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https://www.readbyqxmd.com/read/28817234/osmium-mediated-4su-to-c-transformation-as-key-to-study-rna-dynamics-by-sequencing-tuc-seq
#1
Christian Riml, Thomas Amort, Dietmar Rieder, Catherina Gasser, Alexandra Lusser, Ronald Micura
To understand the functional roles of RNA in the cell, it is essential to elucidate the dynamics of their production, processing and decay. A recent method for assessing mRNA dynamics is metabolic labeling with 4-thiouridine (4sU), followed by thio-selective attachment of affinity tags. Detection of labeled transcripts by affinity purification and hybridization to microarrays or by deep sequencing then reveals RNA expression levels. Here, we present a novel sequencing method that eliminates affinity purification and allows for direct assessment of 4sU labeled RNA...
August 17, 2017: Angewandte Chemie
https://www.readbyqxmd.com/read/28806774/fibroblast-growth-factor-receptor-1-mediates-internalization-of-pathogenic-spotted-fever-rickettsiae-into-host-endothelium
#2
Abha Sahni, Jignesh Patel, Hema P Narra, Casey L C Schroeder, David H Walker, Sanjeev K Sahni
Rickettsial infections continue to cause serious morbidity and mortality in severe human cases around the world. Host cell adhesion and invasion is an essential requisite for intracellular growth, replication, and subsequent dissemination of pathogenic rickettsiae. Heparan sulfate proteoglycans [HSPGs] facilitate the interactions between fibroblast growth factor(s) and their tyrosine kinase receptors resulting in receptor dimerization/activation and have been implicated in bacterial adhesion to target host cells...
2017: PloS One
https://www.readbyqxmd.com/read/28800637/biologically-active-recombinant-human-erythropoietin-expressed-in-hairy-root-cultures-and-regenerated-plantlets-of-nicotiana-tabacum-l
#3
Poornima Devi Gurusamy, Holger Schäfer, Siva Ramamoorthy, Michael Wink
Hairy root culture is a potential alternative to conventional mammalian cell culture to produce recombinant proteins due to its ease in protein recovery, low costs and absence of potentially human pathogenic contaminants. The current study focussed to develop a new platform of a hairy root culture system from Nicotiana tabacum for the production of recombinant human EPO (rhEPO), which is regularly produced in mammalian cells. The human EPO construct was amplified with C-terminal hexahistidine tag from a cDNA of Caco-2 cells...
2017: PloS One
https://www.readbyqxmd.com/read/28769958/oryza-sativa-brassinosteroid-upregulated1-like1-induces-the-expression-of-a-gene-encoding-a-small-leucine-rich-repeat-protein-to-positively-regulate-lamina-inclination-and-grain-size-in-rice
#4
Seonghoe Jang, Hsing-Yi Li
Oryza sativa BRASSINOSTEROID UPREGULATED1 LIKE1 (OsBUL1) positively affects lamina inclination and grain size. OsBUL1 knock-out (osbul1) plants as well as transgenic rice with reduced level of OsBUL1 expression produce erect leaves and small grains. Here, we identified a putative downstream gene of OsBUL1, OsBUL1 DOWNSTREAM GENE1 (OsBDG1) encoding a small protein with short leucine-rich-repeats by cDNA microarray analyses in the lamina joint and panicles of wild-type and osbul1 plants. Transgenic rice plants with increased OsBDG1 expression exhibit increased leaf angle and grain size, which is similar to an OsBDG1 activation tagging line whereas double stranded RNA interference (dsRNAi) lines for OsBDG1 knock-down generate erect leaves with smaller grains...
2017: Frontiers in Plant Science
https://www.readbyqxmd.com/read/28768715/architecture-and-dynamics-of-overlapped-rna-regulatory-networks
#5
Christopher P Lapointe, Melanie A Preston, Daniel Wilinski, Harriet A J Saunders, Zachary T Campbell, Marvin Wickens
A single protein can bind and regulate many mRNAs. Multiple proteins with similar specificities often bind and control overlapping sets of mRNAs. Yet, little is known about the architecture or dynamics of overlapped networks. We focused on three proteins with similar structures and related RNA-binding specificities ‒ Puf3p, Puf4p, and Puf5p of S. cerevisiae. Using RNA Tagging, we identified a "super-network" comprised of four sub-networks: Puf3p, Puf4p, and Puf5p sub-networks, and one controlled by both Puf4p and Puf5p...
