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Ruslan Aphasizhev, Takuma Suematsu, Liye Zhang, Inna Aphasizheva
RNA uridylation is a significant transcriptome-shaping factor in protists, fungi, metazoans, and plants. The 3' U-additions are catalyzed by terminal uridyltransferases (TUTases), a diverse group of enzymes that along with non-canonical poly(A) polymerases form a distinct group in the superfamily of DNA polymerase β-like nucleotidyl transferases. Within and across studied organisms and subcellular compartments, TUTases differ in nucleotide triphosphate selectivity, interacting partners, and RNA targets. A general premise linking RNA uridylation to 3'-5' degradation received support from several studies of small RNAs and mRNA turnover...
October 7, 2016: RNA Biology
Katie Schaukowitch, Jae-Yeol Joo, Tae-Kyung Kim
With the many advances in genome-wide sequencing, it has been discovered that much more of the genome is transcribed into RNA than previously appreciated. These nonprotein-coding RNAs (ncRNAs) come in many different forms, and they have been shown to have a variety of functions within the cell, influencing processes such as gene expression, mRNA splicing, and transport, just as a few examples. As we delve deeper into studying their mechanisms of action, it becomes important to understand how they play these roles, in particular by understanding what proteins these ncRNAs interact with...
2017: Methods in Molecular Biology
Miriam Gagliardi, Maria R Matarazzo
The relevance of RNA-protein interactions in modulating mRNA and noncoding RNA function is increasingly appreciated and several methods have been recently developed to map them. The RNA immunoprecipitation (RIP) is a powerful method to study the physical association between individual proteins and RNA molecules in vivo. The basic principles of RIP are very similar to those of chromatin immunoprecipitation (ChIP), a largely used tool in the epigenetic field, but with some important caveats. The approach is based on the use of a specific antibody raised against the protein of interest to pull down the RNA-binding protein (RBP) and target-RNA complexes...
2016: Methods in Molecular Biology
Tomoko Fujino, Takeru Suzuki, Koudai Okada, Kanako Kogashi, Ken-Ichi Yasumoto, Kazuhiro Sogawa, Hiroyuki Isobe
A method for the synthesis of chimeric oligonucleotides was developed to incorporate purine nucleobases and multiple triazole linkers in natural, phosphate-linked structures of RNA. A solution-phase synthesis method for triazole-linked RNA oligomers via copper-catalyzed azide-alkyne cycloaddition reaction was optimized and tolerated purine nucleobases and protecting groups for further transformations. Three (TL)RNA trinucleotides with 5'-protected hydroxy and 3'-phosphoramidite groups were prepared, and one congener with a representative sequence was subjected to automated, solid-phase phosphoramidite synthesis...
October 7, 2016: Journal of Organic Chemistry
Arkady Mustaev, Christal L Vitiello, Max E Gottesman
The coliphage HK022 protein Nun transcription elongation arrest factor inhibits RNA polymerase translocation. In vivo, Nun acts specifically to block transcription of the coliphage λ chromosome. Using in vitro assays, we demonstrate that Nun cross-links RNA in an RNA:DNA hybrid within a ternary elongation complex (TEC). Both the 5' and the 3' ends of the RNA cross-link Nun, implying that Nun contacts RNA polymerase both at the upstream edge of the RNA:DNA hybrid and in the vicinity of the catalytic center...
August 2, 2016: Proceedings of the National Academy of Sciences of the United States of America
Katharina Höfer, Sisi Li, Florian Abele, Jens Frindert, Jasmin Schlotthauer, Julia Grawenhoff, Jiamu Du, Dinshaw J Patel, Andres Jäschke
RNA capping and decapping are thought to be distinctive features of eukaryotes. The redox cofactor NAD was recently discovered to be attached to small regulatory RNAs in bacteria in a cap-like manner, and Nudix hydrolase NudC was found to act as a NAD-decapping enzyme in vitro and in vivo. Here, crystal structures of Escherichia coli NudC in complex with substrate NAD and with cleavage product NMN reveal the catalytic residues lining the binding pocket and principles underlying molecular recognition of substrate and product...
September 2016: Nature Chemical Biology
Deborah L Johnson, Bangyan L Stiles
PTEN is a critical tumor suppressor whose dysregulation leads to metabolic disease and cancer. How these diseases are linked at a molecular level is poorly understood. Maf1 is a novel PTEN target that connects PTEN's ability to repress intracellular lipid accumulation with its tumor suppressor function. Maf1 represses the expression of rRNAs and tRNAs to restrain biosynthetic capacity and oncogenic transformation. Recent studies demonstrate that Maf1 also controls intracellular lipid accumulation. In animal models, dysregulation of RNA polymerase I- and III-dependent transcription, and subsequent upregulation of rRNAs and tRNAs, leads to altered lipid metabolism and storage...
