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Wendy W K Mok, Mark P Brynildsen
In this issue, Nelson and colleagues (2017) determined that guanidine, the prevalent protein denaturant, is the long-lost ligand sensed by the ykkC class of riboswitches, and identified that members of its regulon are involved in guanidine detoxification and export.
January 19, 2017: Molecular Cell
Robert A Battaglia, Ian R Price, Ailong Ke
Regulation of gene expression by cis-encoded riboswitches is a prevalent theme in bacteria. Of the hundreds of riboswitch families identified, the majority of them remain as orphans, without a clear ligand assignment. The ykkC orphan family was recently characterized as guanidine-sensing riboswitches. Herein we present a 2.04 Å crystal structure of the guanidine-bound ykkC riboswitch from Dickeya dadantii. The riboswitch folds into a boot-shaped structure, with a co-axially stacked P1/P2 stem forming the boot, and a 3'-P3 stem-loop forming the heel...
January 17, 2017: RNA
Ely B Porter, Jacob T Polaski, Makenna M Morck, Robert T Batey
Allosteric RNA devices are increasingly being viewed as important tools capable of monitoring enzyme evolution, optimizing engineered metabolic pathways, facilitating gene discovery and regulators of nucleic acid-based therapeutics. A key bottleneck in the development of these platforms is the availability of small-molecule-binding RNA aptamers that robustly function in the cellular environment. Although aptamers can be raised against nearly any desired target through in vitro selection, many cannot easily be integrated into devices or do not reliably function in a cellular context...
January 16, 2017: Nature Chemical Biology
Irina A Rodionova, Matthew W Vetting, Xiaoqing Li, Steven C Almo, Andrei L Osterman, Dmitry A Rodionov
Riboflavin (vitamin B2) is the precursor of flavin mononucleotide (FMN) and flavin adenine dinucleotide, which are essential coenzymes in all free-living organisms. Riboflavin biosynthesis in many Bacteria but not in Archaea is controlled by FMN-responsive riboswitches. We identified a novel bifunctional riboflavin kinase/regulator (RbkR), which controls riboflavin biosynthesis and transport genes in major lineages of Crenarchaeota, Euryarchaeota and Thaumarchaeota. RbkR proteins are composed of the riboflavin kinase domain and a DNA-binding winged helix-turn-helix-like domain...
January 9, 2017: Nucleic Acids Research
Adrien Chauvier, Frédéric Picard-Jean, Jean-Christophe Berger-Dancause, Laurène Bastet, Mohammad Reza Naghdi, Audrey Dubé, Pierre Turcotte, Jonathan Perreault, Daniel A Lafontaine
On the basis of nascent transcript sequencing, it has been postulated but never demonstrated that transcriptional pausing at translation start sites is important for gene regulation. Here we show that the Escherichia coli thiamin pyrophosphate (TPP) thiC riboswitch contains a regulatory pause site in the translation initiation region that acts as a checkpoint for thiC expression. By biochemically probing nascent transcription complexes halted at defined positions, we find a narrow transcriptional window for metabolite binding, in which the downstream boundary is delimited by the checkpoint...
January 10, 2017: Nature Communications
Erik R Van Vlack, Shana Topp, Jessica C Seeliger
: We report here the behavior of naturally occurring and rationally engineered preQ1 riboswitches and their application to inducible gene regulation in mycobacteria. Because mycobacteria lack preQ1 biosynthetic genes, we hypothesized that preQ1 could be used as an exogenous non-metabolite ligand to control riboswitches in mycobacteria. Selected naturally occurring preQ1 riboswitches were assayed and successfully drove preQ1-dependent repression of a GFP reporter in Mycobacterium smegmatis Using structure-based design, we engineered three preQ1 riboswitches from Thermoanaerobacter tencongensis, Bacillus subtilis, and Lactobacillus rhamnosus towards achieving higher response ratios and increased repression...