August 2, 2017: RNA
https://www.readbyqxmd.com/read/28767232/rna-polymerase-tags-to-monitor-multidimensional-protein-protein-interactions-reveal-pharmacological-engagement-of-bcl-2-proteins
#6
Jinyue Pu, Jeffrey A Dewey, Abbas Hadji, James L LaBelle, Bryan C Dickinson
We report the development of a new technology for monitoring multidimensional protein-protein interactions (PPIs) inside live mammalian cells using split RNA polymerase (RNAP) tags. In this new system, a protein-of-interest is tagged with an N-terminal split RNAP (RNAPN), and multiple potential binding partners are each fused to orthogonal C-terminal RNAPs (RNAPC). Assembly of RNAPN with each RNAPC is highly dependent on interactions between the tagged proteins. Each PPI-mediated RNAPN-RNAPC assembly transcribes from a separate promoter on a supplied DNA substrate, thereby generating a unique RNA output signal for each PPI...
August 15, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28766289/purification-of-mrna-processing-complexes-using-an-rna-affinity-approach
#7
Xiuye Wang, Yongsheng Shi
Multiple mRNA processing steps, including splicing and 3' processing, take place in macromolecular complexes that contain many proteins and sometimes RNA molecules. A key challenge in the mRNA processing field has been to define the structure-function relationship of these sophisticated molecular machines. A prerequisite for addressing this challenge is to develop tools for purifying mRNA processing complexes in their native and intact forms that are suitable for functional and structural studies. Among many methods that have been developed, RNA affinity-based methods are most widely applied...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28765217/rip-seq-suggests-translational-regulation-by-l7ae-in-archaea
#8
Michael Daume, Michael Uhl, Rolf Backofen, Lennart Randau
L7Ae is a universal archaeal protein that recognizes and stabilizes kink-turn (k-turn) motifs in RNA substrates. These structural motifs are widespread in nature and are found in many functional RNA species, including ribosomal RNAs. Synthetic biology approaches utilize L7Ae/k-turn interactions to control gene expression in eukaryotes. Here, we present results of comprehensive RNA immunoprecipitation sequencing (RIP-Seq) analysis of genomically tagged L7Ae from the hyperthermophilic archaeon Sulfolobus acidocaldarius A large set of interacting noncoding RNAs was identified...
August 1, 2017: MBio
https://www.readbyqxmd.com/read/28761931/unc5c-variants-are-associated-with-cerebral-amyloid-angiopathy
#9
Hyun-Sik Yang, Charles C White, Lori B Chibnik, Hans-Ulrich Klein, Julie A Schneider, David A Bennett, Philip L De Jager
OBJECTIVE: To determine whether common genetic variants in UNC5C, a recently identified late-onset Alzheimer disease (LOAD) dementia susceptibility gene, are associated with AD susceptibility or AD-related clinical/pathologic phenotypes. METHODS: We used data from deceased individuals of European descent who participated in the Religious Orders Study or the Rush Memory and Aging Project (n = 1,288). We examined whether there were associations between single nucleotide polymorphisms (SNPs) within ±100 kb of the UNC5C gene and a diagnosis of AD dementia, global cognitive decline, a pathologic diagnosis of AD, β-amyloid load, neuritic plaque count, diffuse plaque count, paired helical filament tau density, neurofibrillary tangle count, and cerebral amyloid angiopathy (CAA) score...
August 2017: Neurology. Genetics
https://www.readbyqxmd.com/read/28757447/the-phylogeny-of-extant-starfish-asteroidea-echinodermata-including-xyloplax-based-on-comparative-transcriptomics
#10
Gregorio V Linchangco, David W Foltz, Rob Reid, John Williams, Conor Nodzak, Alexander M Kerr, Allison K Miller, Rebecca Hunter, Nerida G Wilson, William J Nielsen, Christopher L Mah, Greg W Rouse, Gregory A Wray, Daniel A Janies
Multi-locus phylogenetic studies of echinoderms based on Sanger and RNA-seq technologies and the fossil record have provided evidence for the Asterozoa-Echinozoa hypothesis. This hypothesis posits a sister relationship between asterozoan classes (Asteroidea and Ophiuroidea) and a similar relationship between echinozoan classes (Echinoidea and Holothuroidea). Despite this consensus around Asterozoa-Echinozoa, phylogenetic relationships within the class Asteroidea (sea stars or starfish) have been controversial for over a century...
