scholarly journals Modeling the time course of ComX: towards molecular process control for Bacillus wild-type cultivations

AMB Express ◽  
2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Chantal Treinen ◽  
Olivia Magosch ◽  
Mareen Hoffmann ◽  
Peter Klausmann ◽  
Berit Würtz ◽  
...  
Keyword(s):  
Quorum Sensing ◽  
Genetic Engineering ◽  
Process Control ◽  
Time Course ◽  
Control Strategies ◽  
Mathematic Model ◽  
Pilot Scale ◽  
Industrial Biotechnology ◽  
Wild Type ◽  
Molecular Process

AbstractWild-type cultivations are of invaluable relevance for industrial biotechnology when it comes to the agricultural or food sector. Here, genetic engineering is hardly applicable due to legal barriers and consumer’s demand for GMO-free products. An important pillar for wild-type cultivations displays the genus Bacillus. One of the challenges for Bacillus cultivations is the global ComX-dependent quorum sensing system. Here, molecular process control can serve as a tool to optimize the production process without genetic engineering. To realize this approach, quantitative knowledge of the mechanism is essential, which, however, is often available only to a limited extent. The presented work provides a case study based on the production of cyclic lipopeptide surfactin, whose expression is in dependence of ComX, using natural producer B. subtilis DSM 10 T. First, a surfactin reference process with 40 g/L of glucose was performed as batch fermentation in a pilot scale bioreactor system to gain novel insights into kinetic behavior of ComX in relation to surfactin production. Interestingly, the specific surfactin productivity did not increase linearly with ComX activity. The data were then used to derive a mathematic model for the time course of ComX in dependence of existing biomass, biomass growth as well as a putative ComX-specific protease. The newly adapted model was validated and transferred to other batch fermentations, employing 20 and 60 g/L glucose. The applied approach can serve as a model system for molecular process control strategies, which can thus be extended to other quorum sensing dependent wild-type cultivations.

scholarly journals The ComX Quorum Sensing Peptide of Bacillus subtilis Affects Biofilm Formation Negatively and Sporulation Positively

2020 ◽  
Vol 8 (8) ◽  
pp. 1131 ◽  
Author(s):  
Mihael Špacapan ◽  
Tjaša Danevčič ◽  
Polonca Štefanic ◽  
Michael Porter ◽  
Nicola R. Stanley-Wall ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Quorum Sensing ◽  
Biofilm Formation ◽  
Matrix Protein ◽  
Control Strategies ◽  
Wild Type ◽  
Fibrous Matrix ◽  
Sensing System ◽  
Quorum Sensing System ◽  
Biofilm Development

Quorum sensing (QS) is often required for the formation of bacterial biofilms and is a popular target of biofilm control strategies. Previous studies implicate the ComQXPA quorum sensing system of Bacillus subtilis as a promoter of biofilm formation. Here, we report that ComX signaling peptide deficient mutants form thicker and more robust pellicle biofilms that contain chains of cells. We confirm that ComX positively affects the transcriptional activity of the PepsA promoter, which controls the synthesis of the major matrix polysaccharide. In contrast, ComX negatively controls the PtapA promoter, which drives the production of TasA, a fibrous matrix protein. Overall, the biomass of the mutant biofilm lacking ComX accumulates more monosaccharide and protein content than the wild type. We conclude that this QS phenotype might be due to extended investment into growth rather than spore development. Consistent with this, the ComX deficient mutant shows a delayed activation of the pre-spore specific promoter, PspoIIQ, and a delayed, more synchronous commitment to sporulation. We conclude that ComX mediated early commitment to sporulation of the wild type slows down biofilm formation and modulates the coexistence of multiple biological states during the early stages of biofilm development.

Arabidopsis 4-COUMAROYL-COA LIGASE 8 contributes to the biosynthesis of the benzenoid ring of coenzyme Q in peroxisomes

2019 ◽  
Vol 476 (22) ◽  
pp. 3521-3532
Author(s):  
Eric Soubeyrand ◽  
Megan Kelly ◽  
Shea A. Keene ◽  
Ann C. Bernert ◽  
Scott Latimer ◽  
...  
Keyword(s):  
Oxidative Metabolism ◽  
Time Course ◽  
Reverse Genetics ◽  
De Novo ◽  
Coenzyme Q ◽  
Control Level ◽  
Wild Type ◽  
Fluorescent Reporters ◽  
Coa Ligase ◽  
Peroxisomal Targeting