January 9, 2017: Journal of Bacteriology
Stefanie Sandra Krajewski, Isabelle Isoz, Jörgen Johansson
The emerging development of antibiotic resistant bacteria calls for novel types of antibacterial agents. In this work we examined the putative antibacterial effect of purine analogs in Listeria monocytogenes We show that, among several tested purine analogs, only 6-N-hydroxylaminopurine (6-N-HAP) reduces the viability of the Gram-positive pathogen Listeria monocytogenes As in Bacillus subtilis, 6-N-HAP terminates expression at guanine riboswitches in L. monocytogenes hence preventing expression of their downstream genes...
January 6, 2017: Nucleic Acids Research
Ronald R Breaker, Ruben M Atilho, Sarah N Malkowski, James W Nelson, Madeline E Sherlock
No abstract text is available yet for this article.
January 6, 2017: Biochemistry
Elnaz Mehdizadeh Aghdam, Malte Sinn, Vahideh Tarhriz, Abolfazl Barzegar, Jörg S Hartig, Mohammad Saeid Hejazi
Riboswitches are located in non-coding areas of mRNAs and act as sensors of cellular small molecules, regulating gene expression in response to ligand binding. The TPP riboswitch is the most widespread riboswitch occurring in all three domains of life. However, it has been rarely characterized in environmental bacteria other than Escherichia coli and Bacillus subtilis. In this study, TPP riboswitches located in the 5' UTR of thiC operon from Alishewanella tabrizica and Alishewanella aestuarii were identified and characterized...
January 2017: Microbiological Research
Caroline W Reiss, Yong Xiong, Scott A Strobel
The guanidine-I riboswitch is a conserved RNA element with approximately 2,000 known examples across four phyla of bacteria. It exists upstream of nitrogen metabolism and multidrug resistance transporter genes and alters expression through the specific recognition of a free guanidinium cation. Here we report the structure of a guanidine riboswitch aptamer from Sulfobacillus acidophilus at 2.7 Å resolution. Helices P1, P1a, P1b, and P2 form a coaxial stack that acts as a scaffold for ligand binding. A previously unidentified P3 helix docks into P1a to form the guanidinium binding pocket, which is completely enclosed...
December 18, 2016: Structure
Savannah Colameco, Marie A Elliot
Antibiotics inhibit a wide range of essential processes in the bacterial cell, including replication, transcription, translation and cell wall synthesis. In many instances, these antibiotics exert their effects through association with non-coding RNAs. This review highlights many classical antibiotic targets (e.g. rRNAs and the ribosome), explores a number of emerging targets (e.g. tRNAs, RNase P, riboswitches and small RNAs), and discusses the future directions and challenges associated with non-coding RNAs as antibiotic targets...
December 22, 2016: Biochemical Pharmacology
Johanna Roßmanith, Franz Narberhaus
Due to their simple architecture and control mechanism, regulatory RNA modules are attractive building blocks in synthetic biology. This is especially true for riboswitches, which are natural ligand-binding regulators of gene expression. The discovery of various tandem riboswitches inspired the design of combined RNA modules with activities not yet found in nature. Riboswitches were placed in tandem or in combination with a ribozyme or temperature-responsive RNA thermometer resulting in new functionalities...
December 23, 2016: RNA Biology
Madeline E Sherlock, Ronald R Breaker
Recently, it was determined that representatives of the riboswitch candidates called ykkC and mini-ykkC directly bind free guanidine. These riboswitches regulate the expression of genes whose protein products are implicated in overcoming the toxic effects of this ligand. Thus, the relevant ykkC motif and mini-ykkC motif RNAs have been classified as guanidine-I and guanidine-II riboswitch RNAs, respectively. Moreover, we had previously noted that a third candidate riboswitch class, called ykkC-III, was associated with a distribution of genes similar to those of the other two motifs...
January 6, 2017: Biochemistry
Madeline E Sherlock, Sarah N Malkowski, Ronald R Breaker
Recently, it was determined that representatives of the riboswitch candidates called ykkC and ykkC-III directly bind free guanidine. Guanidine-binding ykkC motif RNAs, now renamed guanidine-I riboswitches, were demonstrated to commonly regulate the expression of genes encoding guanidine carboxylases, as well as others encoding guanidine efflux proteins such as EmrE and SugE. Likewise, genes encoding similar efflux proteins are associated with ykkC-III motif RNAs, which have now been renamed guanidine-III riboswitches...
