scholarly journals The ParB-parS Chromosome Segregation System Modulates Competence Development in Streptococcus pneumoniae

mBio ◽  
2015 ◽  
Vol 6 (4) ◽  
Author(s):  
Laetitia Attaiech ◽  
Anita Minnen ◽  
Morten Kjos ◽  
Stephan Gruber ◽  
Jan-Willem Veening
Keyword(s):  
Streptococcus Pneumoniae ◽  
Chromosome Segregation ◽  
Dna Sequences ◽  
Molecular Mechanisms ◽  
Developmental Process ◽  
Competence Development ◽  
Human Pathogen ◽  
Transcription Analysis ◽  
Exogenous Dna ◽  
Content Type

ABSTRACT ParB proteins bind centromere-like DNA sequences called parS sites and are involved in plasmid and chromosome segregation in bacteria. We previously showed that the opportunistic human pathogen Streptococcus pneumoniae contains four parS sequences located close to the origin of replication which are bound by ParB. Using chromatin immunoprecipitation (ChIP), we found here that ParB spreads out from one of these parS sites, parS(−1.6°), for more than 5 kb and occupies the nearby comCDE operon, which drives competence development. Competence allows S. pneumoniae to take up DNA from its environment, thereby mediating horizontal gene transfer, and is also employed as a general stress response. Mutating parS(−1.6°) or deleting parB resulted in transcriptional up-regulation of comCDE and ssbB (a gene belonging to the competence regulon), demonstrating that ParB acts as a repressor of competence. However, genome-wide transcription analysis showed that ParB is not a global transcriptional regulator. Different factors, such as the composition of the growth medium and antibiotic-induced stress, can trigger the sensitive switch driving competence. This work shows that the ParB-parS chromosome segregation machinery also influences this developmental process. IMPORTANCE Streptococcus pneumoniae (pneumococcus) is an important human pathogen responsible for more than a million deaths each year. Like all other organisms, S. pneumoniae must be able to segregate its chromosomes properly. Not only is understanding the molecular mechanisms underlying chromosome segregation in S. pneumoniae therefore of fundamental importance, but also, this knowledge might offer new leads for ways to target this pathogen. Here, we identified a link between the pneumococcal chromosome segregation system and the competence-developmental system. Competence allows S. pneumoniae to take up and integrate exogenous DNA in its chromosome. This process plays a crucial role in successful adaptation to—and escape from—host defenses, antibiotic treatments, and vaccination strategies. We show that the chromosome segregation protein ParB acts as a repressor of competence. To the best of our knowledge, this is the first example of a ParB protein controlling bacterial competence.

scholarly journals Refining the Pneumococcal Competence Regulon by RNA Sequencing

2019 ◽  
Vol 201 (13) ◽  
Author(s):  
Jelle Slager ◽  
Rieza Aprianto ◽  
Jan-Willem Veening
Keyword(s):  
Antibiotic Resistance ◽  
Streptococcus Pneumoniae ◽  
Rna Sequencing ◽  
Genome Annotation ◽  
Stress Factors ◽  
Microarray Technology ◽  
Human Pathogen ◽  
Exogenous Dna ◽  
Content Type ◽  
Opportunistic Human Pathogen

