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Thao Nguyen Le Lam, Claire Morvan, Wenfeng Liu, Chantal Bohn, Yan Jaszczyszyn, Philippe Bouloc
Bacteria optimize their fitness in response to a changing environment by tight regulation of gene expression. Regulation can be controlled at both transcriptional and post-transcriptional levels via key players such as sigma factors, regulatory proteins and regulatory RNAs. The identification of phenotypes associated with gene deletions is the established method for finding gene functions but may require testing many conditions for each studied mutant. As regulatory RNAs often contribute to fine-tuning gene expression, phenotypes associated with their inactivation are often weak and difficult to detect...
December 1, 2016: Methods: a Companion to Methods in Enzymology
Jia Wen, John R Harp, Elizabeth M Fozo
Many bacterial type I toxin mRNAs possess a long 5' untranslated region (UTR) that serves as the target site of the corresponding antitoxin sRNA. This is the case for the zorO-orzO type I system where the OrzO antitoxin base pairs to the 174-nucleotide zorO 5' UTR. Here, we demonstrate that the full-length 5' UTR of the zorO type I toxin hinders its own translation independent of the sRNA whereas a processed 5' UTR (zorO Δ28) promotes translation. The full-length zorO 5' UTR folds into an extensive secondary structure sequestering the ribosome binding site (RBS)...
November 29, 2016: Nucleic Acids Research
Agata Groszewska, Zuzanna Wroblewska, Mikolaj Olejniczak
Small bacterial RNAs (sRNAs) regulate translation by pairing with complementary sequences in their target mRNAs, in a process which is often dependent on the Hfq protein. Here, the secondary structure of a 95-nt long fragment of Salmonella fadL mRNA containing RybB sRNA binding site in the coding region was analyzed. The data indicated local rearrangements in this mRNA structure after the annealing of RybB. The filter retention data had shown that Hfq bound both RybB and the fadL mRNA fragment with tight affinities...
November 25, 2016: Acta Biochimica Polonica
Hannah Gaimster, Lisa Chalklen, Mark Alston, John T Munnoch, David J Richardson, Andrew J Gates, Gary Rowley
Nitrous oxide (N2O) is a stable, ozone depleting greenhouse gas. Emissions of N2O into the atmosphere continue to rise, primarily due to the use of nitrogen-containing fertilizers by soil denitrifying microbes. It is clear more effective mitigation strategies are required to reduce emissions. One way to help develop future mitigation strategies is to address the currently poor understanding of transcriptional regulation of the enzymes used to produce and consume N2O. With this ultimate aim in mind we performed RNA-seq on a model soil denitrifier, Paracoccus denitrificans, cultured anaerobically under high N2O and low N2O emitting conditions, and aerobically under zero N2O emitting conditions to identify small RNAs (sRNAs) with potential regulatory functions transcribed under these conditions...
2016: Frontiers in Microbiology
Sabrina Shore, Jordana M Henderson, Alexandre Lebedev, Michelle P Salcedo, Gerald Zon, Anton P McCaffrey, Natasha Paul, Richard I Hogrefe
For most sample types, the automation of RNA and DNA sample preparation workflows enables high throughput next-generation sequencing (NGS) library preparation. Greater adoption of small RNA (sRNA) sequencing has been hindered by high sample input requirements and inherent ligation side products formed during library preparation. These side products, known as adapter dimer, are very similar in size to the tagged library. Most sRNA library preparation strategies thus employ a gel purification step to isolate tagged library from adapter dimer contaminants...
2016: PloS One
Sarah Maria Hücker, Svenja Simon, Siegfried Scherer, Klaus Neuhaus
The enteric pathogen Escherichia coli O157:H7 Sakai (EHEC) is able to grow at lower temperatures compared to commensal E. coli Growth at environmental conditions displays complex challenges different to those in a host. EHEC was grown at 37°C (control) and at 14°C with 4% NaCl, a combination of cold and osmotic stress as present in the food chain. Comparison of RNAseq and RIBOseq data provided a snap shot of ongoing transcription and translation, differentiating transcriptional and post-transcriptional gene regulation, respectively...
November 16, 2016: FEMS Microbiology Letters
Jiandong Chen, Susan Gottesman
: The L-arabinose-inducible araBAD promoter (PBAD) allows tightly controlled and tunable expression of genes of interest in a broad range of bacterial species. It has been successfully used to study bacterial sRNA regulation, where PBAD drives expression of target mRNA translational fusions. Here we report that in Escherichia coli, Spot 42 sRNA can regulate PBAD promoter activity by affecting arabinose uptake. We demonstrate that Spot 42 sRNA represses araF, a gene encoding the AraF subunit of the high-affinity low-capacity arabinose transporter AraFGH, through direct base pairing interactions...
