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Neerupma Bhardwaj, Kirtimaan Syal, Dipankar Chatterji
ppGpp, an alarmone for stringent response, plays an important role in the reprogramming of the transcription complex at the time of stress. In Escherichia coli, ppGpp mediates its action by binding to at least two different sites on RNA polymerase (RNAP). One of the sites to which ppGpp binds to RNAP is at the β'-ω interface; however, the underlying molecular mechanism and the physiological relevance of ppGpp binding to this site remain unclear. In this study, we have performed UV cross-linking experiments using32 P azido-labeled ppGpp to probe its association with RNAP in the absence and presence of ω, and observed weaker binding of ppGpp to the RNAP without ω...
March 15, 2018: Genes to Cells: Devoted to Molecular & Cellular Mechanisms
Hwa Young Kim, Sang-Mi Yu, Sang Chul Jeong, Sang Sun Yoon, Young Taek Oh
The stringent response (SR), which is activated by accumulation of (p)ppGpp under conditions of growth-inhibiting stresses, plays an important role ongrowth and virulence in Vibrio cholerae . Herein, we carried out a genome-wide screen using transposon random mutagenesis to identify genes controlled by SR in (p)ppGpp-overproducing mutant strain. One of the identified SR target genes was the flaC encoding flagellin. Genetic studies using flaC and SR mutants demonstrated that FlaC involved in bacterial growth, toxin production, and normal flagellum function under conditions of high (p)ppGpp levels, suggesting FlaC plays an important role in SR-induced pathogenicityin V...
March 15, 2018: Journal of Microbiology and Biotechnology
Yong Zhang, Eva Zborníková, Dominik Rejman, Kenn Gerdes
The alarmone (p)ppGpp plays pivotal roles in basic bacterial stress responses by increasing tolerance of various nutritional limitations and chemical insults, including antibiotics. Despite intensive studies since (p)ppGpp was discovered over 4 decades ago, (p)ppGpp binding proteins have not been systematically identified in Escherichia coli We applied DRaCALA (<u>d</u>ifferential<u>ra</u>dial<u>c</u>apillary<u>a</u>ction of<u>l</u>igand<u>a</u>ssay) to identify (p)ppGpp-protein interactions...
March 6, 2018: MBio
Sophie E Irving, Rebecca M Corrigan
The stringent response is a conserved bacterial stress response mechanism that allows bacteria to respond to nutritional challenges. It is mediated by the alarmones pppGpp and ppGpp, nucleotides that are synthesized and hydrolyzed by members of the RSH superfamily. Whilst there are key differences in the binding targets for (p)ppGpp between Gram-negative and Gram-positive bacterial species, the transient accumulation of (p)ppGpp caused by nutritional stresses results in a global change in gene expression in all species...
March 2018: Microbiology
Bin Bin Chen, Meng Li Liu, Lei Zhan, Chun Mei Li, Chengzhi Huang
Highly selective and sensitive detection of guanosine 3'-diphosphate-5'-diphosphate (ppGpp), namely the stringent in plants or microorganisms responding to strict or extreme environmental conditions such as stress and starvation, which plays an important role in gene expression, rRNA and antibiotics production, regulations of virulence of bacteria and growth of plants, faces a great challenge owing to its extreme similarity to normal nucleotides. By modifying the surface groups of a facile two-step hydrothermal route prepared carbon dots (CDs) with terbium ions (Tb3+) in this contribution, a novel fluorescent probe with excellent properties such as highly physical and chemical stability, narrow emission and excitation wavelength-independent emission was prepared...
March 1, 2018: Analytical Chemistry
Vadim Molodtsov, Elena Sineva, Lu Zhang, Xuhui Huang, Michael Cashel, Sarah E Ades, Katsuhiko S Murakami
DksA and ppGpp are the central players in the stringent response and mediate a complete reprogramming of the transcriptome. A major component of the response is a reduction in ribosome synthesis, which is accomplished by the synergistic action of DksA and ppGpp bound to RNA polymerase (RNAP) inhibiting transcription of rRNAs. Here, we report the X-ray crystal structures of Escherichia coli RNAP in complex with DksA alone and with ppGpp. The structures show that DksA accesses the template strand at the active site and the downstream DNA binding site of RNAP simultaneously and reveal that binding of the allosteric effector ppGpp reshapes the RNAP-DksA complex...
February 20, 2018: Molecular Cell
Galina V Smirnova, Aleksey V Tyulenev, Nadezda G Muzyka, Oleg N Oktyabrsky
Amino acid starvation causes an RelA-dependent increase in the regulatory nucleotide (p)ppGpp that leads to pleiotropic changes in Escheria coli metabolism, but the role of (p)ppGpp in regulation of respiration remains unclear. Here we demonstrate that amino acid starvation is accompanied by sharp RelA-dependent inhibition of respiration. The sharp phase of inhibition is absent in relA mutants, and can be prevented by translation inhibitors chloramphenicol and tetracycline, which abolish accumulation of (p)ppGpp...
