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  1. Article ; Online: CRISPR interference to interrogate genes that control biofilm formation in Pseudomonas fluorescens

    Marie-Francoise Noirot-Gros / Sara Forrester / Grace Malato / Peter E. Larsen / Philippe Noirot

    Scientific Reports, Vol 9, Iss 1, Pp 1-

    2019  Volume 14

    Abstract: Abstract Bacterial biofilm formation involves signaling and regulatory pathways that control the transition from motile to sessile lifestyle, production of extracellular polymeric matrix, and maturation of the biofilm 3D structure. Biofilms are ... ...

    Abstract Abstract Bacterial biofilm formation involves signaling and regulatory pathways that control the transition from motile to sessile lifestyle, production of extracellular polymeric matrix, and maturation of the biofilm 3D structure. Biofilms are extensively studied because of their importance in biomedical, ecological and industrial settings. Gene inactivation is a powerful approach for functional studies but it is often labor intensive, limiting systematic gene surveys to the most tractable bacterial hosts. Here, we adapted the CRISPR interference (CRISPRi) system for use in diverse strain isolates of P. fluorescens, SBW25, WH6 and Pf0-1. We found that CRISPRi is applicable to study complex phenotypes such as cell morphology, motility and biofilm formation over extended periods of time. In SBW25, CRISPRi-mediated silencing of genes encoding the GacA/S two-component system and regulatory proteins associated with the cylic di-GMP signaling messenger produced swarming and biofilm phenotypes similar to those obtained after gene inactivation. Combined with detailed confocal microscopy of biofilms, our study also revealed novel phenotypes associated with extracellular matrix biosynthesis as well as the potent inhibition of SBW25 biofilm formation mediated by the PFLU1114 operon. We conclude that CRISPRi is a reliable and scalable approach to investigate gene networks in the diverse P. fluorescens group.
    Keywords Medicine ; R ; Science ; Q
    Subject code 572
    Language English
    Publishing date 2019-11-01T00:00:00Z
    Publisher Nature Publishing Group
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  2. Article ; Online: Constitutive Stringent Response Restores Viability of Bacillus subtilis Lacking Structural Maintenance of Chromosome Protein.

    Camille Benoist / Cyprien Guérin / Philippe Noirot / Etienne Dervyn

    PLoS ONE, Vol 10, Iss 11, p e

    2015  Volume 0142308

    Abstract: Bacillus subtilis mutants lacking the SMC-ScpAB complex are severely impaired for chromosome condensation and partitioning, DNA repair, and cells are not viable under standard laboratory conditions. We isolated suppressor mutations that restored the ... ...

    Abstract Bacillus subtilis mutants lacking the SMC-ScpAB complex are severely impaired for chromosome condensation and partitioning, DNA repair, and cells are not viable under standard laboratory conditions. We isolated suppressor mutations that restored the capacity of a smc deletion mutant (Δsmc) to grow under standard conditions. These suppressor mutations reduced chromosome segregation defects and abrogated hypersensitivity to gyrase inhibitors of Δsmc. Three suppressor mutations were mapped in genes involved in tRNA aminoacylation and maturation pathways. A transcriptomic survey of isolated suppressor mutations pointed to a potential link between suppression of Δsmc and induction of the stringent response. This link was confirmed by (p)ppGpp quantification which indicated a constitutive induction of the stringent response in multiple suppressor strains. Furthermore, sublethal concentrations of arginine hydroxamate (RHX), a potent inducer of stringent response, restored growth of Δsmc under non permissive conditions. We showed that production of (p)ppGpp alone was sufficient to suppress the thermosensitivity exhibited by the Δsmc mutant. Our findings shed new light on the coordination between chromosome dynamics mediated by SMC-ScpAB and other cellular processes during rapid bacterial growth.
    Keywords Medicine ; R ; Science ; Q
    Language English
    Publishing date 2015-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  3. Article ; Online: Tracking the Elusive Function of Bacillus subtilis Hfq.

    Tatiana Rochat / Olivier Delumeau / Nara Figueroa-Bossi / Philippe Noirot / Lionello Bossi / Etienne Dervyn / Philippe Bouloc

    PLoS ONE, Vol 10, Iss 4, p e

    2015  Volume 0124977

    Abstract: RNA-binding protein Hfq is a key component of the adaptive responses of many proteobacterial species including Escherichia coli, Salmonella enterica and Vibrio cholera. In these organisms, the importance of Hfq largely stems from its participation to ... ...

