Publications by authors named "Mahmoud M Al-Bassam"

20 Publications

  • Page 1 of 1

Competition-based screening helps to secure the evolutionary stability of a defensive microbiome.

BMC Biol 2021 09 15;19(1):205. Epub 2021 Sep 15.

School of Biological Sciences, Norwich Research Park, University of East Anglia, Norwich, Norfolk, NR4 7TJ, UK.

Background: The cuticular microbiomes of Acromyrmex leaf-cutting ants pose a conundrum in microbiome biology because they are freely colonisable, and yet the prevalence of the vertically transmitted bacteria Pseudonocardia, which contributes to the control of Escovopsis fungus garden disease, is never compromised by the secondary acquisition of other bacterial strains. Game theory suggests that competition-based screening can allow the selective recruitment of antibiotic-producing bacteria from the environment, by providing abundant resources to foment interference competition between bacterial species and by using Pseudonocardia to bias the outcome of competition in favour of antibiotic producers.

Results: Here, we use RNA-stable isotope probing (RNA-SIP) to confirm that Acromyrmex ants can maintain a range of microbial symbionts on their cuticle by supplying public resources. We then used RNA sequencing, bioassays, and competition experiments to show that vertically transmitted Pseudonocardia strains produce antibacterials that differentially reduce the growth rates of other microbes, ultimately biassing the bacterial competition to allow the selective establishment of secondary antibiotic-producing strains while excluding non-antibiotic-producing strains that would parasitise the symbiosis.

Conclusions: Our findings are consistent with the hypothesis that competition-based screening is a plausible mechanism for maintaining the integrity of the co-adapted mutualism between the leaf-cutting ant farming symbiosis and its defensive microbiome. Our results have broader implications for explaining the stability of other complex symbioses involving horizontal acquisition.
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http://dx.doi.org/10.1186/s12915-021-01142-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8444595PMC
September 2021

Specialized and shared functions of diguanylate cyclases and phosphodiesterases in Streptomyces development.

Mol Microbiol 2020 11 14;114(5):808-822. Epub 2020 Aug 14.

Department of Biology/Microbiology, Humboldt-Universität zu Berlin, Berlin, Germany.

The second messenger bis-3,5-cyclic di-guanosine monophosphate (c-di-GMP) determines when Streptomyces initiate sporulation. c-di-GMP signals are integrated into the genetic differentiation network by the regulator BldD and the sigma factor σ . However, functions of the development-specific diguanylate cyclases (DGCs) CdgB and CdgC, and the c-di-GMP phosphodiesterases (PDEs) RmdA and RmdB, are poorly understood. Here, we provide biochemical evidence that the GGDEF-EAL domain protein RmdB from S. venezuelae is a monofunctional PDE that hydrolyzes c-di-GMP to 5'pGpG. Despite having an equivalent GGDEF-EAL domain arrangement, RmdA cleaves c-di-GMP to GMP and exhibits residual DGC activity. We show that an intact EAL motif is crucial for the in vivo function of both enzymes since strains expressing protein variants with an AAA motif instead of EAL are delayed in development, similar to null mutants. Transcriptome analysis of ∆cdgB, ∆cdgC, ∆rmdA, and ∆rmdB strains revealed that the c-di-GMP specified by these enzymes has a global regulatory role, with about 20% of all S. venezuelae genes being differentially expressed in the cdgC mutant. Our data suggest that the major c-di-GMP-controlled targets determining the timing and mode of sporulation are genes involved in cell division and the production of the hydrophobic sheath that covers Streptomyces aerial hyphae and spores.
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http://dx.doi.org/10.1111/mmi.14581DOI Listing
November 2020

Developmentally regulated volatiles geosmin and 2-methylisoborneol attract a soil arthropod to Streptomyces bacteria promoting spore dispersal.

Nat Microbiol 2020 06 6;5(6):821-829. Epub 2020 Apr 6.

Department of Biology, Lund University, Lund, Sweden.