February 22, 2018: Research in Microbiology
Hwa Young Kim, Junhyeok Go, Kang-Mu Lee, Young Taek Oh, Sang Sun Yoon
The pathogen Vibrio cholerae is the causative agent of cholera. Emergence of antibiotic-resistant V. cholerae strains is increasing, but the underlying mechanisms remain unclear. Herein, we report that the stringent response regulator and stress alarmone guanosine tetra- and pentaphosphate ([p]ppGpp) significantly contributes to antibiotic tolerance in V. cholerae We found that N16961, a pandemic V. cholerae strain and its isogenic (p)ppGpp-overexpressing mutant ΔrelAΔspoT are both more antibiotic resistant than (p)ppGpp0 ( ΔrelAΔrelVΔspoT ) and ΔdksA mutants, which cannot produce or utilize (p)ppGpp, respectively...
February 23, 2018: Journal of Biological Chemistry
Etienne Maisonneuve, Manuela Castro-Camargo, Kenn Gerdes
No abstract text is available yet for this article.
February 22, 2018: Cell
Kaneyoshi Yamamoto, Yuki Yamanaka, Tomohiro Shimada, Paramita Sarkar, Myu Yoshida, Neerupma Bhardwaj, Hiroki Watanabe, Yuki Taira, Dipankar Chatterji, Akira Ishihama
The RNA polymerase (RNAP) of Escherichia coli K-12 is a complex enzyme consisting of the core enzyme with the subunit structure α2 ββ'ω and one of the σ subunits with promoter recognition properties. The smallest subunit, omega (the rpoZ gene product), participates in subunit assembly by supporting the folding of the largest subunit, β', but its functional role remains unsolved except for its involvement in ppGpp binding and stringent response. As an initial approach for elucidation of its functional role, we performed in this study ChIP-chip (chromatin immunoprecipitation with microarray technology) analysis of wild-type and rpoZ -defective mutant strains...
January 2018: MSystems
Shanshan Zhang, Shuang Liu, Nan Wu, Youhua Yuan, Wenhong Zhang, Ying Zhang
As dormant phenotypic variants of bacteria, persisters account for many chronic infections affecting human health. Despite numerous studies, the role of small non-coding RNA (sRNA) in bacterial persistence has not been reported. To investigate the role of Hfq-interacting sRNA in persistence, we constructed the deletion mutants of 20 Hfq-interacting sRNAs (RyhB, GcvB, MgrR, RybB, MicF, SgrS, RprA, DicF, SsrS, FnrS, GadY, DsrA, OmrB, ArcZ, RyeB, RydC, OmrA, MicA, MicC, and ChiX) to assess their persistence capacity in uropathogenic Escherichia coli strain UTI89 and identified a new sRNA RyhB being involved in persister formation...
2018: Frontiers in Microbiology
Sousuke Imamura, Yuhta Nomura, Tokiaki Takemura, Imran Pancha, Keiko Taki, Kazuki Toguchi, Yuzuru Tozawa, Kan Tanaka
Chloroplasts are plant organelles that carry out oxygenic photosynthesis. Chloroplast biogenesis depends upon chloroplast ribosomes and their translational activity. However, regulation of chloroplast ribosome biogenesis remains an important unanswered question. In this study, we found that inhibition of target of rapamycin (TOR), a general eukaryotic checkpoint kinase, results in a decline in chloroplast ribosomal RNA (rRNA) transcription in the unicellular red alga, Cyanidioschyzon merolae. Upon TOR inhibition, transcriptomics and other analyses revealed increased expression of a nuclear-encoded chloroplast RelA-SpoT homolog (RSH) gene (CmRSH4b), which encodes a homolog of the guanosine 3'-diphosphate 5'-diphosphate (ppGpp) synthetases that modulate rRNA synthesis in bacteria...
February 14, 2018: Plant Journal: for Cell and Molecular Biology
A E Stella, D Luz, R M F Piazza, B Spira
No abstract text is available yet for this article.
February 6, 2018: Epidemiology and Infection
Wieland Steinchen, Marian S Vogt, Florian Altegoer, Pietro I Giammarinaro, Petra Horvatek, Christiane Wolz, Gert Bange
The nutritional alarmones ppGpp and pppGpp (collectively: (p)ppGpp) are nucleotide-based second messengers enabling bacteria to respond to environmental and stress conditions. Several bacterial species contain two highly homologous (p)ppGpp synthetases named RelP (SAS2, YwaC) and RelQ (SAS1, YjbM). It is established that RelQ forms homotetramers that are subject to positive allosteric regulation by pppGpp, but structural and mechanistic insights into RelP lack behind. Here we present a structural and mechanistic characterization of RelP...