    Abstract RNA-binding protein Hfq is a key component of the adaptive responses of many proteobacterial species including Escherichia coli, Salmonella enterica and Vibrio cholera. In these organisms, the importance of Hfq largely stems from its participation to regulatory mechanisms involving small non-coding RNAs. In contrast, the function of Hfq in Gram-positive bacteria has remained elusive and somewhat controversial. In the present study, we have further addressed this point by comparing growth phenotypes and transcription profiles between wild-type and an hfq deletion mutant of the model Gram-positive bacterium, Bacillus subtilis. The absence of Hfq had no significant consequences on growth rates under nearly two thousand metabolic conditions and chemical treatments. The only phenotypic difference was a survival defect of B. subtilis hfq mutant in rich medium in stationary phase. Transcriptomic analysis correlated this phenotype with a change in the levels of nearly one hundred transcripts. Albeit a significant fraction of these RNAs (36%) encoded sporulation-related functions, analyses in a strain unable to sporulate ruled out sporulation per se as the basis of the hfq mutant's stationary phase fitness defect. When expressed in Salmonella, B. subtilis hfq complemented the sharp loss of viability of a degP hfq double mutant, attenuating the chronic σE-activated phenotype of this strain. However, B. subtilis hfq did not complement other regulatory deficiencies resulting from loss of Hfq-dependent small RNA activity in Salmonella indicating a limited functional overlap between Salmonella and B. subtilis Hfqs. Overall, this study confirmed that, despite structural similarities with other Hfq proteins, B. subtilis Hfq does not play a central role in post-transcriptional regulation but might have a more specialized function connected with stationary phase physiology. This would account for the high degree of conservation of Hfq proteins in all 17 B. subtilis strains whose genomes have been sequenced.
    Keywords Medicine ; R ; Science ; Q
    Language English
    Publishing date 2015-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  4. Article: Quantitative prediction of genome-wide resource allocation in bacteria

    Goelzer, Anne / Dörte Becher / Eric Prestel / Jan Muntel / Mahendra Mariadassou / Matthieu Jules / Michael Hecker / Philippe Noirot / Rolf Nölker / Stéphane Aymerich / Victor Chubukov / Vincent Fromion

    International Metabolic Engineering Society Metabolic engineering. 2015 Nov., v. 32

    2015  

    Abstract: Predicting resource allocation between cell processes is the primary step towards decoding the evolutionary constraints governing bacterial growth under various conditions. Quantitative prediction at genome-scale remains a computational challenge as ... ...

    Abstract Predicting resource allocation between cell processes is the primary step towards decoding the evolutionary constraints governing bacterial growth under various conditions. Quantitative prediction at genome-scale remains a computational challenge as current methods are limited by the tractability of the problem or by simplifying hypotheses. Here, we show that the constraint-based modeling method Resource Balance Analysis (RBA), calibrated using genome-wide absolute protein quantification data, accurately predicts resource allocation in the model bacterium Bacillus subtilis for a wide range of growth conditions. The regulation of most cellular processes is consistent with the objective of growth rate maximization except for a few suboptimal processes which likely integrate more complex objectives such as coping with stressful conditions and survival. As a proof of principle by using simulations, we illustrated how calibrated RBA could aid rational design of strains for maximizing protein production, offering new opportunities to investigate design principles in prokaryotes and to exploit them for biotechnological applications.
    Keywords Bacillus subtilis ; bacteria ; microbial growth ; models ; prediction ; prokaryotic cells ; resource allocation
    Language English
    Dates of publication 2015-11
    Size p. 232-243.
    Publishing place Elsevier Inc.
    Document type Article
    ZDB-ID 1470383-x
    ISSN 1096-7184 ; 1096-7176
    ISSN (online) 1096-7184
    ISSN 1096-7176
    DOI 10.1016/j.ymben.2015.10.003
    Database NAL-Catalogue (AGRICOLA)

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  5. Article ; Online: The cyst-dividing bacterium Ramlibacter tataouinensis TTB310 genome reveals a well-stocked toolbox for adaptation to a desert environment.

