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https://www.readbyqxmd.com/read/28812884/controlling-bdellovibrio-bacteriovorus-gene-expression-and-predation-using-synthetic-riboswitches
#1
Mohammed Dwidar, Yohei Yokobayashi
Bdellovibrio bacteriovorus is a predatory bacterium that feeds on Gram-negative bacteria including a wide range of pathogens and thus has potential applications as a biocontrol agent. Owing to its unique life cycle, however, there are limited tools that enable genetic manipulation of B. bacteriovorus. This work describes our first steps toward engineering the predatory bacterium for practical applications by developing basic genetic parts to control gene expression. Specifically, we evaluated four robust promoters that are active during the attack phase of B...
August 18, 2017: ACS Synthetic Biology
https://www.readbyqxmd.com/read/28803467/not-gate-genetic-circuits-to-control-gene-expression-in-cyanobacteria
#2
Arnaud Taton, Amy T Ma, Mizuho Ota, Susan Golden, James W Golden
To downregulate gene expression in cyanobacteria, we constructed NOT gate genetic circuits using orthogonal promoters and their cognate repressors regulated translationally by synthetic riboswitches. Four NOT gates were tested and characterized in five cyanobacterial strains using fluorescent reporter-gene assays. In comparison to alternative systems used to downregulate gene expression in cyanobacteria, these NOT gates performed well, reducing YFP reporter expression by 4 to 50-fold. We further evaluated these NOT gates by controlling the expression of the ftsZ gene, which encodes a prokaryotic tubulin homolog that is required for cell division and is essential for Synechococcus elongatus PCC 7942...
August 14, 2017: ACS Synthetic Biology
https://www.readbyqxmd.com/read/28798404/an-in-vivo-high-throughput-screening-for-riboswitch-ligands-using-a-reverse-reporter-gene-system
#3
Marion Kirchner, Kenji Schorpp, Kamyar Hadian, Sabine Schneider
Riboswitches are bacterial RNA elements that regulate gene expression in response to metabolite or ion abundance and are considered as potential drug targets. In recent years a number of methods to find non-natural riboswitch ligands have been described. Here we report a high-throughput in vivo screening system that allows identifying OFF-riboswitch modulators in a 384 well bioluminescence assay format. We use a reverse reporter gene setup in Bacillus subtilis, consisting of a primary screening assay, a secondary assay as well as counter assays to detect compounds in a library of 1,280 molecules that act on the guanine-responsive xpt riboswitch from B...
August 10, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28793208/investigation-of-the-melting-behavior-of-dna-three-way-junctions-in-the-closed-and-open-states
#4
Carolyn E Carr, Luis A Marky
Intramolecular three-way junctions are commonly found in both DNA and RNA. These structures are functionally relevant in ribozymes, riboswitches, rRNA, and during replication. In this work, we present a thermodynamic description of the unfolding of DNA intramolecular three-way junctions. We used a combination of spectroscopic and calorimetric techniques to investigate the folding/unfolding thermodynamics of two three-way junctions with a closed (Closed-J) or open (Open-J) junction and their appropriate control stem-loop motifs (GAAATT-Hp, CTATC-Hp, and Dumbbell)...
August 8, 2017: Biophysical Journal
https://www.readbyqxmd.com/read/28792118/rna-mediated-signal-perception-in-pathogenic-bacteria
#5
REVIEW
Dmitriy Ignatov, Jörgen Johansson
Bacterial pathogens encounter several different environments during an infection, many of them possibly being detrimental. In order to sense its surroundings and adjust the gene expression accordingly, different regulatory schemes are undertaken. With these, the bacterium appropriately can differentiate between various environmental cues to express the correct virulence factor at the appropriate time and place. An attractive regulator device is RNA, which has an outstanding ability to alter its structure in response to external stimuli, such as metabolite concentration or alterations in temperature, to control its downstream gene expression...
