scholarly journals GAUGE-Annotated Microbial Transcriptomic Data Facilitate Parallel Mining and High-Throughput Reanalysis To Form Data-Driven Hypotheses

mSystems ◽  
2021 ◽  
Vol 6 (2) ◽  
Author(s):  
Zhongyou Li ◽  
Katja Koeppen ◽  
Victoria I. Holden ◽  
Samuel L. Neff ◽  
Liviu Cengher ◽  
...  
Keyword(s):  
Gene Expression ◽  
High Throughput ◽  
Biofilm Formation ◽  
Data Driven ◽  
Data Sets ◽  
Web Interface ◽  
Content Type ◽  
Transcriptomic Data ◽  
Parallel Mining ◽  
Differential Gene

ABSTRACT The NCBI Gene Expression Omnibus (GEO) provides tools to query and download transcriptomic data. However, less than 4% of microbial experiments include the sample group annotations required to assess differential gene expression for high-throughput reanalysis, and data deposited after 2014 universally lack these annotations. Our algorithm GAUGE (general annotation using text/data group ensembles) automatically annotates GEO microbial data sets, including microarray and RNA sequencing studies, increasing the percentage of data sets amenable to analysis from 4% to 33%. Eighty-nine percent of GAUGE-annotated studies matched group assignments generated by human curators. To demonstrate how GAUGE annotation can lead to scientific insight, we created GAPE (GAUGE-annotated Pseudomonas aeruginosa and Escherichia coli transcriptomic compendia for reanalysis), a Shiny Web interface to analyze 73 GAUGE-annotated P. aeruginosa studies, three times more than previously available. GAPE analysis revealed that PA3923, a gene of unknown function, was frequently differentially expressed in more than 50% of studies and significantly coregulated with genes involved in biofilm formation. Follow-up wet-bench experiments demonstrate that PA3923 mutants are indeed defective in biofilm formation, consistent with predictions facilitated by GAUGE and GAPE. We anticipate that GAUGE and GAPE, which we have made freely available, will make publicly available microbial transcriptomic data easier to reuse and lead to new data-driven hypotheses. IMPORTANCE GEO archives transcriptomic data from over 5,800 microbial experiments and allows researchers to answer questions not directly addressed in published papers. However, less than 4% of the microbial data sets include the sample group annotations required for high-throughput reanalysis. This limitation blocks a considerable amount of microbial transcriptomic data from being reused easily. Here, we demonstrate that the GAUGE algorithm could make 33% of microbial data accessible to parallel mining and reanalysis. GAUGE annotations increase statistical power and, thereby, make consistent patterns of differential gene expression easier to identify. In addition, we developed GAPE (GAUGE-annotated Pseudomonas aeruginosa and Escherichia coli transcriptomic compendia for reanalysis), a Shiny Web interface that performs parallel analyses on P. aeruginosa and E. coli compendia. Source code for GAUGE and GAPE is freely available and can be repurposed to create compendia for other bacterial species.

scholarly journals RNA Sequencing Data Sets Identifying Differentially Expressed Transcripts during Campylobacter jejuni Biofilm Formation

2020 ◽  
Vol 9 (1) ◽  
Author(s):  
Greg Tram ◽  
William P. Klare ◽  
Joel A. Cain ◽  
Basem Mourad ◽  
Stuart J. Cordwell ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Sequencing ◽  
Campylobacter Jejuni ◽  
Biofilm Formation ◽  
Foodborne Pathogen ◽  
Data Sets ◽  
Sequencing Data ◽  
Content Type ◽  
Differential Gene ◽  
Insight Into

Campylobacter jejuni is a foodborne pathogen and an important contributor to gastroenteritis in humans. C. jejuni readily forms biofilms which may play a role in the transmission of the pathogen from animals to humans. Herein, we present RNA sequencing data investigating differential gene expression in biofilm and planktonic C. jejuni. These data provide insight into pathways which may be important to biofilm formation in this organism.

scholarly journals Reversible Gene Expression Control inYersinia pestisby Using an Optimized CRISPR Interference System

2019 ◽  
Vol 85 (12) ◽  
Author(s):  
Tong Wang ◽  
Min Wang ◽  
Qingwen Zhang ◽  
Shiyang Cao ◽  
Xiang Li ◽  
...  
Keyword(s):  
Gene Expression ◽  
High Throughput ◽  
Biofilm Formation ◽  
Prevention And Control ◽  
Target Gene ◽  
Gene Knockdown ◽  
Content Type ◽  
Short Palindromic Repeat ◽  
And Control