Volatile compounds emitted by bacteria are often sensed by other organisms as odours, but their ecological roles are poorly understood. Well-known examples are the soil-smelling terpenoids geosmin and 2-methylisoborneol (2-MIB), which humans and various animals sense at extremely low concentrations. The conservation of geosmin biosynthesis genes among virtually all species of Streptomyces bacteria (and genes for the biosynthesis of 2-MIB in about 50%), suggests that the volatiles provide a selective advantage for these soil microbes. We show, in the present study, that these volatiles mediate interactions of apparent mutual benefit between streptomycetes and springtails (Collembola). In field experiments, springtails were attracted to odours emitted by Streptomyces colonies. Geosmin and 2-MIB in these odours induce electrophysiological responses in the antennae of the model springtail Folsomia candida, which is also attracted to both compounds. Moreover, the genes for geosmin and 2-MIB synthases are under the direct control of sporulation-specific transcription factors, constraining emission of the odorants to sporulating colonies. F. candida feeds on the Streptomyces colonies and disseminates spores both via faecal pellets and through adherence to its hydrophobic cuticle. The results indicate that geosmin and 2-MIB production is an integral part of the sporulation process, completing the Streptomyces life cycle by facilitating dispersal of spores by soil arthropods.
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http://dx.doi.org/10.1038/s41564-020-0697-xDOI Listing
June 2020

c-di-AMP hydrolysis by the phosphodiesterase AtaC promotes differentiation of multicellular bacteria.

Proc Natl Acad Sci U S A 2020 03 18;117(13):7392-7400. Epub 2020 Mar 18.

Department of Microbiology, Humboldt-Universität zu Berlin, 10115 Berlin, Germany;

Antibiotic-producing use the diadenylate cyclase DisA to synthesize the nucleotide second messenger c-di-AMP, but the mechanism for terminating c-di-AMP signaling and the proteins that bind the molecule to effect signal transduction are unknown. Here, we identify the AtaC protein as a c-di-AMP-specific phosphodiesterase that is also conserved in pathogens such as and AtaC is monomeric in solution and binds Mn to specifically hydrolyze c-di-AMP to AMP via the intermediate 5'-pApA. As an effector of c-di-AMP signaling, we characterize the RCK_C domain protein CpeA. c-di-AMP promotes interaction between CpeA and the predicted cation/proton antiporter, CpeB, linking c-di-AMP signaling to ion homeostasis in Actinobacteria. Hydrolysis of c-di-AMP is critical for normal growth and differentiation in , connecting ionic stress to development. Thus, we present the discovery of two components of c-di-AMP signaling in bacteria and show that precise control of this second messenger is essential for ion balance and coordinated development in .
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http://dx.doi.org/10.1073/pnas.1917080117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7132281PMC
March 2020

Author Correction: Environmental stimuli drive a transition from cooperation to competition in synthetic phototrophic communities.

Nat Microbiol 2019 Dec;4(12):2578

Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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http://dx.doi.org/10.1038/s41564-019-0621-4DOI Listing
December 2019

Environmental stimuli drive a transition from cooperation to competition in synthetic phototrophic communities.

Nat Microbiol 2019 12 7;4(12):2184-2191. Epub 2019 Oct 7.

Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA.

Phototrophic communities of photosynthetic algae or cyanobacteria and heterotrophic bacteria or fungi are pervasive throughout the environment. How interactions between members contribute to the resilience and affect the fitness of phototrophic communities is not fully understood. Here, we integrated metatranscriptomics, metabolomics and phenotyping with computational modelling to reveal condition-dependent secretion and cross-feeding of metabolites in a synthetic community. We discovered that interactions between members are highly dynamic and are driven by the availability of organic and inorganic nutrients. Environmental factors, such as ammonia concentration, influenced community stability by shifting members from collaborating to competing. Furthermore, overall fitness was dependent on genotype and streamlined genomes improved growth of the entire community. Our mechanistic framework provides insights into the physiology and metabolic response to environmental and genetic perturbation of these ubiquitous microbial associations.
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http://dx.doi.org/10.1038/s41564-019-0567-6DOI Listing
December 2019

Functional and Proteomic Analysis of Virulence Upon Loss of Its Native Cas9 Nuclease.