Plants have evolved the ability to derive the benzenoid moiety of the respiratory cofactor and antioxidant, ubiquinone (coenzyme Q), either from the β-oxidative metabolism of p-coumarate or from the peroxidative cleavage of kaempferol. Here, isotopic feeding assays, gene co-expression analysis and reverse genetics identified Arabidopsis 4-COUMARATE-COA LIGASE 8 (4-CL8; At5g38120) as a contributor to the β-oxidation of p-coumarate for ubiquinone biosynthesis. The enzyme is part of the same clade (V) of acyl-activating enzymes than At4g19010, a p-coumarate CoA ligase known to play a central role in the conversion of p-coumarate into 4-hydroxybenzoate. A 4-cl8 T-DNA knockout displayed a 20% decrease in ubiquinone content compared with wild-type plants, while 4-CL8 overexpression boosted ubiquinone content up to 150% of the control level. Similarly, the isotopic enrichment of ubiquinone's ring was decreased by 28% in the 4-cl8 knockout as compared with wild-type controls when Phe-[Ring-13C6] was fed to the plants. This metabolic blockage could be bypassed via the exogenous supply of 4-hydroxybenzoate, the product of p-coumarate β-oxidation. Arabidopsis 4-CL8 displays a canonical peroxisomal targeting sequence type 1, and confocal microscopy experiments using fused fluorescent reporters demonstrated that this enzyme is imported into peroxisomes. Time course feeding assays using Phe-[Ring-13C6] in a series of Arabidopsis single and double knockouts blocked in the β-oxidative metabolism of p-coumarate (4-cl8; at4g19010; at4g19010 × 4-cl8), flavonol biosynthesis (flavanone-3-hydroxylase), or both (at4g19010 × flavanone-3-hydroxylase) indicated that continuous high light treatments (500 µE m−2 s−1; 24 h) markedly stimulated the de novo biosynthesis of ubiquinone independently of kaempferol catabolism.

Postnatal growth rate and gonadal development in circadian tau mutant hamsters reared in constant dim red light

Reproduction ◽  
2000 ◽  
pp. 327-330 ◽  
Author(s):  
RJ Lucas ◽  
JA Stirland ◽  
YN Mohammad ◽  
AS Loudon
Keyword(s):  
Time Course ◽  
Red Light ◽  
Somatic Growth ◽  
Postnatal Growth ◽  
Gonadal Development ◽  
Testicular Development ◽  
Wild Type ◽  
Similar Time ◽  
Syrian Hamsters ◽  
Body Weights

The role of the circadian clock in the reproductive development of Syrian hamsters (Mesocricetus auratus was examined in wild type and circadian tau mutant hamsters reared from birth to 26 weeks of age under constant dim red light. Testis diameter and body weights were determined at weekly intervals in male hamsters from 4 weeks of age. In both genotypes, testicular development, subsequent regression and recrudescence exhibited a similar time course. The age at which animals displayed reproductive photosensitivity, as exhibited by testicular regression, was unrelated to circadian genotype (mean +/- SEM: 54 +/- 3 days for wild type and 59 +/- 5 days for tau mutants). In contrast, our studies revealed a significant impact of the mutation on somatic growth, such that tau mutants weighed 18% less than wild types at the end of the experiment. Our study reveals that the juvenile onset of reproductive photoperiodism in Syrian hamsters is not timed by the circadian system.

Optimization and Control Petroleum Refinery Wastewater Treatment Systems — Status, Trends and Needs

1989 ◽  
Vol 24 (3) ◽  
pp. 463-477
Author(s):  
Stephen G. Nutt
Keyword(s):  
Wastewater Treatment ◽  
Process Control ◽  
Petroleum Refinery ◽  
Control Strategies ◽  
Refinery Wastewater ◽  
Statistical Process ◽  
Petroleum Refinery Wastewater ◽  
Optimization And Control ◽  
And Control ◽  
Wastewater Treatment Systems

Abstract Based on discussions in workshop sessions, several recurring themes became evident with respect to the optimization and control of petroleum refinery wastewater treatment systems to achieve effective removal of toxic contaminants. It was apparent that statistical process control (SPC) techniques are finding more widespread use and have been found to be effective. However, the implementation of real-time process control strategies in petroleum refinery wastewater treatment systems is in its infancy. Considerable effort will need to be expended to demonstrate the practicality of on-line sensors, and the utility of automated process control in petroleum refinery wastewater treatment systems. This paper provides a summary of the discussions held at the workshop.