ABSTRACTMany genes in the bacterial pathogenYersinia pestis, the causative agent of three plague pandemics, remain uncharacterized, greatly hampering the development of measures for plague prevention and control. Clustered regularly interspaced short palindromic repeat interference (CRISPRi) has been shown to be an effective tool for gene knockdown in model bacteria. In this system, a catalytically dead Cas9 (dCas9) and a small guide RNA (sgRNA) form a complex, binding to the specific DNA target through base pairing, thereby impeding RNA polymerase binding and causing target gene repression. Here, we introduce an optimized CRISPRi system usingStreptococcus pyogenesCas9-derived dCas9 for gene knockdown inY. pestis. Multiple genes harbored on either the chromosome or plasmids ofY. pestiswere efficiently knocked down (up to 380-fold) in a strictly anhydrotetracycline-inducible manner using this CRISPRi approach. Knockdown ofhmsH(responsible for biofilm formation) orcspB(encoding a cold shock protein) resulted in greatly decreased biofilm formation or impaired cold tolerance inin vitrophenotypic assays. Furthermore, silencing of the virulence-associated genesyscBorailusing this CRISPRi system resulted in attenuation of virulence in HeLa cells and mice similar to that previously reported foryscBandailnull mutants. Taken together, our results confirm that this optimized CRISPRi system can reversibly and efficiently repress the expression of target genes inY. pestis, providing an alternative to conventional gene knockdown techniques, as well as a strategy for high-throughput phenotypic screening ofY. pestisgenes with unknown functions.IMPORTANCEYersiniapestisis a lethal pathogen responsible for millions of human deaths in history. It has also attracted much attention for potential uses as a bioweapon or bioterrorism agent, against which new vaccines are desperately needed. However, manyY. pestisgenes remain uncharacterized, greatly hampering the development of measures for plague prevention and control. Clustered regularly interspaced short palindromic repeat interference (CRISPRi) has been successfully used in a variety of bacteria in functional genomic studies, but no such genetic tool has been reported inY. pestis. Here, we systematically optimized the CRISPRi approach for use inY. pestis, which ultimately repressed target gene expression with high efficiency in a reversible manner. Knockdown of functional genes using this method produced phenotypes that were readily detected byin vitroassays, cell infection assays, and mouse infection experiments. This is a report of a CRISPRi approach inY. pestisand highlights the potential use of this approach in high-throughput functional genomics studies of this pathogen.

scholarly journals Horizontal meta-analysis identifies common deregulated genes across AML subgroups providing a robust prognostic signature

2020 ◽  
Vol 4 (20) ◽  
pp. 5322-5335
Author(s):  
Ali Nehme ◽  
Hassan Dakik ◽  
Frédéric Picou ◽  
Meyling Cheok ◽  
Claude Preudhomme ◽  
...  
Keyword(s):  
Gene Expression ◽  
Microarray Data ◽  
Meta Analysis ◽  
Prognostic Score ◽  
Gene Expression Omnibus ◽  
Response To Therapy ◽  
Data Sets ◽  
Dna Hypermethylation ◽  
Transcriptomic Data ◽  
Differential Gene

Abstract Advances in transcriptomics have improved our understanding of leukemic development and helped to enhance the stratification of patients. The tendency of transcriptomic studies to combine AML samples, regardless of cytogenetic abnormalities, could lead to bias in differential gene expression analysis because of the differential representation of AML subgroups. Hence, we performed a horizontal meta-analysis that integrated transcriptomic data on AML from multiple studies, to enrich the less frequent cytogenetic subgroups and to uncover common genes involved in the development of AML and response to therapy. A total of 28 Affymetrix microarray data sets containing 3940 AML samples were downloaded from the Gene Expression Omnibus database. After stringent quality control, transcriptomic data on 1534 samples from 11 data sets, covering 10 AML cytogenetically defined subgroups, were retained and merged with the data on 198 healthy bone marrow samples. Differentially expressed genes between each cytogenetic subgroup and normal samples were extracted, enabling the unbiased identification of 330 commonly deregulated genes (CODEGs), which showed enriched profiles of myeloid differentiation, leukemic stem cell status, and relapse. Most of these genes were downregulated, in accordance with DNA hypermethylation. CODEGs were then used to create a prognostic score based on the weighted sum of expression of 22 core genes (CODEG22). The score was validated with microarray data of 5 independent cohorts and by quantitative real time-polymerase chain reaction in a cohort of 142 samples. CODEG22-based stratification of patients, globally and into subpopulations of cytologically healthy and elderly individuals, may complement the European LeukemiaNet classification, for a more accurate prediction of AML outcomes.