Front Microbiol 2019 22;10:1967. Epub 2019 Aug 22.

Division of Host-Microbe Systems and Therapeutics, Department of Pediatrics, School of Medicine, University of California, San Diego, La Jolla, CA, United States.

The public health impact of (group A , GAS) as a top 10 cause of infection-related mortality in humans contrasts with its benefit to biotechnology as the main natural source of Cas9 nuclease, the key component of the revolutionary CRISPR-Cas9 gene editing platform. Despite widespread knowledge acquired in the last decade on the molecular mechanisms by which GAS Cas9 achieves precise DNA targeting, the functions of Cas9 in the biology and pathogenesis of its native organism remain unknown. In this study, we generated an isogenic serotype M1 GAS mutant deficient in Cas9 protein and compared its behavior and phenotypes to the wild-type parent strain. Absence of Cas9 was linked to reduced GAS epithelial cell adherence, reduced growth in human whole blood , and attenuation of virulence in a murine necrotizing skin infection model. Virulence defects of the GAS Δ strain were explored through quantitative proteomic analysis, revealing a significant reduction in the abundance of key GAS virulence determinants. Similarly, deletion of affected the expression of several known virulence regulatory proteins, indicating that Cas9 impacts the global architecture of GAS gene regulation.
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http://dx.doi.org/10.3389/fmicb.2019.01967DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6714885PMC
August 2019

Sensing and responding to diverse extracellular signals: an updated analysis of the sensor kinases and response regulators of species.

Microbiology (Reading) 2019 09;165(9):929-952

School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich, Norfolk NR4 7TJ, UK.

is a Gram-positive, filamentous actinomycete with a complex developmental life cycle. Genomic analysis revealed that encodes a large number of two-component systems (TCSs): these consist of a membrane-bound sensor kinase (SK) and a cognate response regulator (RR). These proteins act together to detect and respond to diverse extracellular signals. Some of these systems have been shown to regulate antimicrobial biosynthesis in species, making them very attractive to researchers. The ability of to sporulate in both liquid and solid cultures has made it an increasingly popular model organism in which to study these industrially and medically important bacteria. Bioinformatic analysis identified 58 TCS operons in with an additional 27 orphan SK and 18 orphan RR genes. A broader approach identified 15 of the 58 encoded TCSs to be highly conserved in 93 species for which high-quality and complete genome sequences are available. This review attempts to unify the current work on TCS in the streptomycetes, with an emphasis on .
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http://dx.doi.org/10.1099/mic.0.000817DOI Listing
September 2019

Predicting proteome allocation, overflow metabolism, and metal requirements in a model acetogen.

PLoS Comput Biol 2019 03 7;15(3):e1006848. Epub 2019 Mar 7.

Department of Pediatrics, University of California, San Diego, La Jolla, California, United States of America.

The unique capability of acetogens to ferment a broad range of substrates renders them ideal candidates for the biotechnological production of commodity chemicals. In particular the ability to grow with H2:CO2 or syngas (a mixture of H2/CO/CO2) makes these microorganisms ideal chassis for sustainable bioproduction. However, advanced design strategies for acetogens are currently hampered by incomplete knowledge about their physiology and our inability to accurately predict phenotypes. Here we describe the reconstruction of a novel genome-scale model of metabolism and macromolecular synthesis (ME-model) to gain new insights into the biology of the model acetogen Clostridium ljungdahlii. The model represents the first ME-model of a Gram-positive bacterium and captures all major central metabolic, amino acid, nucleotide, lipid, major cofactors, and vitamin synthesis pathways as well as pathways to synthesis RNA and protein molecules necessary to catalyze these reactions, thus significantly broadens the scope and predictability. Use of the model revealed how protein allocation and media composition influence metabolic pathways and energy conservation in acetogens and accurately predicted secretion of multiple fermentation products. Predicting overflow metabolism is of particular interest since it enables new design strategies, e.g. the formation of glycerol, a novel product for C. ljungdahlii, thus broadening the metabolic capability for this model microbe. Furthermore, prediction and experimental validation of changing secretion rates based on different metal availability opens the window into fermentation optimization and provides new knowledge about the proteome utilization and carbon flux in acetogens.
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http://dx.doi.org/10.1371/journal.pcbi.1006848DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6430413PMC
March 2019