November 14, 2016: Journal of Bacteriology
Ajay Kumar Mishra, Ganesh Selvaraj Duraisamy, Jaroslav Matoušek, Sebastjan Radisek, Branka Javornik, Jernej Jakse
BACKGROUND: Hop (Humulus lupulus L.) plants are grown primarily for the brewing industry and have been used as a traditional medicinal herb for a long time. Severe hop stunt disease caused by the recently discovered Citrus bark cracking viroid (CBCVd) is one of the most devastating diseases among other viroid infections in hop. MicroRNAs (miRNAs) are a class of non-coding small RNAs that play important roles in gene expression regulation. To identify miRNAs in hop and their response to CBCVd-infection, two small RNA (sRNA) libraries were prepared from healthy and CBCVd-infected hop plants and were investigated by high throughput sequencing...
November 15, 2016: BMC Genomics
Veronica Ancona, Jae Hoon Lee, Youfu Zhao
The GacS/GacA two-component system (also called GrrS/GrrA) is a global regulatory system which is highly conserved among gamma-proteobacteria. This system positively regulates non-coding small regulatory RNA csrB, which in turn binds to the RNA-binding protein CsrA. However, how GacS/GacA-Csr system regulates virulence traits in E. amylovora remains unknown. Results from mutant characterization showed that the csrB mutant was hypermotile, produced higher amount of exopolysaccharide amylovoran, and had increased expression of type III secretion (T3SS) genes in vitro...
November 15, 2016: Scientific Reports
Shafagh A Waters, Sean P McAteer, Grzegorz Kudla, Ignatius Pang, Nandan P Deshpande, Timothy G Amos, Kai Wen Leong, Marc R Wilkins, Richard Strugnell, David L Gally, David Tollervey, Jai J Tree
RNA sequencing studies have identified hundreds of non-coding RNAs in bacteria, including regulatory small RNA (sRNA). However, our understanding of sRNA function has lagged behind their identification due to a lack of tools for the high-throughput analysis of RNA-RNA interactions in bacteria. Here we demonstrate that in vivo sRNA-mRNA duplexes can be recovered using UV-crosslinking, ligation and sequencing of hybrids (CLASH). Many sRNAs recruit the endoribonuclease, RNase E, to facilitate processing of mRNAs...
November 11, 2016: EMBO Journal
Yi Zheng, Shan Gao, Chellappan Padmanabhan, Rugang Li, Marco Galvez, Dina Gutierrez, Segundo Fuentes, Kai-Shu Ling, Jan Kreuze, Zhangjun Fei
Accurate detection of viruses in plants and animals is critical for agriculture production and human health. Deep sequencing and assembly of virus-derived small interfering RNAs has proven to be a highly efficient approach for virus discovery. Here we present VirusDetect, a bioinformatics pipeline that can efficiently analyze large-scale small RNA (sRNA) datasets for both known and novel virus identification. VirusDetect performs both reference-guided assemblies through aligning sRNA sequences to a curated virus reference database and de novo assemblies of sRNA sequences with automated parameter optimization and the option of host sRNA subtraction...
November 5, 2016: Virology
Divya Balasubramanian, Preethi T Ragunathan, Jingyi Fei, Carin K Vanderpool
Hundreds of small RNAs (sRNAs) have been identified in diverse bacterial species, and while the functions of most remain unknown, some regulate key processes, particularly stress responses. The sRNA DicF was identified over 25 years ago as an inhibitor of cell division but since then has remained uncharacterized. DicF consists of 53 nucleotides and is encoded by a gene carried on a prophage (Qin) in the genomes of many Escherichia coli strains. We demonstrated that DicF inhibits cell division via direct base pairing with ftsZ mRNA to repress translation and prevent new synthesis of the bacterial tubulin homolog FtsZ...
January 2016: MSystems
Yuyan Sun, Yang Qiu, Mengmeng Duan, Jinglei Wang, Xiaohui Zhang, Haiping Wang, Jiangping Song, Xixiang Li
Anthocyanins are widely distributed water-soluble phytochemical pigments belonging to the flavonoid group. To date, limited knowledge is available about the regulatory roles of miRNAs in anthocyanin biosynthesis in plants. To identify the miRNAs associated with anthocyanin biosynthesis in radish, five small RNA (sRNA) libraries constructed from 'Xinlimei' radish roots at 11, 21, 44, 56 and 73 days (d) were examined using high-throughput sequencing technology. A total of 102.02 million (M) clean reads were generated, from which 483 known and 1415 novel miRNAs were identified...