ABSTRACTCompetence for genetic transformation allows the opportunistic human pathogenStreptococcus pneumoniaeto take up exogenous DNA for incorporation into its own genome. This ability may account for the extraordinary genomic plasticity of this bacterium, leading to antigenic variation, vaccine escape, and the spread of antibiotic resistance. The competence system has been thoroughly studied, and its regulation is well understood. Additionally, over the last decade, several stress factors have been shown to trigger the competent state, leading to the activation of several stress response regulons. The arrival of next-generation sequencing techniques allowed us to update the competence regulon, the latest report on which still depended on DNA microarray technology. Enabled by the availability of an up-to-date genome annotation, including transcript boundaries, we assayed time-dependent expression of all annotated features in response to competence induction, were able to identify the affected promoters, and produced a more complete overview of the various regulons activated during the competence state. We show that 4% of all annotated genes are under direct control of competence regulators ComE and ComX, while the expression of a total of up to 17% of all genes is affected, either directly or indirectly. Among the affected genes are various small RNAs with an as-yet-unknown function. Besides the ComE and ComX regulons, we were also able to refine the CiaR, VraR (LiaR), and BlpR regulons, underlining the strength of combining transcriptome sequencing (RNA-seq) with a well-annotated genome.IMPORTANCEStreptococcus pneumoniaeis an opportunistic human pathogen responsible for over a million deaths every year. Although both vaccination programs and antibiotic therapies have been effective in prevention and treatment of pneumococcal infections, respectively, the sustainability of these solutions is uncertain. The pneumococcal genome is highly flexible, leading to vaccine escape and antibiotic resistance. This flexibility is predominantly facilitated by competence, a state allowing the cell to take up and integrate exogenous DNA. Thus, it is essential to obtain a detailed overview of gene expression during competence. This is stressed by the fact that administration of several classes of antibiotics can lead to competence. Previous studies on the competence regulon were performed with microarray technology and were limited to an incomplete set of known genes. Using RNA sequencing combined with an up-to-date genome annotation, we provide an updated overview of competence-regulated genes.

scholarly journals Serum Albumin and Ca2+Are Natural Competence Inducers in the Human Pathogen Acinetobacter baumannii

2016 ◽  
Vol 60 (8) ◽  
pp. 4920-4929 ◽  
Author(s):  
German Matias Traglia ◽  
Brettni Quinn ◽  
Sareda T. J. Schramm ◽  
Alfonso Soler-Bistue ◽  
Maria Soledad Ramirez
Keyword(s):  
Acinetobacter Baumannii ◽  
Natural Transformation ◽  
Hospital Environment ◽  
Human Pathogen ◽  
Nosocomial Pathogen ◽  
Natural Competence ◽  
Exogenous Dna ◽  
Content Type ◽  
Resistance Determinants ◽  
Main Protein

ABSTRACTThe increasing frequency of bacteria showing antimicrobial resistance (AMR) raises the menace of entering into a postantibiotic era. Horizontal gene transfer (HGT) is one of the prime reasons for AMR acquisition.Acinetobacter baumanniiis a nosocomial pathogen with outstanding abilities to survive in the hospital environment and to acquire resistance determinants. Its capacity to incorporate exogenous DNA is a major source of AMR genes; however, few studies have addressed this subject. The transformation machinery as well as the factors that induce natural competence inA. baumanniiare unknown. In this study, we demonstrate that naturally competent strain A118 increases its natural transformation frequency upon the addition of Ca2+or albumin. We show thatcomEAandpilQare involved in this process since their expression levels are increased upon the addition of these compounds. An unspecific protein, like casein, does not reproduce this effect, showing that albumin's effect is specific. Our work describes the first specific inducers of natural competence inA. baumannii. Overall, our results suggest that the main protein in blood enhances HGT inA. baumannii, contributing to the increase of AMR in this threatening human pathogen.

scholarly journals Pathways of Pathogenicity: Transcriptional Stages of Germination in the Fatal Fungal PathogenRhizopus delemar

mSphere ◽  
2018 ◽  
Vol 3 (5) ◽  
Author(s):  
Poppy C. S. Sephton-Clark ◽  
Jose F. Muñoz ◽  
Elizabeth R. Ballou ◽  
Christina A. Cuomo ◽  
Kerstin Voelz
Keyword(s):  
Molecular Mechanisms ◽  
Developmental Stages ◽  
Developmental Process ◽  
Fungal Spores ◽  
Hyphal Growth ◽  
Large Shift ◽  
Rhizopus Delemar ◽  
Content Type ◽  
Transcriptional Changes ◽  
Dormant Spore