BldC Delays Entry into Development To Produce a Sustained Period of Vegetative Growth in Streptomyces venezuelae.

mBio 2019 02 5;10(1). Epub 2019 Feb 5.

Department of Molecular Microbiology, John Innes Centre, Norwich, United Kingdom

Streptomycetes are filamentous bacteria that differentiate by producing spore-bearing reproductive structures called aerial hyphae. The transition from vegetative to reproductive growth is controlled by the (bald) loci, and mutations in genes prevent the formation of aerial hyphae, either by blocking entry into development (typically mutations in activators) or by inducing precocious sporulation in the vegetative mycelium (typically mutations in repressors). One of the genes, , encodes a 68-residue DNA-binding protein related to the DNA-binding domain of MerR-family transcription factors. Recent work has shown that BldC binds DNA by a novel mechanism, but there is less insight into its impact on development. Here we used ChIP-seq coupled with RNA-seq to define the BldC regulon in the model species , showing that BldC can function both as a repressor and as an activator of transcription. Using electron microscopy and time-lapse imaging, we show that mutants are bald because they initiate development prematurely, bypassing the formation of aerial hyphae. This is consistent with the premature expression of BldC target genes encoding proteins with key roles in development (e.g., , , ), chromosome condensation and segregation (e.g., , ), and sporulation-specific cell division (e.g., ), suggesting that BldC-mediated repression is critical to maintain a sustained period of vegetative growth prior to sporulation. We discuss the possible significance of BldC as an evolutionary link between MerR family transcription factors and DNA architectural proteins. Understanding the mechanisms that drive bacterial morphogenesis depends on the dissection of the regulatory networks that underpin the cell biological processes involved. Recently, has emerged as an attractive model system for the study of morphological differentiation in This has led to significant progress in identifying the genes controlled by the transcription factors that regulate aerial mycelium formation (Bld regulators) and sporulation (Whi regulators). Taking advantage of , we used ChIP-seq coupled with RNA-seq to identify the genes directly under the control of BldC. Because sporulates in liquid culture, the complete spore-to-spore life cycle can be examined using time-lapse microscopy, and we applied this technique to the mutant. These combined approaches reveal BldC to be a member of an emerging class of Bld regulators that function principally to repress key sporulation genes, thereby extending vegetative growth and blocking the onset of morphological differentiation.
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http://dx.doi.org/10.1128/mBio.02812-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6428758PMC
February 2019

Expression Patterns, Genomic Conservation and Input Into Developmental Regulation of the GGDEF/EAL/HD-GYP Domain Proteins in .

Front Microbiol 2018 23;9:2524. Epub 2018 Oct 23.

Institut für Biologie/Mikrobiologie, Humboldt-Universität zu Berlin, Berlin, Germany.

To proliferate, antibiotic-producing undergo a complex developmental transition from vegetative growth to the production of aerial hyphae and spores. This morphological switch is controlled by the signaling molecule cyclic bis-(3',5') di-guanosine-mono-phosphate (c-di-GMP) that binds to the master developmental regulator, BldD, leading to repression of key sporulation genes during vegetative growth. However, a systematical analysis of all the GGDEF/EAL/HD-GYP proteins that control c-di-GMP levels in is still lacking. Here, we have FLAG-tagged all 10 c-di-GMP turnover proteins in and characterized their expression patterns throughout the life cycle, revealing that the diguanylate cyclase (DGC) CdgB and the phosphodiesterase (PDE) RmdB are the most abundant GGDEF/EAL proteins. Moreover, we have deleted all the genes coding for c-di-GMP turnover enzymes individually and analyzed morphogenesis of the mutants in macrocolonies. We show that the composite GGDEF-EAL protein CdgC is an active DGC and that deletion of the DGCs and enhance sporulation whereas deletion of the PDEs and delay development in . By comparing the pan genome of 93 fully sequenced species we show that the DGCs CdgA, CdgB, and CdgC, and the PDE RmdB represent the most conserved c-di-GMP-signaling proteins in the genus .
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http://dx.doi.org/10.3389/fmicb.2018.02524DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6205966PMC
October 2018