November 5, 2016: Molecular Genetics and Genomics: MGG
Ibukun A Akinyemi, Fang Wang, Benguo Zhou, Shuishui Qi, Qingfa Wu
BACKGROUND: The invasion of plant by viruses cause major damage to plants and reduces crop yield and integrity. Devastating plant virus infection has been experienced at different times all over the world, which are attributed to different events of mutation, re-assortment and recombination occurring in the viruses. Strategies for proper virus management has been mostly limited to eradicating the vectors that spreads the plant viruses. However, development of prompt and effective diagnostic methods are required to monitor emerging and re-emerging diseases that may be symptomatic or asymptomatic in the plant as well as the genetic variation and evolution in the plant viruses...
November 4, 2016: Virology Journal
Lennart Weber, Clemens Thoelken, Marcel Volk, Bernhard Remes, Marcus Lechner, Gabriele Klug
Cell division and cell wall synthesis mechanisms are similarly conserved among bacteria. Consequently some bacterial species have comparable sets of genes organized in the dcw (division and cell wall) gene cluster. Dcw genes, their regulation and their relative order within the cluster are outstandingly conserved among rod shaped and gram negative bacteria to ensure an efficient coordination of growth and division. A well studied representative is the dcw gene cluster of E. coli. The first promoter of the gene cluster (mraZ1p) gives rise to polycistronic transcripts containing a 38 nt long 5' UTR followed by the first gene mraZ...
2016: PloS One
Franciele Maboni Siqueira, Guilherme Loss de Morais, Susan Higashi, Laura Scherer Beier, Gabriela Merker Breyer, Caio Padoan de Sá Godinho, Marie-France Sagot, Irene Silveira Schrank, Arnaldo Zaha, Ana Tereza Ribeiro de Vasconcelos
BACKGROUND: Bacterial non-coding RNAs act by base-pairing as regulatory elements in crucial biological processes. We performed the identification of trans-encoded small RNAs (sRNA) from the genomes of Mycoplama hyopneumoniae, Mycoplasma flocculare and Mycoplasma hyorhinis, which are Mycoplasma species that have been identified in the porcine respiratory system. RESULTS: A total of 47, 15 and 11 putative sRNAs were predicted in M. hyopneumoniae, M. flocculare and M...
October 25, 2016: BMC Genomics
Ian K Greaves, Steven R Eichten, Michael Groszmann, Aihua Wang, Hua Ying, W James Peacock, Elizabeth S Dennis
Hybrid Arabidopsis plants undergo epigenetic reprogramming producing decreased levels of 24-nt siRNAs and altered patterns of DNA methylation that can affect gene expression. Driving the changes in methylation are the processes trans-chromosomal methylation (TCM) and trans-chromosomal demethylation (TCdM). In TCM/TCdM the methylation state of one allele is altered to resemble the other allele. We show that Pol IV-dependent sRNAs are required to establish TCM events. The changes in DNA methylation and the associated changes in sRNA levels in the F1 hybrid can be maintained in subsequent generations and affect hundreds of regions in the F2 epigenome...
October 17, 2016: Proceedings of the National Academy of Sciences of the United States of America
Nhan Tu, Ronan K Carroll, Andy Weiss, Lindsey N Shaw, Gael Nicolas, Sarah Thomas, Amorce Lima, Udoka Okaro, Burt Anderson
Bartonella henselae is a gram-negative zoonotic bacterium that causes infections in humans including endocarditis and bacillary angiomatosis. B. henselae has been shown to grow as large aggregates and form biofilms in vitro. The aggregative growth and the angiogenic host response requires the trimeric autotransporter adhesin BadA. We examined the transcriptome of the Houston-1 strain of B. henselae using RNA-seq revealing nine novel, highly-expressed intergenic transcripts (Bartonella regulatory transcript, Brt1-9)...
October 28, 2016: MicrobiologyOpen
Chao Wu, Xinyue Li, Song Guo, Sek-Man Wong
In order to identify specific plant anti-viral genes related to the miRNA regulatory pathway, RNA-Seq and sRNA-Seq were performed using Arabidopsis WT and dcl1-9 mutant line. A total of 5,204 DEGs were identified in TCV-infected WT plants. In contrast, only 595 DEGs were obtained in the infected dcl1-9 mutant plants. GO enrichment analysis of the shared DEGs and dcl1-9 unique DEGs showed that a wide range of biological processes were affected in the infected WT plants. In addition, miRNAs displayed different patterns between mock and infected WT plants...
October 26, 2016: Scientific Reports
Sofía Hernández-Arranz, Dione Sánchez-Hevia, Fernando Rojo, Renata Moreno
In Pseudomonas putida, the Hfq and Crc proteins regulate the expression of many genes in response to nutritional and environmental cues, by binding to mRNAs that bear specific target motifs and inhibiting their translation. The effect of these two proteins is antagonized by the CrcZ and CrcY small RNAs (sRNAs), the levels of which vary greatly according to growth conditions. The crcZ and crcY genes are transcribed from promoters PcrcZ and PcrcY, respectively, a process that relies on the CbrB transcriptional activator and the RpoN σ factor...
December 2016: RNA
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