ABSTRACTRhizopus delemaris an invasive fungal pathogen responsible for the frequently fatal disease mucormycosis. Germination, a crucial mechanism by which infectious spores ofRhizopus delemarcause disease, is a key developmental process that transforms the dormant spore state into a vegetative one. The molecular mechanisms that underpin this transformation may be key to controlling mucormycosis; however, the regulation of germination remains poorly understood. This study describes the phenotypic and transcriptional changes that take place over the course of germination. This process is characterized by four distinct stages: dormancy, isotropic swelling, germ tube emergence, and hyphal growth. Dormant spores are shown to be transcriptionally unique, expressing a subset of transcripts absent in later developmental stages. A large shift in the expression profile is prompted by the initiation of germination, with genes involved in respiration, chitin, cytoskeleton, and actin regulation appearing to be important for this transition. A period of transcriptional consistency can be seen throughout isotropic swelling, before the transcriptional landscape shifts again at the onset of hyphal growth. This study provides a greater understanding of the regulation of germination and highlights processes involved in transformingRhizopus delemarfrom a single-cellular to multicellular organism.IMPORTANCEGermination is key to the growth of many organisms, including fungal spores. Mucormycete spores exist abundantly within the environment and germinate to form hyphae. These spores are capable of infecting immunocompromised individuals, causing the disease mucormycosis. Germination from spore to hyphae within patients leads to angioinvasion, tissue necrosis, and often fatal infections. This study advances our understanding of how spore germination occurs in the mucormycetes, identifying processes we may be able to inhibit to help prevent or treat mucormycosis.

scholarly journals Draft Genome Sequence of Pediatric Otitis Media Isolate Streptococcus pneumoniae Strain EF3030, Which Forms In Vitro Biofilms That Closely Mimic In Vivo Biofilms

2019 ◽  
Vol 8 (2) ◽  
Author(s):  
Edgar J. Scott ◽  
Nicole R. Luke-Marshall ◽  
Anthony A. Campagnari ◽  
David W. Dyer
Keyword(s):  
Streptococcus Pneumoniae ◽  
Otitis Media ◽  
Genome Sequence ◽  
Dna Sequences ◽  
Draft Genome ◽  
Draft Genome Sequence ◽  
Draft Assembly ◽  
Content Type

Here, we report the draft genome sequence of Streptococcus pneumoniae EF3030, a pediatric otitis media isolate active in biofilm assays of epithelial colonization. The final draft assembly included 2,209,198 bp; the annotation predicted 2,120 coding DNA sequences (CDSs), 4 complete rRNA operons, 58 tRNAs, 3 noncoding RNAs (ncRNAs), and 199 pseudogenes.

scholarly journals The Protein Kinase A-Dependent Phosphoproteome of the Human Pathogen Aspergillus fumigatus Reveals Diverse Virulence-Associated Kinase Targets

mBio ◽  
2020 ◽  
Vol 11 (6) ◽  
pp. e02880-20
Author(s):  
E. Keats Shwab ◽  
Praveen R. Juvvadi ◽  
Greg Waitt ◽  
Shareef Shaheen ◽  
John Allen ◽  
...  
Keyword(s):  
Infectious Disease ◽  
Protein Kinase ◽  
Aspergillus Fumigatus ◽  
Protein Kinase A ◽  
Molecular Mechanisms ◽  
Targeted Mutagenesis ◽  
Human Pathogen ◽  
Content Type ◽  
Kinase A ◽  
Microbial Disease