Optimization of carbon and energy utilization through differential translational efficiency.

Nat Commun 2018 10 26;9(1):4474. Epub 2018 Oct 26.

Department of Pediatrics, Division of Host-Microbe Systems and Therapeutics, University of California San Diego, 9500 Gilman Drive, La Jolla, CA, 92093, USA.

Control of translation is vital to all species. Here we employ a multi-omics approach to decipher condition-dependent translational regulation in the model acetogen Clostridium ljungdahlii. Integration of data from cells grown autotrophically or heterotrophically revealed that pathways critical to carbon and energy metabolism are under strong translational regulation. Major pathways involved in carbon and energy metabolism are not only differentially transcribed and translated, but their translational efficiencies are differentially elevated in response to resource availability under different growth conditions. We show that translational efficiency is not static and that it changes dynamically in response to mRNA expression levels. mRNAs harboring optimized 5'-untranslated region and coding region features, have higher translational efficiencies and are significantly enriched in genes encoding carbon and energy metabolism. In contrast, mRNAs enriched in housekeeping functions harbor sub-optimal features and have lower translational efficiencies. We propose that regulation of translational efficiency is crucial for effectively controlling resource allocation in energy-deprived microorganisms.
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http://dx.doi.org/10.1038/s41467-018-06993-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6203783PMC
October 2018

Group B Streptococcus Biofilm Regulatory Protein A Contributes to Bacterial Physiology and Innate Immune Resistance.

J Infect Dis 2018 10;218(10):1641-1652

Division of Host-Microbe Systems and Therapeutics, Department of Pediatrics, University of California-San Diego, La Jolla.

Background: Streptococcus agalactiae (group B Streptococcus [GBS]) asymptomatically colonizes approximately 20% of adults; however, GBS causes severe disease in susceptible populations, including newborns, pregnant women, and elderly individuals. In shifting between commensal and pathogenic states, GBS reveals multiple mechanisms of virulence factor control. Here we describe a GBS protein that we named "biofilm regulatory protein A" (BrpA) on the basis of its homology with BrpA from Streptococcus mutans.

Methods: We coupled phenotypic assays, RNA sequencing, human neutrophil and whole-blood killing assays, and murine infection models to investigate the contribution of BrpA to GBS physiology and virulence.

Results: Sequence analysis identified BrpA as a LytR-CpsA-Psr enzyme. Targeted mutagenesis yielded a GBS mutant (ΔbrpA) with normal ultrastructural morphology but a 6-fold increase in chain length, a biofilm defect, and decreased acid tolerance. GBS ΔbrpA stimulated increased neutrophil reactive oxygen species and proved more susceptible to human and murine blood and neutrophil killing. Notably, the wild-type parent outcompeted ΔbrpA GBS in murine sepsis and vaginal colonization models. RNA sequencing of ΔbrpA uncovered multiple differences from the wild-type parent, including pathways of cell wall synthesis and cellular metabolism.

Conclusions: We propose that BrpA is an important virulence regulator and potential target for design of novel antibacterial therapeutics against GBS.
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http://dx.doi.org/10.1093/infdis/jiy341DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6173572PMC
October 2018

Watasemycin biosynthesis in : thiazoline C-methylation by a type B radical-SAM methylase homologue.

Chem Sci 2017 Apr 19;8(4):2823-2831. Epub 2017 Jan 19.