October 2016: Trends in Endocrinology and Metabolism: TEM
Simon Ketterer, Lukas Gladis, Adnan Kozica, Matthias Meier
A set of 12 fluorogenic glycine riboswitches with different thermodynamic and kinetic response properties was engineered. For the design of functional riboswitches, a three-part RNA approach was applied based on the idea of linking a RNA sensor, transmitter and actuator part together. For the RNA sensor and actuator part, we used the tandem glycine aptamer structure from Bacillus subtillis, and fluorogenic aptamer Spinach, respectively. To achieve optimal signal transduction from the sensor to the actuator, a riboswitch library with variable transmitter was screened with a microfluidic large-scale integration chip...
July 8, 2016: Nucleic Acids Research
Albert Remus R Rosana, Denise S Whitford, Richard P Fahlman, George W Owttrim
UNLABELLED: The cyanobacterium Synechocystis sp. strain PCC 6803 encodes a single DEAD box RNA helicase, CrhR, whose expression is tightly autoregulated in response to cold stress. Subcellular localization and proteomic analysis results indicate that CrhR localizes to both the cytoplasmic and thylakoid membrane regions and cosediments with polysome and RNA degradosome components. Evidence is presented that either functional RNA helicase activity or a C-terminal localization signal was required for polysome but not thylakoid membrane localization...
August 1, 2016: Journal of Bacteriology
Walid Mahrez, Juhyun Shin, Rafael Muñoz-Viana, Duarte D Figueiredo, Minerva S Trejo-Arellano, Vivien Exner, Alexey Siretskiy, Wilhelm Gruissem, Claudia Köhler, Lars Hennig
Several pathways control time to flowering in Arabidopsis thaliana through transcriptional and posttranscriptional gene regulation. In recent years, mRNA processing has gained interest as a critical regulator of flowering time control in plants. However, the molecular mechanisms linking RNA splicing to flowering time are not well understood. In a screen for Arabidopsis early flowering mutants we identified an allele of BRR2a. BRR2 proteins are components of the spliceosome and highly conserved in eukaryotes...
April 2016: PLoS Genetics
Wenhu Zhou, Jinsong Ding, Juewen Liu
RNA can form two types of linkage. In addition to the predominant 3'-5' linkage, 2'-5'-linked RNA is also important in biology, medicine, and prebiotic studies. Here, in vitro selection was used to isolate a DNAzyme that specifically cleaves 2'-5' RNA by using Ce(3+) as the metal cofactor, but leaves the 3'-5' counterpart intact. This Ce5 DNAzyme requires trivalent light lanthanide ions and shows a rate of 0.16 min(-1) in the presence of 10 μm Ce(3+) ; the activity decreases with heavier lanthanide ions...
May 17, 2016: Chembiochem: a European Journal of Chemical Biology
Maïwen Caudron-Herger, Teresa Pankert, Jeanette Seiler, Attila Németh, Renate Voit, Ingrid Grummt, Karsten Rippe
Non-coding RNAs play a key role in organizing the nucleus into functional subcompartments. By combining fluorescence microscopy and RNA deep-sequencing-based analysis, we found that RNA polymerase II transcripts originating from intronic Alu elements (aluRNAs) were enriched in the nucleolus. Antisense-oligo-mediated depletion of aluRNAs or drug-induced inhibition of RNA polymerase II activity disrupted nucleolar structure and impaired RNA polymerase I-dependent transcription of rRNA genes. In contrast, overexpression of a prototypic aluRNA sequence increased both nucleolus size and levels of pre-rRNA, suggesting a functional link between aluRNA, nucleolus integrity and pre-rRNA synthesis...
November 12, 2015: EMBO Journal
Loyal A Goff, John L Rinn
The regulatory potential of RNA has never ceased to amaze: from RNA catalysis, to RNA-mediated splicing, to RNA-based silencing of an entire chromosome during dosage compensation. More recently, thousands of long noncoding RNA (lncRNA) transcripts have been identified, the majority with unknown function. Thus, it is tempting to think that these lncRNAs represent a cadre of new factors that function through ribonucleic mechanisms. Some evidence points to several lncRNAs with tantalizing physiological contributions and thought-provoking molecular modalities...
October 2015: Genome Research
Andrea B Eberle, Antonio Jordán-Pla, Antoni Gañez-Zapater, Viktoria Hessle, Gilad Silberberg, Anne von Euler, Rebecca A Silverstein, Neus Visa
RNA surveillance factors are involved in heterochromatin regulation in yeast and plants, but less is known about the possible roles of ribonucleases in the heterochromatin of animal cells. Here we show that RRP6, one of the catalytic subunits of the exosome, is necessary for silencing heterochromatic repeats in the genome of Drosophila melanogaster. We show that a fraction of RRP6 is associated with heterochromatin, and the analysis of the RRP6 interaction network revealed physical links between RRP6 and the heterochromatin factors HP1a, SU(VAR)3-9 and RPD3...