July 27, 2017: Molecular Phylogenetics and Evolution
https://www.readbyqxmd.com/read/28752326/modified-whole-mount-in-situ-hybridisation-and-immunohistochemistry-protocols-without-removal-of-the-vitelline-membrane-in-the-appendicularian-oikopleura-dioica
#11
Takeshi A Onuma, Masaki Matsuo, Hiroki Nishida
The appendicularian Oikopleura dioica is a planktonic chordate that retains a tadpole shape throughout its life. Its simple and transparent body, invariant cell lineages, fast development and available genome and transcriptome resources make it a promising model organism for research in developmental biology. However, large-scale analysis of gene expression in O. dioica is limited owing to the laborious and time-consuming process of manual removal of the vitelline membrane, because devitellinisation of pre-hatching embryos causes failure of normal development...
July 27, 2017: Development Genes and Evolution
https://www.readbyqxmd.com/read/28752295/nmr-based-metabolomics-reveals-the-metabolite-profiles-of-vibrio-parahaemolyticus-under-ferric-iron-stimulation
#12
Jun Zhou, Chenyang Lu, Dijun Zhang, Chennv Ma, Xiurong Su
Vibrio parahaemolyticus is a halophilic bacterium endemic to coastal areas, and its pathogenicity has caused widespread seafood poisoning. In our previous research, the protein expression of V. parahaemolyticus in Fe(3+) medium was determined using isobaric tags for relative and absolute quantitation (iTRAQ). Here, nuclear magnetic resonance (NMR) was used to detect changes in the V. parahaemolyticus metabolome. NMR spectra were obtained using methanol-water extracts of intracellular metabolites from V. parahaemolyticus under various culture conditions, and 62 metabolites were identified, including serine, arginine, alanine, ornithine, tryptophan, glutamine, malate, NAD(+), NADP(+), oxypurinol, xanthosine, dCTP, uracil, thymine, hypoxanthine, and betaine...
August 2017: Journal of Microbiology / the Microbiological Society of Korea
https://www.readbyqxmd.com/read/28749687/site-specific-covalent-conjugation-of-modified-mrna-by-trna-guanine-transglycosylase
#13
Fabian Ehret, Cun Yu Zhou, Seth C Alexander, Dongyang Zhang, Neal K Devaraj
Modified messenger RNA (mod-mRNA) has recently been widely studied as the form of RNA useful for therapeutic applications due to its high stability and lowered immune response. Herein we extend the scope of the recently established RNA-TAG (transglycosylation at guanosine) methodology, a novel approach for genetically encoded site-specific labeling of large mRNA transcripts, by employing mod-mRNA as substrate. As a proof of concept, we covalently attached a fluorescent probe to mCherry encoding mod-mRNA transcripts bearing 5-methylcytidine and/or pseudouridine substitutions with high labeling efficiencies...
July 27, 2017: Molecular Pharmaceutics
https://www.readbyqxmd.com/read/28747322/ribonucleoprotein-purification-and-characterization-using-rna-mango
#14
Shyam Ss Panchapakesan, Matthew L Ferguson, Eric J Hayden, Xin Chen, Aaron A Hoskins, Peter J Unrau
The characterization of RNA-protein complexes (RNPs) is a difficult but increasingly important problem in modern biology. By combining the compact RNA Mango aptamer with a fluorogenic thiazole orange desthiobiotin (TO1-Dtb or TO3-Dtb) ligand, we have created an RNA tagging system that simplifies the purification and subsequent characterization of endogenous RNPs. Mango-tagged RNP complexes can be immobilized on a streptavidin solid support and recovered in their native state by the addition of free biotin. Furthermore, Mango-based RNP purification can be adapted to different scales of RNP isolation ranging from pull-down assays to the isolation of large amounts of biochemically defined cellular RNPs...
July 26, 2017: RNA
https://www.readbyqxmd.com/read/28745416/a-role-for-host-cell-exocytosis-in-inlb-mediated-internalization-of-listeria-monocytogenes
#15
Hoan Van Ngo, Manmeet Bhalla, Da-Yuan Chen, Keith Ireton
The bacterial surface protein InlB mediates internalization of Listeria monocytogenes into human cells through interaction with the host receptor tyrosine kinase, Met. InlB-mediated entry requires localized polymerization of the host actin cytoskeleton. Apart from actin polymerization, roles for other host processes in Listeria entry are unknown. Here we demonstrate that exocytosis in the human cell promotes InlB-dependent internalization. Using a probe consisting of VAMP3 with an exofacial Green Fluorescent Protein (GFP) tag, focal exocytosis was detected during InlB-mediated entry...