January 6, 2017: Biochemistry
Gesine Domin, Sven Findeiß, Manja Wachsmuth, Sebastian Will, Peter F Stadler, Mario Mörl
Riboswitches have gained attention as tools for synthetic biology, since they enable researchers to reprogram cells to sense and respond to exogenous molecules. In vitro evolutionary approaches produced numerous RNA aptamers that bind such small ligands, but their conversion into functional riboswitches remains difficult. We previously developed a computational approach for the design of synthetic theophylline riboswitches based on secondary structure prediction. These riboswitches have been constructed to regulate ligand-dependent transcription termination in Escherichia coli Here, we test the usability of this design strategy by applying the approach to tetracycline and streptomycin aptamers...
December 19, 2016: Nucleic Acids Research
James W Nelson, Ruben M Atilho, Madeline E Sherlock, Randy B Stockbridge, Ronald R Breaker
The guanidyl moiety is a component of fundamental metabolites, including the amino acid arginine, the energy carrier creatine, and the nucleobase guanine. Curiously, reports regarding the importance of free guanidine in biology are sparse, and no biological receptors that specifically recognize this compound have been previously identified. We report that many members of the ykkC motif RNA, the longest unresolved riboswitch candidate, naturally sense and respond to guanidine. This RNA is found throughout much of the bacterial domain of life, where it commonly controls the expression of proteins annotated as urea carboxylases and multidrug efflux pumps...
January 19, 2017: Molecular Cell
Wenjing Cui, Jintao Cheng, Shengnan Miao, Li Zhou, Zhongmei Liu, Junling Guo, Zhemin Zhou
Tuneable gene expression controlled by synthetic biological elements is of great importance to biotechnology and synthetic biology. The synthetic riboswitch is a pivotal type of elements that can easily control the heterologous gene expression in diverse bacteria. In this study, the theophylline-dependent synthetic riboswitch and the corresponding variants with varied spacings between Shine-Dalgarno (SD) sequence and start codon were employed to comprehensively characterize the induction and regulation properties through combining a strong promoter aprE in Bacillus subtilis...
December 16, 2016: Applied Microbiology and Biotechnology
Magali Boutard, Laurence Ettwiller, Tristan Cerisy, Adriana Alberti, Karine Labadie, Marcel Salanoubat, Ira Schildkraut, Andrew C Tolonen
Bacteria respond to their environment by regulating mRNA synthesis, often by altering the genomic sites at which RNA polymerase initiates transcription. Here, we investigate genome-wide changes in transcription start site (TSS) usage by Clostridium phytofermentans, a model bacterium for fermentation of lignocellulosic biomass. We quantify expression of nearly 10,000 TSS at single base resolution by Capp-Switch sequencing, which combines capture of synthetically capped 5' mRNA fragments with template-switching reverse transcription...
December 16, 2016: Nature Communications
Atsushi Ogawa, Yuta Murashige, Junichiro Tabuchi, Taiki Omatsu
We have rationally constructed a novel regulation-type of artificial riboswitch that ligand-dose dependently upregulates translation initiation mediated by a 3' cap-independent translation element (3' CITE) with no major hybridization switches in a plant expression system (wheat germ extract).
December 15, 2016: Molecular BioSystems
Suzanne G Rzuczek, Lesley A Colgan, Yoshio Nakai, Michael D Cameron, Denis Furling, Ryohei Yasuda, Matthew D Disney
Excluding the ribosome and riboswitches, developing small molecules that selectively target RNA is a longstanding problem in chemical biology. A typical cellular RNA is difficult to target because it has little tertiary, but abundant secondary structure. We designed allele-selective compounds that target such an RNA, the toxic noncoding repeat expansion (r(CUG)(exp)) that causes myotonic dystrophy type 1 (DM1). We developed several strategies to generate allele-selective small molecules, including non-covalent binding, covalent binding, cleavage and on-site probe synthesis...
February 2017: Nature Chemical Biology
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