August 9, 2017: Wiley Interdisciplinary Reviews. RNA
https://www.readbyqxmd.com/read/28761163/the-basal-translation-rate-of-authentic-hiv-1-rna-is-regulated-by-5-utr-nt-pairings-at-junction-of-r-and-u5
#6
I Boeras, B Seufzer, S Brady, A Rendahl, X Heng, K Boris-Lawrie
The paradigm protein synthesis rate is regulated by structural complexity of the 5'untranslated region (UTR) derives from bacterial and other riboswitches. In-solution, HIV-1 5'UTR forms two interchangeable long-range nucleotide (nt) -pairings, one sequesters the gag start codon promoting dimerization while the other sequesters the dimer initiation signal preventing dimerization. While the effect of these nt-pairings on dimerization and packaging has been documented their effect on authentic HIV translation in cellulo has remained elusive until now...
July 31, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28754999/identification-of-an-operon-involved-in-fluoride-resistance-in-enterobacter-cloacae-frm
#7
Xiaoqing Liu, Jian Tian, Lihui Liu, Tao Zhu, Xiaoxia Yu, Xiaoyu Chu, Bin Yao, Ningfeng Wu, Yunliu Fan
Fluorine is ubiquitous and the most active non-metal element in nature. While many microorganisms have developed fluoride resistance as a result of the widespread and prolonged application of oral hygiene products, the mechanisms used by these organisms to overcome fluoride toxicity are incompletely understood. In this study, a fluoride-resistant strain, Enterobacter cloacae FRM, was identified which could grow well at a fluoride concentration of 4,000 mg/L. According to comparative genomics, transcriptome under fluoride stress, and sequence analyses of two fluoride-resistant fosmid clones, the genomic island GI3 was found to be important for fluoride resistance...
July 28, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28749656/development-of-artificial-riboswitches-for-monitoring-of-naringenin-in-vivo
#8
Sungho Jang, Sungyeon Jang, Yu Xiu, Taek Jin Kang, Sang-Hyeup Lee, Mattheos A G Koffas, Gyoo Yeol Jung
Microbial strains are considered promising hosts for production of flavonoids because of their rapid growth rate and suitability for large-scale manufacturing. However, productivity and titer of current recombinant strains still do not meet the requirements of industrial processes. Genetically encoded biosensors have been applied for high-throughput screening or dynamic regulation of biosynthetic pathways for enhancing the performance of microbial strains that produce valuable chemicals. Currently, few protein sensor-regulators for flavonoids exist...
August 10, 2017: ACS Synthetic Biology
https://www.readbyqxmd.com/read/28746845/cooperation-between-magnesium-and-metabolite-controls-collapse-of-the-sam-i-riboswitch
#9
Susmita Roy, José N Onuchic, Karissa Y Sanbonmatsu
The S-adenosylmethionine (SAM)-I riboswitch is a noncoding RNA that regulates the transcription termination process in response to metabolite (SAM) binding. The aptamer portion of the riboswitch may adopt an open or closed state depending on the presence of metabolite. Although the transition between the open and closed states is critical for the switching process, its atomistic details are not well understood. Using atomistic simulations, we calculate the effect of SAM and magnesium ions on the folding free energy landscape of the SAM-I riboswitch...
July 25, 2017: Biophysical Journal
https://www.readbyqxmd.com/read/28719589/an-excited-state-underlies-gene-regulation-of-a-transcriptional-riboswitch
#10
Bo Zhao, Sharon L Guffy, Benfeard Williams, Qi Zhang
Riboswitches control gene expression through ligand-dependent structural rearrangements of the sensing aptamer domain. However, we found that the Bacillus cereus fluoride riboswitch aptamer adopts identical tertiary structures in solution with and without ligand. Using chemical-exchange saturation transfer (CEST) NMR spectroscopy, we revealed that the structured ligand-free aptamer transiently accesses a low-populated (∼1%) and short-lived (∼3 ms) excited conformational state that unravels a conserved 'linchpin' base pair to signal transcription termination...
