNA (miR)-21 is an important suppressor of T-cell apoptosis that is also overexpressed in many types of cancers. The exact mechanisms underlying the antiapoptotic effects of miR-21 are not well understood. In this study, we used the Jurkat T-cell line as a model to identify apoptosis-associated miR-21 target genes. We showed that expression of miR-21 rapidly increases upon αCD3/αCD28 activation of Jurkat cells. Inhibition of miR-21 reduced cell growth which could be explained by an increase in apoptosis...
January 11, 2017: FEBS Journal
https://www.readbyqxmd.com/read/28053119/short-intron-derived-ncrnas
#12
Florent Hubé, Damien Ulveling, Alain Sureau, Sabrina Forveille, Claire Francastel
Introns represent almost half of the human genome, although they are eliminated from transcripts through RNA splicing. Yet, different classes of non-canonical miRNAs have been proposed to originate directly from intron splicing. Here, we considered the alternative splicing of introns as an interesting source of miRNAs, compatible with a developmental switch. We report computational prediction of new Short Intron-Derived ncRNAs (SID), defined as precursors of smaller ncRNAs like miRNAs and snoRNAs produced directly by splicing, and tested their dependence on each key factor in canonical or alternative miRNAs biogenesis (Drosha, DGCR8, DBR1, snRNP70, U2AF65, PRP8, Dicer, Ago2)...
January 3, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28030800/the-role-of-mir-17-92-in-the-miregulatory-landscape-of-ewing-sarcoma
#13
Raphaela Schwentner, David Herrero-Martin, Maximilian O Kauer, Cornelia N Mutz, Anna M Katschnig, Grzegorz Sienski, Javier Alonso, Dave Nt Aryee, Heinrich Kovar
MicroRNAs serve to fine-tune gene expression and play an important regulatory role in tissue specific gene networks. The identification and validation of miRNA target genes in a tissue still poses a significant problem since the presence of a seed sequence in the 3'UTR of an mRNA and its expression modulation upon ectopic expression of the miRNA do not reliably predict regulation under physiological conditions. The chimeric oncoprotein EWS-FLI1 is the driving pathogenic force in Ewing sarcoma. MiR-17-92, one of the most potent oncogenic miRNAs, was recently reported to be among the top EWS-FLI1 activated miRNAs...
December 22, 2016: Oncotarget
https://www.readbyqxmd.com/read/28008821/cooperative-enhancement-of-translation-by-two-adjacent-microrna-122-argonaute-2-complexes-binding-to-the-5%C3%A2-untranslated-region-of-hepatitis-c-virus-rna
#14
Anika Nieder-Röhrmann, Nadia Dünnes, Gesche K Gerresheim, Lyudmila A Shalamova, Andreas Herchenröther, Michael Niepmann
The liver-specific microRNA-122 (miR-122) binds to two conserved binding sites in the 5´-untranslated region (5´UTR) of Hepatitis C Virus (HCV) RNA. This binding was reported to enhance HCV RNA replication, translation and stability. We have analyzed binding of miR-122/Argonaute 2 (Ago2) complexes to these sites using anti-Ago2-co-immunoprecipitation of radioactively labeled HCV RNAs along with ectopic miR-122 in HeLa cells. Our results show that the miR-122 target sites can be addressed separately. When both target sites were addressed simultaneously, we observed a synergistic binding of both miR/Ago2 complexes...
December 23, 2016: Journal of General Virology
https://www.readbyqxmd.com/read/28003477/argonaute-proteins-regulate-hiv-1-multiply-spliced-rna-and-viral-production-in-a-dicer-independent-manner
#15
Agathe Eckenfelder, Emmanuel Ségéral, Natalia Pinzón, Damien Ulveling, Céline Amadori, Marine Charpentier, Sabine Nidelet, Jean-Paul Concordet, Jean-François Zagury, Jean-Christophe Paillart, Clarisse Berlioz-Torrent, Hervé Seitz, Stéphane Emiliani, Sarah Gallois-Montbrun
Argonaute (Ago) proteins associate with microRNAs (miRNAs) to form the core of the RNA-induced silencing complex (RISC) that mediates post-transcriptional gene silencing of target mRNAs. As key players in anti-viral defense, Ago proteins are thought to have the ability to interact with human immunodeficiency virus type 1 (HIV-1) RNA. However, the role of this interaction in regulating HIV-1 replication has been debated. Here, we used high throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP) to explore the interaction between Ago2 and HIV-1 RNA in infected cells...