July 26, 2017: Cellular Microbiology
https://www.readbyqxmd.com/read/28743737/real-time-visualization-of-chromatin-modification-in-isolated-nuclei
#16
Luca Sardo, Angel Lin, Svetlana Khakhina, Lucas Beckman, Luis Ricon, Weam Elbezanti, Tara Jaison, Harshad Vishwasrao, Hari Shroff, Christopher Janetopoulos, Zachary A Klase
Chromatin modification is traditionally queried by biochemical assays that provide average measurements of static events since the analysis requires components from many cells. Microscopy can visualize single cells, but the cell body and organelles can hamper staining and visualization of the nucleus. Normally, chromatin is visualized by immunostaining a fixed sample or by expressing exogenous fluorescently tagged proteins in a live cell. Alternative microscopy tools to observe changes of endogenous chromatin in real-time are needed...
July 25, 2017: Journal of Cell Science
https://www.readbyqxmd.com/read/28743675/role-of-nmd-and-nas-in-the-mrna-pattern-of-two-new-%C3%AE-thalassemia-mutants
#17
Giovanna Cardiero, Clelia Scarano, Gennaro Musollino, Francesca Di Noce, Romeo Prezioso, Sabrina Dembech, Gaetana La Porta, Maria Grazia Bisconte, Rosario Colella, Giuseppina Lacerra
The α-thalassemia is a common disease due in prevalence to deletional mutants. We have identified two new α-thalassemia pointform mutants: α1 cod22 GGC > GGT Gly > Gly creating a 5' splicing sequence and: α1 cod23 GAG >TAG Glu > stop. We perform a qualitative and semi-quantitative analysis of the mRNA molecules, from blood of the carriers, to define the molecular mechanisms giving rise to thalassemia phenotype. In vitro analysis using minigenes and cycloheximide was performed to evaluate if the mutants are substrate of nonsense-mediated mRNA decay...
July 22, 2017: International Journal of Biochemistry & Cell Biology
https://www.readbyqxmd.com/read/28743433/scavengers-for-bacteria-rainbow-trout-have-two-functional-variants-of-marco-that-bind-to-gram-negative-and-positive-bacteria
#18
Sarah J Poynter, Andrea L Monjo, Gabriella Micheli, Stephanie J DeWitte-Orr
Class A scavenger receptors (SR-As) are a family of surface-expressed receptors who bind a wide range of polyanionic ligands including bacterial components and nucleic acids and play a role in innate immunity. Macrophage receptor with collagenous structure (MARCO) is a SR-A family member that has been studied in mammals largely for its role in binding bacteria. To date there is little information about SR-As in general and MARCO specifically in fish, particularly what ligands individual SR-A family members bind remains largely unknown...
July 22, 2017: Developmental and Comparative Immunology
https://www.readbyqxmd.com/read/28736700/profiling-and-characterization-of-a-longissimus-dorsi-muscle-microrna-dataset-from-an-f2-duroc%C3%A2-%C3%A3-%C3%A2-pietrain-pig-resource-population
#19
Kaitlyn R Daza, Juan P Steibel, Deborah Velez-Irizarry, Nancy E Raney, Ronald O Bates, Catherine W Ernst
To elucidate the effects of microRNA (miRNA) regulation in skeletal muscle of adult pigs, miRNA expression profiling was performed with RNA extracted from longissimus dorsi (LD) muscle samples from 174 F2 pigs (~ 5.5 months of age) from a Duroc × Pietrain resource population. Total RNA was extracted from LD samples, and libraries were sequenced on an Illumina HiSeq 2500 platform in 1 × 50 bp format. After processing, 232,826,977 total reads were aligned to the Sus scrofa reference genome (v10.2.79), with 74...
September 2017: Genomics Data
https://www.readbyqxmd.com/read/28736360/-s100a4-a-potential-therapeutic-target-on-bladder-cancer-stem-cells
#20
Chun-Yan Wang, Qing-Wen Nie, Xuan Zhou, Da-Xiong Huang, Wei-Qiang Xiao, Yong-Tong Zhu
OBJECTIVE: To observe the effect of S100A4 gene silencing mediated by small interfering RNA (siRNA) on the proliferation of bladder cancer stem cells (CSCs) and their capacity of xenograft tumor formation. METHODS: MB49 bladder cancer stem cells (MCSCs) were isolated and identified. The differentially expressed protein S100A4 was identified in MCSCs using isobaric tags for relative and absolute quantitation technology (iTRAQ). A siRNA targeting S100A4 was constructed and transfected into MCSCs, and its inhibitory effects on S100A4 expression in MCSCs were assessed with Western blotting and qPCR...
July 20, 2017: Nan Fang Yi Ke da Xue Xue Bao, Journal of Southern Medical University
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