Department of Chemistry , University of Warwick , Coventry , CV4 7AL , UK . Email:

2-Hydroxyphenylthiazolines are a family of iron-chelating nonribosomal peptide natural products that function as virulence-conferring siderophores in various Gram-negative bacteria. They have also been reported as metabolites of Gram-positive species. Transcriptional analyses of ATCC 10712 revealed that its genome contains a putative 2-hydroxyphenylthiazoline biosynthetic gene cluster. Heterologous expression of the gene cluster in M1152 showed that the mono- and dimethylated derivatives, thiazostatin and watasemycin, respectively, of the 2-hydroxyphenylthiazoline enantiopyochelin are two of its metabolic products. In addition, isopyochelin, a novel isomer of pyochelin containing a C-methylated thiazolidine, was identified as a third metabolic product of the cluster. Metabolites with molecular formulae corresponding to aerugine and pulicatins A/B were also detected. The structure and stereochemistry of isopyochelin were confirmed by comparison with synthetic standards. The role of two genes in the cluster encoding homologues of PchK, which is proposed to catalyse thiazoline reduction in the biosynthesis of enantiopyochelin in , was investigated. One was required for the production of all the metabolic products of the cluster, whereas the other appears not to be involved in the biosynthesis of any of them. Deletion of a gene in the cluster encoding a type B radical-SAM methylase homologue abolished the production of watasemycin, but not thiazostatin or isopyochelin. Feeding of thiazostatin to the mutant lacking the functional PchK homologue resulted in complete conversion to watasemycin, demonstrating that thiazoline C-methylation by the type B radical-SAM methylase homologue is the final step in watasemycin biosynthesis.
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http://dx.doi.org/10.1039/c6sc03533gDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5427693PMC
April 2017

Networks of energetic and metabolic interactions define dynamics in microbial communities.

Proc Natl Acad Sci U S A 2015 Dec 30;112(50):15450-5. Epub 2015 Nov 30.

Department of Bioengineering, University of California, San Diego, CA 92091

Microorganisms form diverse communities that have a profound impact on the environment and human health. Recent technological advances have enabled elucidation of community diversity at high resolution. Investigation of microbial communities has revealed that they often contain multiple members with complementing and seemingly redundant metabolic capabilities. An understanding of the communal impacts of redundant metabolic capabilities is currently lacking; specifically, it is not known whether metabolic redundancy will foster competition or motivate cooperation. By investigating methanogenic populations, we identified the multidimensional interspecies interactions that define composition and dynamics within syntrophic communities that play a key role in the global carbon cycle. Species-specific genomes were extracted from metagenomic data using differential coverage binning. We used metabolic modeling leveraging metatranscriptomic information to reveal and quantify a complex intertwined system of syntrophic relationships. Our results show that amino acid auxotrophies create additional interdependencies that define community composition and control carbon and energy flux through the system while simultaneously contributing to overall community robustness. Strategic use of antimicrobials further reinforces this intricate interspecies network. Collectively, our study reveals the multidimensional interactions in syntrophic communities that promote high species richness and bolster community stability during environmental perturbations.
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http://dx.doi.org/10.1073/pnas.1506034112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4687543PMC
December 2015

NsrR from Streptomyces coelicolor is a nitric oxide-sensing [4Fe-4S] cluster protein with a specialized regulatory function.

J Biol Chem 2015 May 14;290(20):12689-704. Epub 2015 Mar 14.