scholarly journals N-3-Hydroxy Dodecanoyl-DL-homoserine Lactone (OH-dDHL) Triggers Apoptosis of Bone Marrow-Derived Macrophages through the ER- and Mitochondria-Mediated Pathways

2021 ◽  
Vol 22 (14) ◽  
pp. 7565
Author(s):  
Kyungho Woo ◽  
Dong Ho Kim ◽  
Man Hwan Oh ◽  
Ho Sung Park ◽  
Chul Hee Choi
Keyword(s):  
Bone Marrow ◽  
Quorum Sensing ◽  
Deletion Mutant ◽  
Transmembrane Protein ◽  
Cell Communication ◽  
Homoserine Lactone ◽  
Host Responses ◽  
Wild Type ◽  
Mutant Strains

Quorum sensing of Acinetobacter nosocomialis for cell-to-cell communication produces N-3-hydroxy dodecanoyl-DL-homoserine lactone (OH-dDHL) by an AnoR/I two-component system. However, OH-dDHL-driven apoptotic mechanisms in hosts have not been clearly defined. Here, we investigated the induction of apoptosis signaling pathways in bone marrow-derived macrophages treated with synthetic OH-dDHL. Moreover, the quorum-sensing system for virulence regulation was evaluated in vivo using wild-type and anoI-deletion mutant strains. OH-dDHL decreased the viability of macrophage and epithelial cells in dose- and time-dependent manners. OH-dDHL induced Ca2+ efflux and caspase-12 activation by ER stress transmembrane protein (IRE1 and ATF6a p50) aggregation and induced mitochondrial dysfunction through reactive oxygen species (ROS) production, which caused cytochrome c to leak. Pretreatment with a pan-caspase inhibitor reduced caspase-3, -8, and -9, which were activated by OH-dDHL. Pro-inflammatory cytokine and paraoxonase-2 (PON2) gene expression were increased by OH-dDHL. We showed that the anoI-deletion mutant strains have less intracellular invasion compared to the wild-type strain, and their virulence, such as colonization and dissemination, was decreased in vivo. Consequently, these findings revealed that OH-dDHL, as a virulence factor, contributes to bacterial infection and survival as well as the modification of host responses in the early stages of infection.

scholarly journals Quorum Sensing: a Transcriptional Regulatory System Involved in the Pathogenicity of Burkholderia mallei

2004 ◽  
Vol 72 (11) ◽  
pp. 6589-6596 ◽  
Author(s):  
Ricky L. Ulrich ◽  
David DeShazer ◽  
Harry B. Hines ◽  
Jeffrey A. Jeddeloh
Keyword(s):  
Quorum Sensing ◽  
Virulence Factor ◽  
Regulatory System ◽  
In Silico Analysis ◽  
Homoserine Lactone ◽  
Burkholderia Mallei ◽  
Wild Type ◽  
Transcriptional Regulatory ◽  
A Cell

ABSTRACT Numerous gram-negative bacterial pathogens regulate virulence factor expression by using a cell density mechanism termed quorum sensing (QS). An in silico analysis of the Burkholderia mallei ATCC 23344 genome revealed that it encodes at least two luxI and four luxR homologues. Using mass spectrometry, we showed that wild-type B. mallei produces the signaling molecules N-octanoyl-homoserine lactone and N-decanoyl-homoserine lactone. To determine if QS is involved in the virulence of B. mallei, we generated mutations in each putative luxIR homologue and tested the pathogenicities of the derivative strains in aerosol BALB/c mouse and intraperitoneal hamster models. Disruption of the B. mallei QS alleles, especially in RJ16 (bmaII) and RJ17 (bmaI3), which are luxI mutants, significantly reduced virulence, as indicated by the survival of mice who were aerosolized with 104 CFU (10 50% lethal doses [LD50s]). For the B. mallei transcriptional regulator mutants (luxR homologues), mutation of the bmaR5 allele resulted in the most pronounced decrease in virulence, with 100% of the challenged animals surviving a dose of 10 LD50s. Using a Syrian hamster intraperitoneal model of infection, we determined the LD50s for wild-type B. mallei and each QS mutant. An increase in the relative LD50 was found for RJ16 (bmaI1) (>967 CFU), RJ17 (bmaI3) (115 CFU), and RJ20 (bmaR5) (151 CFU) compared to wild-type B. mallei (<13 CFU). These findings demonstrate that B. mallei carries multiple luxIR homologues that either directly or indirectly regulate the biosynthesis of an essential virulence factor(s) that contributes to the pathogenicity of B. mallei in vivo.