September 2015: PLoS Genetics
Romina J Gentier, Fred W van Leeuwen
Amyloid β (Aβ) plaque formation is a prominent cellular hallmark of Alzheimer's disease (AD). To date, immunization trials in AD patients have not been effective in terms of curing or ameliorating dementia. In addition, γ-secretase inhibitor strategies await clinical improvements in AD. These approaches were based upon the idea that autosomal dominant mutations in amyloid precursor protein (APP) and Presenilin 1 (PS1) genes are predictive for treatment of all AD patients. However most AD patients are of the sporadic form which partly explains the failures to treat this multifactorial disease...
2015: Frontiers in Molecular Neuroscience
Tea Kaartokallio, Alejandra Cervera, Anjuska Kyllönen, Krista Laivuori
Pre-eclampsia is a common and complex pregnancy disorder that often involves impaired placental development. In order to identify altered gene expression in pre-eclamptic placenta, we sequenced placental transcriptomes of nine pre-eclamptic and nine healthy pregnant women in pools of three. The differential gene expression was tested both by including all the pools in the analysis and by excluding some of the pools based on phenotypic characteristics. From these analyses, we identified altogether 53 differently expressed genes, a subset of which was validated by qPCR in 20 cases and 19 controls...
2015: Scientific Reports
Deepak Sumbria, Lachhman Das Singla, Amrita Sharma
A cross-sectional study was conducted in Submountain undulating, Undulating plain, Western and Western plain agro-climatic zones of Punjab province, India, to determine the prevalence, agreement between diagnostic tests and associated related risk factors of Theileria equi and Babesia caballi infection in equids (horses, donkey, mules). An overall prevalence of 14.14 and 0.0% of T. equi and B. caballi was recorded by multiplex polymerase chain reaction targeting 18S ribosomal RNA (rRNA) for both the parasites and 75 and 1...
January 2016: Tropical Animal Health and Production
Werner E Bischoff, Rebecca J McNall, Maria W Blevins, JoLyn Turner, Elena N Lopareva, Paul A Rota, John R Stehle
Measles virus (MeV) is known to be highly contagious, with an infectious period lasting from 4 days before to 4 days after rash onset. An unvaccinated, young, female patient with measles confirmed by direct epidemiologic link was hospitalized on day 5 after rash onset. Environmental samples were collected over the 4-day period of hospitalization in a single room. MeV RNA was detectable in air specimens, on surface specimens, and on respirators on days 5-8 after rash onset. This is the first report of environmental surveillance for MeV, and the results suggest that MeV-infected fomites may be present in healthcare settings...
February 15, 2016: Journal of Infectious Diseases
Mayssa H Mokalled, Kelli J Carroll, Bercin K Cenik, Beibei Chen, Ning Liu, Eric N Olson, Rhonda Bassel-Duby
Myocardin-Related Transcription Factors A and B (MRTF-A and MRTF-B) are highly homologous proteins that function as powerful coactivators of serum response factor (SRF), a ubiquitously expressed transcription factor essential for cardiac development. The SRF/MRTF complex binds to CArG boxes found in the control regions of genes that regulate cytoskeletal dynamics and muscle contraction, among other processes. While SRF is required for heart development and function, the role of MRTFs in the developing or adult heart has not been explored...
October 15, 2015: Developmental Biology
May Christine V Malicdan, Thierry Vilboux, Joshi Stephen, Dino Maglic, Luhe Mian, Daniel Konzman, Jennifer Guo, Deniz Yildirimli, Joy Bryant, Roxanne Fischer, Wadih M Zein, Joseph Snow, Meghana Vemulapalli, James C Mullikin, Camilo Toro, Benjamin D Solomon, John E Niederhuber, William A Gahl, Meral Gunay-Aygun
BACKGROUND: In chicken, loss of TALPID3 results in non-functional cilia and short-rib polydactyly syndrome. This phenotype is caused by a frameshift mutation in the chicken ortholog of the human KIAA0586 gene, which encodes a novel coiled-coil domain protein essential for primary ciliogenesis, suggesting that KIAA0586 can be associated with ciliopathy in human beings. METHODS: In our patients with ciliopathy ( NCT00068224), we have collected extensive clinical and neuroimaging data from affected individuals, and performed whole exome sequencing on DNA from affected individuals and their parents...
December 2015: Journal of Medical Genetics
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