    Gilles De Luca / Mohamed Barakat / Philippe Ortet / Sylvain Fochesato / Cécile Jourlin-Castelli / Mireille Ansaldi / Béatrice Py / Gwennaele Fichant / Pedro M Coutinho / Romé Voulhoux / Olivier Bastien / Eric Maréchal / Bernard Henrissat / Yves Quentin / Philippe Noirot / Alain Filloux / Vincent Méjean / Michael S DuBow / Frédéric Barras /
    Valérie Barbe / Jean Weissenbach / Irina Mihalcescu / André Verméglio / Wafa Achouak / Thierry Heulin

    PLoS ONE, Vol 6, Iss 9, p e

    2011  Volume 23784

    Abstract: Ramlibacter tataouinensis TTB310(T) (strain TTB310), a betaproteobacterium isolated from a semi-arid region of South Tunisia (Tataouine), is characterized by the presence of both spherical and rod-shaped cells in pure culture. Cell division of strain ... ...

    Abstract Ramlibacter tataouinensis TTB310(T) (strain TTB310), a betaproteobacterium isolated from a semi-arid region of South Tunisia (Tataouine), is characterized by the presence of both spherical and rod-shaped cells in pure culture. Cell division of strain TTB310 occurs by the binary fission of spherical "cyst-like" cells ("cyst-cyst" division). The rod-shaped cells formed at the periphery of a colony (consisting mainly of cysts) are highly motile and colonize a new environment, where they form a new colony by reversion to cyst-like cells. This unique cell cycle of strain TTB310, with desiccation tolerant cyst-like cells capable of division and desiccation sensitive motile rods capable of dissemination, appears to be a novel adaptation for life in a hot and dry desert environment. In order to gain insights into strain TTB310's underlying genetic repertoire and possible mechanisms responsible for its unusual lifestyle, the genome of strain TTB310 was completely sequenced and subsequently annotated. The complete genome consists of a single circular chromosome of 4,070,194 bp with an average G+C content of 70.0%, the highest among the Betaproteobacteria sequenced to date, with total of 3,899 predicted coding sequences covering 92% of the genome. We found that strain TTB310 has developed a highly complex network of two-component systems, which may utilize responses to light and perhaps a rudimentary circadian hourglass to anticipate water availability at the dew time in the middle/end of the desert winter nights and thus direct the growth window to cyclic water availability times. Other interesting features of the strain TTB310 genome that appear to be important for desiccation tolerance, including intermediary metabolism compounds such as trehalose or polyhydroxyalkanoate, and signal transduction pathways, are presented and discussed.
    Keywords Medicine ; R ; Science ; Q
    Subject code 572
    Language English
    Publishing date 2011-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  6. Article: Condition-Dependent Transcriptome Reveals High-Level Regulatory Architecture in Bacillus subtilis

    Nicolas, Pierre / Anika March / Anne Goelzer / Annette Hansen / Anthony J. Wilkinson / Aurélie Leduc / Benno Schwikowski / Bernd Rinn / Colin R. Harwood / Dörte Becher / David Noone / Edda Klipp / Elena Bidnenko / Elisabeth Härtig / Elodie Marchadier / Emma L. Denham / Eric Botella / Etienne Dervyn / François Lecointe /
    Franck Samson / Frank Rügheimer / Geoff Doherty / Georg Homuth / Hanne Jarmer / Jörg Stülke / Jan Maarten van Dijl / Kevin M. Devine / Leif Steil / Ludovic Le Chat / Marc Schaffer / Mark Hoebeke / Mark J. Fogg / Matthieu Jules / Michael Hecker / Nathalie Pigeonneau / Olivier Delumeau / Paola Bisicchia / Peter Lewis / Philippe Bessières / Philippe Noirot / Praveen K. Sappa / Priyanka Nannapaneni / Ruben A. T. Mars / Stéphane Aymerich / Susanne Pohl / Tatiana Rochat / Thomas Wiegert / Ulrike Mäder / Uwe Völker / Vincent Fromion / Wolfram Liebermeister

    Science. 2012 Mar. 2, v. 335, no. 6072

    2012  

    Abstract: Bacteria adapt to environmental stimuli by adjusting their transcriptomes in a complex manner, the full potential of which has yet to be established for any individual bacterial species. Here, we report the transcriptomes of Bacillus subtilis exposed to ... ...