From the Centre for Molecular and Structural Biochemistry, School of Chemistry,

The Rrf2 family transcription factor NsrR controls expression of genes in a wide range of bacteria in response to nitric oxide (NO). The precise form of the NO-sensing module of NsrR is the subject of controversy because NsrR proteins containing either [2Fe-2S] or [4Fe-4S] clusters have been observed previously. Optical, Mössbauer, resonance Raman spectroscopies and native mass spectrometry demonstrate that Streptomyces coelicolor NsrR (ScNsrR), previously reported to contain a [2Fe-2S] cluster, can be isolated containing a [4Fe-4S] cluster. ChIP-seq experiments indicated that the ScNsrR regulon is small, consisting of only hmpA1, hmpA2, and nsrR itself. The hmpA genes encode NO-detoxifying flavohemoglobins, indicating that ScNsrR has a specialized regulatory function focused on NO detoxification and is not a global regulator like some NsrR orthologues. EMSAs and DNase I footprinting showed that the [4Fe-4S] form of ScNsrR binds specifically and tightly to an 11-bp inverted repeat sequence in the promoter regions of the identified target genes and that DNA binding is abolished following reaction with NO. Resonance Raman data were consistent with cluster coordination by three Cys residues and one oxygen-containing residue, and analysis of ScNsrR variants suggested that highly conserved Glu-85 may be the fourth ligand. Finally, we demonstrate that some low molecular weight thiols, but importantly not physiologically relevant thiols, such as cysteine and an analogue of mycothiol, bind weakly to the [4Fe-4S] cluster, and exposure of this bound form to O2 results in cluster conversion to the [2Fe-2S] form, which does not bind to DNA. These data help to account for the observation of [2Fe-2S] forms of NsrR.
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http://dx.doi.org/10.1074/jbc.M115.643072DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4432287PMC
May 2015

New insights into chloramphenicol biosynthesis in Streptomyces venezuelae ATCC 10712.

Antimicrob Agents Chemother 2014 Dec 29;58(12):7441-50. Epub 2014 Sep 29.

Department of Molecular Microbiology, John Innes Centre, Norwich Research Park, Norwich, United Kingdom

Comparative genome analysis revealed seven uncharacterized genes, sven0909 to sven0915, adjacent to the previously identified chloramphenicol biosynthetic gene cluster (sven0916-sven0928) of Streptomyces venezuelae strain ATCC 10712 that was absent in a closely related Streptomyces strain that does not produce chloramphenicol. Transcriptional analysis suggested that three of these genes might be involved in chloramphenicol production, a prediction confirmed by the construction of deletion mutants. These three genes encode a cluster-associated transcriptional activator (Sven0913), a phosphopantetheinyl transferase (Sven0914), and a Na(+)/H(+) antiporter (Sven0915). Bioinformatic analysis also revealed the presence of a previously undetected gene, sven0925, embedded within the chloramphenicol biosynthetic gene cluster that appears to encode an acyl carrier protein, bringing the number of new genes likely to be involved in chloramphenicol production to four. Microarray experiments and synteny comparisons also suggest that sven0929 is part of the biosynthetic gene cluster. This has allowed us to propose an updated and revised version of the chloramphenicol biosynthetic pathway.
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http://dx.doi.org/10.1128/AAC.04272-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4249514PMC
December 2014

Response regulator heterodimer formation controls a key stage in Streptomyces development.

PLoS Genet 2014 Aug 7;10(8):e1004554. Epub 2014 Aug 7.

Department of Molecular Microbiology, John Innes Centre, Norwich Research Park, Colney Lane, Norwich, United Kingdom.

The orphan, atypical response regulators BldM and WhiI each play critical roles in Streptomyces differentiation. BldM is required for the formation of aerial hyphae, and WhiI is required for the differentiation of these reproductive structures into mature spores. To gain insight into BldM function, we defined the genome-wide BldM regulon using ChIP-Seq and transcriptional profiling. BldM target genes clustered into two groups based on their whi gene dependency. Expression of Group I genes depended on bldM but was independent of all the whi genes, and biochemical experiments showed that Group I promoters were controlled by a BldM homodimer. In contrast, Group II genes were expressed later than Group I genes and their expression depended not only on bldM but also on whiI and whiG (encoding the sigma factor that activates whiI). Additional ChIP-Seq analysis showed that BldM Group II genes were also direct targets of WhiI and that in vivo binding of WhiI to these promoters depended on BldM and vice versa. We go on to demonstrate that BldM and WhiI form a functional heterodimer that controls Group II promoters, serving to integrate signals from two distinct developmental pathways. The BldM-WhiI system thus exemplifies the potential of response regulator heterodimer formation as a mechanism to expand the signaling capabilities of bacterial cells.
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http://dx.doi.org/10.1371/journal.pgen.1004554DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4125116PMC
August 2014

Household size is critical to varicella-zoster virus transmission in the tropics despite lower viral infectivity.