September 2017: Nature Chemical Biology
https://www.readbyqxmd.com/read/28718632/systematic-approaches-to-efficiently-produce-2-3-butanediol-in-a-marine-cyanobacterium
#11
Nicole E Nozzi, Anna E Case, Austin L Carroll, Shota Atsumi
Cyanobacteria have attracted significant interest as a platform for renewable production of fuel and feedstock chemicals from abundant atmospheric carbon dioxide by way of photosynthesis. While great strides have been made in developing this technology in freshwater cyanobacteria, logistical issues remain in scale-up. Use of the cyanobacterium Synechococcus sp. PCC 7002 (7002) as a chemical production chassis could address a number of these issues given the higher tolerance to salt, light, and heat as well as the fast growth rate of 7002 in comparison to traditional model cyanobacteria such as Synechococcus elongatus PCC 7942 and Synechocystis sp...
August 3, 2017: ACS Synthetic Biology
https://www.readbyqxmd.com/read/28701520/ligand-recognition-and-helical-stacking-formation-are-intimately-linked-in-the-sam-i-riboswitch-regulatory-mechanism
#12
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/28691825/small-molecule-recognition-triggers-secondary-and-tertiary-interactions-in-dna-folding-and-hammerhead-ribozyme-catalysis
#13
Jie Mao, Chris DeSantis, Dennis Bong
We have identified tris(2-aminoethyl)amine (tren)-derived scaffolds with two (t2M) or four (t4M) melamine rings that can target oligo T/U domains in DNA/RNA. Unstructured T-rich DNAs cooperatively fold with the tren derivatives to form hairpin-like structures. Both t2M and t4M act as functional switches in a family of hammerhead ribozymes deactivated by stem or loop replacement with a U-rich sequence. Catalysis of bond scission in these hammerhead ribozymes could be restored by putative t2M/t4M refolding of stem secondary structure or tertiary bridging interactions between loop and stem...
July 13, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28676696/ribo-attenuators-novel-elements-for-reliable-and-modular-riboswitch-engineering
#14
Thomas Folliard, Barbara Mertins, Harrison Steel, Thomas P Prescott, Thomas Newport, Christopher W Jones, George Wadhams, Travis Bayer, Judith P Armitage, Antonis Papachristodoulou, Lynn J Rothschild
Riboswitches are structural genetic regulatory elements that directly couple the sensing of small molecules to gene expression. They have considerable potential for applications throughout synthetic biology and bio-manufacturing as they are able to sense a wide range of small molecules and regulate gene expression in response. Despite over a decade of research they have yet to reach this considerable potential as they cannot yet be treated as modular components. This is due to several limitations including sensitivity to changes in genetic context, low tunability, and variability in performance...
July 4, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28661578/fluoro-carba-sugars-are-glycomimetic-activators-of-the-glms-ribozyme
#15
Günter Mayer, Daniel Matzner, Anna Schüller, Torben Seitz, Valentin Wittmann
The glmS ribozyme is a bacterial gene-regulating riboswitch that controls cell wall synthesis, depending on glucosamine-6-phosphate as a cofactor. Due to the presence of this ribozyme in several human pathogen bacteria (e.g. MRSA, VRSA), the glmS ribozyme represents an attractive target for the development of artificial cofactors. The substitution of the ring oxygen in carbohydrates by functionalized methylene groups leads to a new generation of glycomimetics that exploits distinct interaction possibilities with their target structure in biological systems...
June 29, 2017: Chemistry: a European Journal
https://www.readbyqxmd.com/read/28640851/ligand-mediated-and-tertiary-interactions-cooperatively-stabilize-the-p1-region-in-the-guanine-sensing-riboswitch
#16
Christian A Hanke, Holger Gohlke
Riboswitches are genetic regulatory elements that control gene expression depending on ligand binding. The guanine-sensing riboswitch (Gsw) binds ligands at a three-way junction formed by paired regions P1, P2, and P3. Loops L2 and L3 cap the P2 and P3 helices and form tertiary interactions. Part of P1 belongs to the switching sequence dictating the fate of the mRNA. Previous studies revealed an intricate relationship between ligand binding and presence of the tertiary interactions, and between ligand binding and influence on the P1 region...