    Abstract Bacteria adapt to environmental stimuli by adjusting their transcriptomes in a complex manner, the full potential of which has yet to be established for any individual bacterial species. Here, we report the transcriptomes of Bacillus subtilis exposed to a wide range of environmental and nutritional conditions that the organism might encounter in nature. We comprehensively mapped transcription units (TUs) and grouped 2935 promoters into regulons controlled by various RNA polymerase sigma factors, accounting for approximately 66% of the observed variance in transcriptional activity. This global classification of promoters and detailed description of TUs revealed that a large proportion of the detected antisense RNAs arose from potentially spurious transcription initiation by alternative sigma factors and from imperfect control of transcription termination.
    Keywords Bacillus subtilis ; bacteria ; DNA-directed RNA polymerase ; regulon ; RNA ; sigma factors ; transcription (genetics) ; transcriptome ; variance
    Language English
    Dates of publication 2012-0302
    Size p. 1103-1106.
    Publishing place American Association for the Advancement of Science
    Document type Article
    ZDB-ID 128410-1
    ISSN 1095-9203 ; 0036-8075
    ISSN (online) 1095-9203
    ISSN 0036-8075
    DOI 10.1126/science.1206848
    Database NAL-Catalogue (AGRICOLA)

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  7. Article: Global Network Reorganization During Dynamic Adaptations of Bacillus subtilis Metabolism

    Buescher, Joerg Martin / Anne Goelzer / Annette Hansen / Anthony J. Wilkinson / Aurélie Leduc / Benno Schwikowski / Bernd Hessling / Bernd Rinn / Claus Hultschig / Colin R. Harwood / Dörte Becher / Edda Klipp / Elena Bidnenko / Emma L. Denham / Eric Botella / Etienne Dervyn / François Lecointe / Frank Molina / Frank Rügheimer /
    Geoff Doherty / Hanne Jarmer / Jörg Stelling / Jan Maarten Van Dijl / Jan Muntel / Julian Schnidder / Kevin M. Devine / Liza Felicori / Ludovic Le Chat / Marc Schaffer / Mark J. Fogg / Markus Uhr / Matthieu Jules / Michael Hecker / Nathalie Pigeonneau / Patrick Veiga / Peter Lewis / Philippe Bessieres / Philippe Noirot / Pierre Nicolas / Roelco Jacobus Kleijn / Sabine Peres / Samuel Drulhe / Sean Walsh / Sebastian Hubner / Simon Rasmussen / Sjouke Piersma / Stéphane Aymerich / Susanne Pohl / Ulrike Mäder / Uwe Sauer / Wolfram Liebermeister

    Science. 2012 Mar. 2, v. 335, no. 6072

    2012  

    Abstract: Adaptation of cells to environmental changes requires dynamic interactions between metabolic and regulatory networks, but studies typically address only one or a few layers of regulation. For nutritional shifts between two preferred carbon sources of ... ...

    Abstract Adaptation of cells to environmental changes requires dynamic interactions between metabolic and regulatory networks, but studies typically address only one or a few layers of regulation. For nutritional shifts between two preferred carbon sources of Bacillus subtilis, we combined statistical and model-based data analyses of dynamic transcript, protein, and metabolite abundances and promoter activities. Adaptation to malate was rapid and primarily controlled posttranscriptionally compared with the slow, mainly transcriptionally controlled adaptation to glucose that entailed nearly half of the known transcription regulation network. Interactions across multiple levels of regulation were involved in adaptive changes that could also be achieved by controlling single genes. Our analysis suggests that global trade-offs and evolutionary constraints provide incentives to favor complex control programs.
    Keywords Bacillus subtilis ; carbon ; genes ; glucose ; malates ; metabolism ; metabolites ; transcription (genetics)
    Language English
    Dates of publication 2012-0302
    Size p. 1099-1103.
    Publishing place American Association for the Advancement of Science
    Document type Article
    ZDB-ID 128410-1
    ISSN 1095-9203 ; 0036-8075
    ISSN (online) 1095-9203
    ISSN 0036-8075
    DOI 10.1126/science.1206871
    Database NAL-Catalogue (AGRICOLA)

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