Epidemics 2011 Mar 7;3(1):12-8. Epub 2010 Dec 7.

School of Biological and Chemical Sciences, Queen Mary University of London, UK.

The epidemiology and severity of infections can vary dramatically in different geographical regions. Varicella zoster virus (VZV) is a particularly tractable model for investigating such global differences, since infections can be unambiguously identified. VZV is spread by aerosol to cause chickenpox, which, in temperate countries, is a relatively benign childhood infection; yet in tropical countries it tends to occur at later age, a trend associated with markedly increased severity including complications, hospitalization, and overall burden of care. To investigate global differences in the epidemiology of chickenpox we studied a population in Guinea Bissau, which in contrast to other tropical countries has an unexpectedly early age of infection with VZV, comparable to temperate latitudes. In this study we used detailed records from over 3000 houses during an outbreak of chickenpox, combined with viral genetic information on routes of infection, to obtain precise estimates of disease transmission within and between houses. This community contains many large households in which different families live under a single roof, in living quarters divided by partitions. Our data show that household infectivity in tropical Guinea Bissau is reduced four-fold compared with temperate climates (14.8% versus 61-85%), with an intermediate rate between members of the same family who are in more intimate contact (23.5%). All else being equal, these lower infection rates would be expected to lead to a later age of infection as is commonly seen in other tropical countries. The young age of infection, which had drawn our attention to the Guinea Bissau population, can however be explained by the exceptionally large household sizes (mean 14.5 people). We have combined genetic and demographic data to show that the epidemiology of chickenpox in tropical Guinea Bissau is dependent on the interaction of the social and physical environments. The distinctive clinical presentation of VZV and its ubiquitous distribution make it an attractive model for estimating the variables that contribute to global differences in the transmission of airborne viruses.
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http://dx.doi.org/10.1016/j.epidem.2010.11.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3072572PMC
March 2011

Natural selection for rash-forming genotypes of the varicella-zoster vaccine virus detected within immunized human hosts.

Proc Natl Acad Sci U S A 2007 Jan 20;104(1):208-12. Epub 2006 Dec 20.

Centre for Infectious Disease, Institute for Cell and Molecular Science, Queen Mary's School of Medicine and Dentistry, 4 Newark Street, London E1 2AT, United Kingdom.

The Oka vaccine strain is a live attenuated virus that is routinely administered to children in the United States and Europe to prevent chickenpox. It is effective and safe but occasionally produces a rash. The vaccine virus has accumulated mutations during its attenuation, but the rashes are not explained by their reversion, unlike complications reported for other viral vaccines. Indeed, most of the novel mutations distinguishing the Oka vaccine from the more virulent parental virus have not actually become fixed. Because the parental alleles are still present, the vaccine is polymorphic at >30 loci and therefore contains a mixture of related viruses. The inoculation of >40 million patients has consequently created a highly replicated evolutionary experiment that we have used to assess the competitive ability of these different viral genotypes in a human host. Using virus recovered from rash vesicles, we show that two vaccine mutations, causing amino acid substitutions in the major transactivating protein IE62, are outcompeted by the ancestral alleles. Standard interpretations of varicella disease severity concentrate on the undeniably important effects of host genotype and immune status, yet our results allow us to demonstrate that the viral genotype is associated with virulence and to identify the key sites. We propose that these loci have pleiotropic effects on the immunogenic properties of the virus, rash formation, and its epidemiological spread, which mould the evolution of its virulence. These findings are of practical importance for reducing the incidence of vaccine-associated rash and promoting public acceptance of the vaccine.
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http://dx.doi.org/10.1073/pnas.0605688104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1765436PMC
January 2007
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