2017: PloS One
https://www.readbyqxmd.com/read/28640671/regulation-of-a-muralytic-enzyme-encoding-gene-by-two-non-coding-rnas
#17
Renée J St-Onge, Marie A Elliot
Non-coding regulatory RNAs fine-tune gene expression post-transcriptionally. In the streptomycetes, rpfA - encoding a muralytic enzyme required for establishing and exiting dormancy - is flanked by non-coding regulatory RNA elements both upstream (riboswitch) and downstream [antisense small RNA (sRNA)]. In Streptomyces coelicolor, the upstream riboswitch decreases rpfA transcript abundance in response to the second messenger cyclic di-AMP, itself involved in cell wall metabolism and dormancy. There is, however, no obvious expression platform associated with this riboswitch and consequently, its mechanism of action is entirely unknown...
June 22, 2017: RNA Biology
https://www.readbyqxmd.com/read/28625696/comparative-visualization-of-the-rna-suboptimal-conformational-ensemble-in%C3%A2-vivo
#18
COMPARATIVE STUDY
Chanin T Woods, Lela Lackey, Benfeard Williams, Nikolay V Dokholyan, David Gotz, Alain Laederach
When a ribonucleic acid (RNA) molecule folds, it often does not adopt a single, well-defined conformation. The folding energy landscape of an RNA is highly dependent on its nucleotide sequence and molecular environment. Cellular molecules sometimes alter the energy landscape, thereby changing the ensemble of likely low-energy conformations. The effects of these energy landscape changes on the conformational ensemble are particularly challenging to visualize for large RNAs. We have created a robust approach for visualizing the conformational ensemble of RNAs that is well suited for in vitro versus in vivo comparisons...
July 25, 2017: Biophysical Journal
https://www.readbyqxmd.com/read/28621923/capture-and-release-of-trna-by-the-t-loop-receptor-in-the-function-of-the-t-box-riboswitch
#19
Xianyang Fang, Malgorzata Michnicka, Yikan Zhang, Yun-Xing Wang, Edward P Nikonowicz
In Gram-positive bacteria, the tRNA-dependent T-box riboswitch system regulates expression of amino acid biosynthetic and aminoacyl-tRNA synthetase genes through a transcription attenuation mechanism. Binding of uncharged tRNA "closes" the switch, allowing transcription read-through. Structural studies of the 100-nucleotide stem I domain reveal tRNA utilizes base pairing and stacking interactions to bind the stem, but little is known structurally about the 180-nucleotide riboswitch core (stem I, stem III, and antiterminator stem) in complex with tRNA or the mechanism of coupling of the intermolecular binding domains crucial to T-box function...
July 18, 2017: Biochemistry
https://www.readbyqxmd.com/read/28613837/artificial-off-riboswitches-that-downregulate-internal-ribosome-entry-without-hybridization-switches-in-a-eukaryotic-cell-free-translation-system
#20
Atsushi Ogawa, Hiroki Masuoka, Tsubasa Ota
We constructed novel artificial riboswitches that function in a eukaryotic translation system (wheat germ extract), by rationally implanting an in vitro-selected aptamer into the intergenic internal ribosome entry site (IRES) of Plautia stali intestine virus. These eukaryotic OFF-riboswitches (OFF-eRSs) ligand-dose-dependently downregulate IRES-mediated translation without hybridization switches, which typical riboswitches utilize for gene regulation. The hybridization-switch-free mechanism not only allows for easy design but also requires less energy for regulation, resulting in a higher switching efficiency than hybridization-switch-based OFF-eRSs provide...
June 14, 2017: ACS Synthetic Biology
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