scholarly journals Adaptation of Lactobacillus plantarum IMDO 130201, a Wheat Sourdough Isolate, to Growth in Wheat Sourdough Simulation Medium at Different pH Values through Differential Gene Expression

2011 ◽  
Vol 77 (10) ◽  
pp. 3406-3412 ◽  
Author(s):  
Gino Vrancken ◽  
Luc De Vuyst ◽  
Tom Rimaux ◽  
Joke Allemeersch ◽  
Stefan Weckx
Keyword(s):  
Gene Expression ◽  
Lactobacillus Plantarum ◽  
Differential Gene Expression ◽  
Lipoteichoic Acid ◽  
Low Ph ◽  
Exponential Growth Phase ◽  
Cellular Mechanisms ◽  
Content Type ◽  
Ph Values ◽  
Differential Gene

ABSTRACTSourdough is a very competitive and challenging environment for microorganisms. Usually, a stable microbiota composed of lactic acid bacteria (LAB) and yeasts dominates this ecosystem. Although sourdough is rich in carbohydrates, thus providing an ideal environment for microorganisms to grow, its low pH presents a particular challenge. The nature of the adaptation to this low pH was investigated forLactobacillus plantarumIMDO 130201, an isolate from a laboratory wheat sourdough fermentation. Batch fermentations were carried out in wheat sourdough simulation medium, and total RNA was isolated from mid-exponential-growth-phase cultures, followed by differential gene expression analysis using a LAB functional gene microarray. At low pH values, an increased expression of genes involved in peptide and amino acid metabolism was found as well as that of genes involved in plantaricin production and lipoteichoic acid biosynthesis. The results highlight cellular mechanisms that allowL. plantarumto function at a low environmental pH.

scholarly journals Cyclic Diguanylate Regulates Virulence Factor Genes via Multiple Riboswitches inClostridium difficile

mSphere ◽  
2018 ◽  
Vol 3 (5) ◽  
Author(s):  
Robert W. McKee ◽  
Carissa K. Harvest ◽  
Rita Tamayo
Keyword(s):  
Gene Expression ◽  
Clostridium Difficile ◽  
Biofilm Formation ◽  
Toxin Production ◽  
Intestinal Colonization ◽  
Cyclic Diguanylate ◽  
Signaling Molecule ◽  
Content Type ◽  
Surface Motility

ABSTRACTThe intracellular signaling molecule cyclic diguanylate (c-di-GMP) regulates many processes in bacteria, with a central role in controlling the switch between motile and nonmotile lifestyles. Recent work has shown that inClostridium difficile(also calledClostridioides difficile), c-di-GMP regulates swimming and surface motility, biofilm formation, toxin production, and intestinal colonization. In this study, we determined the transcriptional regulon of c-di-GMP inC. difficile,employing overexpression of a diguanylate cyclase gene to artificially manipulate intracellular c-di-GMP. Consistent with prior work, c-di-GMP regulated the expression of genes involved in swimming and surface motility. c-di-GMP also affected the expression of multiple genes encoding cell envelope proteins, several of which affected biofilm formationin vitro. A substantial proportion of the c-di-GMP regulon appears to be controlled either directly or indirectly via riboswitches. We confirmed the functionality of 11 c-di-GMP riboswitches, demonstrating their effects on downstream gene expression independent of the upstream promoters. The class I riboswitches uniformly functioned as “off” switches in response to c-di-GMP, while class II riboswitches acted as “on” switches. Transcriptional analyses of genes 3′ of c-di-GMP riboswitches over a broad range of c-di-GMP levels showed that relatively modest changes in c-di-GMP levels are capable of altering gene transcription, with concomitant effects on microbial behavior. This work expands the known c-di-GMP signaling network inC. difficileand emphasizes the role of the riboswitches in controlling known and putative virulence factors inC. difficile.IMPORTANCEInClostridium difficile, the signaling molecule c-di-GMP regulates multiple processes affecting its ability to cause disease, including swimming and surface motility, biofilm formation, toxin production, and intestinal colonization. In this study, we used RNA-seq to define the transcriptional regulon of c-di-GMP inC. difficile. Many new targets of c-di-GMP regulation were identified, including multiple putative colonization factors. Transcriptional analyses revealed a prominent role for riboswitches in c-di-GMP signaling. Only a subset of the 16 previously predicted c-di-GMP riboswitches were functionalin vivoand displayed potential variability in their response kinetics to c-di-GMP. This work underscores the importance of studying c-di-GMP riboswitches in a relevant biological context and highlights the role of the riboswitches in controlling gene expression inC. difficile.