December 20, 2016: Nucleic Acids Research
https://www.readbyqxmd.com/read/27928938/sequence-and-structure-based-analysis-of-proteins-involved-in-mirna-biogenesis
#16
Chhaya Sharma, Debasisa Mohanty
miRNA biogenesis is a multistage process for generation of a mature miRNA and involves several different proteins. In this work, we have carried out sequence as well as structure based analysis for crucial proteins involved in miRNA biogenesis, namely, Dicer, Drosha, Argonaute (Ago) and Exportin-5 to understand evolution of these proteins in animal kingdom and also to identify key sequence and structural features which are determinants of their function. Our analysis reveals that in animals the miRNA biogenesis pathway first originated in molluscs...
December 8, 2016: Journal of Biomolecular Structure & Dynamics
https://www.readbyqxmd.com/read/27928054/the-influence-of-the-5-terminal-nucleotide-on-agoshrna-activity-and-biogenesis-importance-of-the-polymerase-iii-transcription-initiation-site
#17
Elena Herrera-Carrillo, Zong-Liang Gao, Alex Harwig, Matthias T Heemskerk, Ben Berkhout
Recent evidence indicates that shRNAs with a relatively short basepaired stem do not require Dicer processing, but instead are processed by the Argonaute 2 protein (Ago2). We named these molecules AgoshRNAs as both their processing and silencing function are mediated by Ago2. This alternative processing yields only a single RNA guide strand, which can avoid off-target effects induced by the passenger strand of regular shRNAs. It is important to understand this alternative processing route in mechanistic detail such that one can design improved RNA reagents...
December 7, 2016: Nucleic Acids Research
https://www.readbyqxmd.com/read/27927008/microrna-296-5p-promotes-invasiveness-through-downregulation-of-nerve-growth-factor-receptor-and-caspase-8
#18
Hong Lee, Chang Hoon Shin, Hye Ree Kim, Kyung Hee Choi, Hyeon Ho Kim
Glioblastomas (GBM) are very difficult to treat and their aggressiveness is one of the main reasons for this as well as for the frequent recurrences. MicroRNAs posttranscriptionally regulate their target genes through interaction between their seed sequence and 3'UTR of the target mRNAs. We previously reported that miR-296-3p is regulated by neurofibromatosis 2 (NF2) and enhances the invasiveness of GBM cells via SOCS2/STAT3. In this study, we investigated whether miR-296-5p, which originates from the same precursor miRNA as miR-296-3p, can increase the invasiveness of GBM cells...
December 8, 2016: Molecules and Cells
https://www.readbyqxmd.com/read/27924488/elucidating-mechanisms-of-molecular-recognition-between-human-argonaute-and-mirna-using-computational-approaches
#19
Hanlun Jiang, Lizhe Zhu, Amélie Héliou, Xin Gao, Julie Bernauer, Xuhui Huang
MicroRNA (miRNA) and Argonaute (AGO) protein together form the RNA-induced silencing complex (RISC) that plays an essential role in the regulation of gene expression. Elucidating the underlying mechanism of AGO-miRNA recognition is thus of great importance not only for the in-depth understanding of miRNA function but also for inspiring new drugs targeting miRNAs. In this chapter we introduce a combined computational approach of molecular dynamics (MD) simulations, Markov state models (MSMs), and protein-RNA docking to investigate AGO-miRNA recognition...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/27924485/small-molecules-targeting-the-mirna-binding-domain-of-argonaute-2-from-computer-aided-molecular-design-to-rna-immunoprecipitation
#20
Teresa Bellissimo, Silvia Masciarelli, Elena Poser, Ilaria Genovese, Alberto Del Rio, Gianni Colotti, Francesco Fazi
The development of small-molecule-based target therapy design for human disease and cancer is object of growing attention. Recently, specific microRNA (miRNA) mimicking compounds able to bind the miRNA-binding domain of Argonaute 2 protein (AGO2) to inhibit miRNA loading and its functional activity were described. Computer-aided molecular design techniques and RNA immunoprecipitation represent suitable approaches to identify and experimentally determine if a compound is able to impair the loading of miRNAs on AGO2 protein...
2017: Methods in Molecular Biology
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