scholarly journals Quorum Sensing Is a Global Regulatory Mechanism in Enterohemorrhagic Escherichia coli O157:H7

2001 ◽  
Vol 183 (17) ◽  
pp. 5187-5197 ◽  
Author(s):  
Vanessa Sperandio ◽  
Alfredo G. Torres ◽  
Jorge A. Girón ◽  
James B. Kaper
Keyword(s):  
Escherichia Coli ◽  
Quorum Sensing ◽  
Type Strain ◽  
Wild Type Strain ◽  
Regulatory Mechanism ◽  
Sos Response ◽  
Enterohemorrhagic Escherichia Coli ◽  
Trna Genes ◽  
Wild Type ◽  
E Coli

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is responsible for outbreaks of bloody diarrhea and hemolytic-uremic syndrome in many countries. EHEC virulence mechanisms include the production of Shiga toxins (Stx) and formation of attaching and effacing (AE) lesions on intestinal epithelial cells. We recently reported that genes involved in the formation of the AE lesion were regulated by quorum sensing through autoinducer-2, which is synthesized by the product of the luxS gene. In this study we hybridized an E. coli gene array with cDNA synthesized from RNA that was extracted from EHEC strain 86-24 and its isogenicluxS mutant. We observed that 404 genes were regulated by luxS at least fivefold, which comprises approximately 10% of the array genes; 235 of these genes were up-regulated and 169 were down-regulated in the wild-type strain compared to in theluxS mutant. Down-regulated genes included several involved in cell division, as well as ribosomal and tRNA genes. Consistent with this pattern of gene expression, theluxS mutant grows faster than the wild-type strain (generation times of 37.5 and 60 min, respectively, in Dulbecco modified Eagle medium). Up-regulated genes included several involved in the expression and assembly of flagella, motility, and chemotaxis. Using operon::lacZ fusions to class I, II, and III flagellar genes, we were able to confirm this transcriptional regulation. We also observed fewer flagella by Western blotting and electron microscopy and decreased motility halos in semisolid agar in the luxS mutant. The average swimming speeds for the wild-type strain and the luxS mutant are 12.5 and 6.6 μm/s, respectively. We also observed an increase in the production of Stx due to quorum sensing. Genes encoding Stx, which are transcribed along with λ-like phage genes, are induced by an SOS response, and genes involved in the SOS response were also regulated by quorum sensing. These results indicate that quorum sensing is a global regulatory mechanism for basic physiological functions of E. coli as well as for virulence factors.

Isolation of a chitinase overproducing mutant of Streptomyces peucetius defective in daunorubicin biosynthesis

2000 ◽  
Vol 46 (10) ◽  
pp. 956-960 ◽  
Author(s):  
Kuzhandhaivel S Vetrivel ◽  
Kuppamuthu Dharmalingam
Keyword(s):  
Biosynthetic Pathway ◽  
Time Course ◽  
Polyketide Synthase ◽  
Protein Profile ◽  
Restriction Enzymes ◽  
Wild Type ◽  
Anthracycline Antibiotic ◽  
Streptomyces Peucetius ◽  
Chitinase Production ◽  
Major Alteration

Streptomyces peucetius, producer of the antitumor anthracycline antibiotic daunorubicin, was mutagenized, and mutants defective in daunorubicin biosynthesis were screened. One mutant (SPVI), which failed to produce daunorubicin, was found to overproduce an extracellular chitinase. Time course analyses of chitinase production and of the extracellular protein profile showed that the increase in activity is due to increased synthesis of the enzyme protein. The production of chitinase in SPVI was repressed by glucose as in the case of wild-type S. peucetius. PFGE analysis of VspI restriction fragments of S. peucetius and SPVI showed that there was no major alteration in the mutant genome. The hybridization pattern of S. peucetius and SPVI genomic DNA digested with various restriction enzymes was identical when probed with dnrUVJI genes of the S. peucetius daunorubicin cluster and chiA of Streptomyces lividans 66. The possible step affected in the daunorubicin biosynthetic pathway could be a polyketide synthase, since aklanonic acid, the earliest detectable intermediate in the daunorubicin pathway, was not synthesized in SPVI.Key words: Streptomyces peucetius, chitinase, daunorubicin, NTG mutagenesis.

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