scholarly journals CsrA (BB0184) Is Not Involved in Activation of the RpoN-RpoS Regulatory Pathway in Borrelia burgdorferi

2014 ◽  
Vol 82 (4) ◽  
pp. 1511-1522 ◽  
Author(s):  
Zhiming Ouyang ◽  
Jianli Zhou ◽  
Michael V. Norgard
Keyword(s):  
Gene Expression ◽  
Borrelia Burgdorferi ◽  
Animal Studies ◽  
Expression System ◽  
Regulatory Pathway ◽  
Content Type ◽  
Mouse Tissues ◽  
Differential Gene ◽  
Carbon Storage Regulator

ABSTRACTBorrelia burgdorferiencodes a homologue of the bacterial carbon storage regulator A (CsrA). Recently, it was reported that CsrA contributes toB. burgdorferiinfectivity and is required for the activation of the central RpoN-RpoS regulatory pathway. However, many questions concerning the function of CsrA inB. burgdorferigene regulation remain unanswered. In particular, there are conflicting reports concerning the molecular details of how CsrA may modulaterpoSexpression and, thus, how CsrA may influence the RpoN-RpoS pathway inB. burgdorferi. To address these key discrepancies, we examined the role of CsrA in differential gene expression in the Lyme disease spirochete. Upon engineering an induciblecsrAexpression system inB. burgdorferi, controlled hyperexpression of CsrA in a merodiploid strain did not significantly alter the protein and transcript levels ofbosR,rpoS, and RpoS-dependent genes (such asospCanddbpA). In addition, we constructed isogeniccsrAmutants in two widely used infectiousB. burgdorferistrains. When expression ofbosR,rpoS,ospC, anddbpAwas compared between thecsrAmutants and their wild-type counterparts, no detectable differences were observed. Finally, animal studies indicated that thecsrAmutants remained infectious for and virulent in mice. Analyses ofB. burgdorferigene expression in mouse tissues showed comparable levels ofrpoStranscripts by thecsrAmutants and the parental strains. Taken together, these results constitute compelling evidence that CsrA is not involved in activation of the RpoN-RpoS pathway and is dispensable for mammalian infectious processes carried out byB. burgdorferi.

scholarly journals Flagellum-Mediated Mechanosensing and RflP Control Motility State of Pathogenic Escherichia coli

mBio ◽  
2020 ◽  
Vol 11 (2) ◽  
Author(s):  
Leanid Laganenka ◽  
María Esteban López ◽  
Remy Colin ◽  
Victor Sourjik
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Porous Medium ◽  
Biofilm Formation ◽  
Liquid Culture ◽  
Content Type ◽  
E Coli ◽  
Surface Contact ◽  
Conditional Inhibition ◽  
Flagellar Genes

ABSTRACT Bacterial flagellar motility plays an important role in many processes that occur at surfaces or in hydrogels, including adhesion, biofilm formation, and bacterium-host interactions. Consequently, expression of flagellar genes, as well as genes involved in biofilm formation and virulence, can be regulated by the surface contact. In a few bacterial species, flagella themselves are known to serve as mechanosensors, where an increased load on flagella experienced during surface contact or swimming in viscous media controls gene expression. In this study, we show that gene regulation by motility-dependent mechanosensing is common among pathogenic Escherichia coli strains. This regulatory mechanism requires flagellar rotation, and it enables pathogenic E. coli to repress flagellar genes at low loads in liquid culture, while activating motility in porous medium (soft agar) or upon surface contact. It also controls several other cellular functions, including metabolism and signaling. The mechanosensing response in pathogenic E. coli depends on the negative regulator of motility, RflP (YdiV), which inhibits basal expression of flagellar genes in liquid. While no conditional inhibition of flagellar gene expression in liquid and therefore no upregulation in porous medium was observed in the wild-type commensal or laboratory strains of E. coli, mechanosensitive regulation could be recovered by overexpression of RflP in the laboratory strain. We hypothesize that this conditional activation of flagellar genes in pathogenic E. coli reflects adaptation to the dual role played by flagella and motility during infection. IMPORTANCE Flagella and motility are widespread virulence factors among pathogenic bacteria. Motility enhances the initial host colonization, but the flagellum is a major antigen targeted by the host immune system. Here, we demonstrate that pathogenic E. coli strains employ a mechanosensory function of the flagellar motor to activate flagellar expression under high loads, while repressing it in liquid culture. We hypothesize that this mechanism allows pathogenic E. coli to regulate its motility dependent on the stage of infection, activating flagellar expression upon initial contact with the host epithelium, when motility is beneficial, but reducing it within the host to delay the immune response.