February 1, 2018: Scientific Reports
Pavel Kudrin, Ievgen Dzhygyr, Kensuke Ishiguro, Jelena Beljantseva, Elena Maksimova, Sofia Raquel Alves Oliveira, Vallo Varik, Roshani Payoe, Andrey L Konevega, Tanel Tenson, Tsutomu Suzuki, Vasili Hauryliuk
During amino acid starvation the Escherichia coli stringent response factor RelA recognizes deacylated tRNA in the ribosomal A-site. This interaction activates RelA-mediated synthesis of alarmone nucleotides pppGpp and ppGpp, collectively referred to as (p)ppGpp. These two alarmones are synthesized by addition of a pyrophosphate moiety to the 3' position of the abundant cellular nucleotide GTP and less abundant nucleotide GDP, respectively. Using untagged native RelA we show that allosteric activation of RelA by pppGpp increases the efficiency of GDP conversion to achieve the maximum rate of (p)ppGpp production...
January 30, 2018: Nucleic Acids Research
Nanjing Ji, Lingxiao Lin, Ling Li, Liying Yu, Yaqun Zhang, Hao Luo, Meizhen Li, Xinguo Shi, Da-Zhi Wang, Senjie Lin
Despite numerous laboratory studies on physiologies of harmful algal bloom (HAB) species, physiologies of these algae during a natural bloom are understudied. Here, we investigated a bloom of the raphidophyte Heterosigma akashiwo in the East China Sea in 2014 using metabarcode (18S rDNA) and metatranscriptome sequencing. Based on 18S rDNA analyses, the phytoplankton community shifted from high diversity in the pre-bloom stage to H. akashiwo predominance during the bloom. A sharp decrease in ambient dissolved inorganic phosphate and strong up-regulation of phosphate and dissolved organic phosphorus (DOP) uptake genes, including the rarely documented (ppGpp)ase, in H...
January 18, 2018: Environmental Microbiology
Farkhondeh Poursina, Jamshid Fagri, Nasrin Mirzaei, Hajieh Ghasemian Safaei
Helicobacter pylori (H. pylori) can convert to coccoid form in unfavorable conditions or as a result of antibiotic treatment. In order to adapt to harsh environments, H. pylori requires a stringent response which, encoded by the spoT gene, has a bifunctional enzyme possessing both (p)ppGpp synthetic and degrading activity. Our goal in this study was to compare spoT gene expression in spiral and induced coccoid forms of H. pylori with use of amoxicillin. First, clinical isolate coccoid forms were induced with amoxicillin; then, the viability test was analyzed by flow cytometer...
January 11, 2018: Folia Microbiologica
Melek Cemre Manav, Jelena Beljantseva, Martin Saxtorph Bojer, Tanel Tenson, Hanne Ingmer, Vasili Hauryliuk, Ditlev Egeskov Brodersen
The stringent response is a global reprogramming of bacterial physiology that renders cells more tolerant to antibiotics and induces virulence gene expression in pathogens in response to stress. This process is driven by accumulation of the intracellular alarmone guanosine-5'-di(tri)phosphate-3'-diphosphate ([p]ppGpp), which is produced by enzymes of the RelA SpoT homologue (RSH) family. The Gram-positive Firmicute pathogen, Staphylococcus aureus, encodes three RSH enzymes: a multi-domain RSH (Rel) that senses amino acid starvation on the ribosome and two small alarmone synthetase (SAS) enzymes, RelQ (SAS1) and RelP (SAS2)...
January 11, 2018: Journal of Biological Chemistry
Priyanka Agrawal, Rajagopal Varada, Shivjee Sah, Souvik Bhattacharyya, Umesh Varshney
Bacteria respond to stressful growth conditions through a conserved phenomenon of 'stringent response' mediated by synthesis of stress alarmones, ppGpp or pppGpp [referred to as (p)ppGpp]. The (p)ppGpp synthesis is known to occur by ribosome associated RelA. In addition, a dual function protein SpoT (with both the synthetase and hydrolase activities) maintains (p)ppGpp homeostasis. The presence of (p)ppGpp is also known to contribute to antibiotic resistance in bacteria. Mycobacterium smegmatis possesses Arr, which inactivates rifampicin by its ADP-ribosylation...
January 8, 2018: Journal of Bacteriology
Ben Field
The hyperphosphorylated nucleotides guanosine pentaphosphate and tetraphosphate (together referred to as (p)ppGpp, or magic spot) orchestrate a signalling cascade in bacteria that controls growth under optimal conditions and in response to environmental stress. (p)ppGpp is also found in the chloroplasts of plants and algae where it is has also been shown to accumulate in response to abiotic stress. Recent studies suggest that (p)ppGpp is a potent inhibitor of chloroplast gene expression in vivo, and is a significant regulator of chloroplast function that can influence both the growth and the development of plants...
December 21, 2017: Journal of Experimental Botany
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