scholarly journals Selective Pressure for Biofilm Formation in Bacillus subtilis: Differential Effect of Mutations in the Master Regulator SinR on Bistability

mBio ◽  
2018 ◽  
Vol 9 (5) ◽  
Author(s):  
Jan Kampf ◽  
Jan Gerwig ◽  
Kerstin Kruse ◽  
Robert Cleverley ◽  
Miriam Dormeyer ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bacillus Subtilis ◽  
Biofilm Formation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Selective Pressure ◽  
Wild Type ◽  
Master Regulator ◽  
Form Complex ◽  
Content Type

ABSTRACT Biofilm formation by Bacillus subtilis requires the expression of genes encoding enzymes for extracellular polysaccharide synthesis and for an amyloid-like protein. The master regulator SinR represses all the corresponding genes, and repression of these key biofilm genes is lifted when SinR interacts with its cognate antagonist proteins. The YmdB phosphodiesterase is a recently discovered factor that is involved in the control of SinR activity: cells lacking YmdB exhibit hyperactive SinR and are unable to relieve the repression of the biofilm genes. In this study, we have examined the dynamics of gene expression patterns in wild-type and ymdB mutant cells by microfluidic analysis coupled to time-lapse microscopy. Our results confirm the bistable expression pattern for motility and biofilm genes in the wild-type strain and the loss of biofilm gene expression in the mutant. Moreover, we demonstrated dynamic behavior in subpopulations of the wild-type strain that is characterized by switches in sets of the expressed genes. In order to gain further insights into the role of YmdB, we isolated a set of spontaneous suppressor mutants derived from ymdB mutants that had regained the ability to form complex colonies and biofilms. Interestingly, all of the mutations affected SinR. In some mutants, large genomic regions encompassing sinR were deleted, whereas others had alleles encoding SinR variants. Functional and biochemical studies with these SinR variants revealed how these proteins allowed biofilm gene expression in the ymdB mutant strains. IMPORTANCE Many bacteria are able to choose between two mutually exclusive lifestyles: biofilm formation and motility. In the model bacterium Bacillus subtilis, this choice is made by each individual cell rather than at the population level. The transcriptional repressor SinR is the master regulator in this decision-making process. The regulation of SinR activity involves complex control of its own expression and of its interaction with antagonist proteins. We show that the YmdB phosphodiesterase is required to allow the expression of SinR-repressed genes in a subpopulation of cells and that such subpopulations can switch between different SinR activity states. Suppressor analyses revealed that ymdB mutants readily acquire mutations affecting SinR, thus restoring biofilm formation. These findings suggest that B. subtilis cells experience selective pressure to form the extracellular matrix that is characteristic of biofilms and that YmdB is required for the homeostasis of SinR and/or its antagonists.

scholarly journals Transcriptomic Data Sets To Determine Gene Expression Changes Mediated by the Presence of PBT2 in Growth Medium of Multidrug-Resistant Neisseria gonorrhoeae WHO Z

2020 ◽  
Vol 9 (21) ◽  
Author(s):  
Freda E.-C. Jen ◽  
Ibrahim M. El-Deeb ◽  
John M. Atack ◽  
Mark von Itzstein ◽  
Michael P. Jennings
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Transcriptome Sequencing ◽  
Multidrug Resistant ◽  
Data Sets ◽  
Sequencing Data ◽  
Transmitted Infection ◽  
High Coverage ◽  
Content Type ◽  
Sexually Transmitted ◽  
Transcriptomic Data

ABSTRACT Neisseria gonorrhoeae causes the sexually transmitted infection gonorrhea. High-coverage (∼3,300-fold) transcriptome sequencing data have been collected from multidrug-resistant N. gonorrhoeae strain WHO Z grown in the presence and absence of PBT2.

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