ABSTRACTProtein kinase A (PKA) signaling plays a critical role in the growth and development of all eukaryotic microbes. However, few direct targets have been characterized in any organism. The fungus Aspergillus fumigatus is a leading infectious cause of death in immunocompromised patients, but the specific molecular mechanisms responsible for its pathogenesis are poorly understood. We used this important pathogen as a platform for a comprehensive and multifaceted interrogation of both the PKA-dependent whole proteome and phosphoproteome in order to elucidate the mechanisms through which PKA signaling regulates invasive microbial disease. Employing advanced quantitative whole-proteomic and phosphoproteomic approaches with two complementary phosphopeptide enrichment strategies, coupled to an independent PKA interactome analysis, we defined distinct PKA-regulated pathways and identified novel direct PKA targets contributing to pathogenesis. We discovered three previously uncharacterized virulence-associated PKA effectors, including an autophagy-related protein, Atg24; a CCAAT-binding transcriptional regulator, HapB; and a CCR4-NOT complex-associated ubiquitin ligase, Not4. Targeted mutagenesis, combined with in vitro kinase assays, multiple murine infection models, structural modeling, and molecular dynamics simulations, was employed to characterize the roles of these new PKA targets in growth, environmental and antimicrobial stress responses, and pathogenesis in a mammalian system. We also elucidated the molecular mechanisms of PKA regulation for these effectors by defining the functionality of phosphorylation at specific PKA target sites. We have comprehensively characterized the PKA-dependent phosphoproteome and validated PKA targets as direct regulators of infectious disease for the first time in any pathogen, providing new insights into PKA signaling and control over microbial pathogenesis.IMPORTANCE PKA is essential for the virulence of eukaryotic human pathogens. Understanding PKA signaling mechanisms is therefore fundamental to deciphering pathogenesis and developing novel therapies. Despite its ubiquitous necessity, specific PKA effectors underlying microbial disease remain unknown. To address this fundamental knowledge gap, we examined the whole-proteomic and phosphoproteomic impacts of PKA on the deadly fungal pathogen Aspergillus fumigatus to uncover novel PKA targets controlling growth and virulence. We also defined the functional consequences of specific posttranslational modifications of these target proteins to characterize the molecular mechanisms of pathogenic effector regulation by PKA. This study constitutes the most comprehensive analysis of the PKA-dependent phosphoproteome of any human pathogen and proposes new and complex roles played by PKA signaling networks in governing infectious disease.

scholarly journals Pneumococcal 6-Phospho-β-Glucosidase (BglA3) Is Involved in Virulence and Nutrient Metabolism

2015 ◽  
Vol 84 (1) ◽  
pp. 286-292 ◽  
Author(s):  
Vanessa Sofia Terra ◽  
Xiangyun Zhi ◽  
Hasan F. Kahya ◽  
Peter W. Andrew ◽  
Hasan Yesilkaya
Keyword(s):  
Hyaluronic Acid ◽  
Streptococcus Pneumoniae ◽  
Cell Membrane ◽  
Human Pathogen ◽  
Wild Type ◽  
Abiotic Surface ◽  
Nutrient Metabolism ◽  
Content Type ◽  
A Cell

For the generation of energy, the important human pathogenStreptococcus pneumoniaerelies on host-derived sugars, including β-glucoside analogs. The catabolism of these nutrients involves the action of 6-phospho-β-glucosidase to convert them into usable monosaccharaides. In this study, we characterized a 6-phospho-β-glucosidase (BglA3) encoded by SPD_0247. We found that this enzyme has a cell membrane localization and is active only against a phosphorylated substrate. A mutated pneumococcal ΔSPD0247 strain had reduced 6-phospho-glucosidase activity and was attenuated in growth on cellobiose and hyaluronic acid compared to the growth of wild-type D39. ΔSPD0247-infected mice survived significantly longer than the wild-type-infected cohort, and the colony counts of the mutant were lower than those of the wild type in the lungs. The expression of SPD_0247 inS. pneumoniaeharvested from infected tissues was significantly increased relative to its expressionin vitroon glucose. Additionally, ΔSPD0247 is severely impaired in its attachment to an abiotic surface. These results indicate the importance of β-glucoside metabolism in pneumococcal survival and virulence.

scholarly journals Mechanism of β-Lactam Action in Streptococcus pneumoniae: the Piperacillin Paradox

2014 ◽  
Vol 59 (1) ◽  
pp. 609-621 ◽  
Author(s):  
Jules Philippe ◽  
Benoit Gallet ◽  
Cécile Morlot ◽  
Dalia Denapaite ◽  
Regine Hakenbeck ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Streptococcus Pneumoniae ◽  
Mode Of Action ◽  
Time Course ◽  
Binding Proteins ◽  
Epifluorescence Microscopy ◽  
Human Pathogen ◽  
Content Type ◽  
Penicillin Binding Proteins

ABSTRACTThe human pathogenStreptococcus pneumoniaehas been treated for decades with β-lactam antibiotics. Its resistance is now widespread, mediated by the expression of mosaic variants of the target enzymes, the penicillin-binding proteins (PBPs). Understanding the mode of action of β-lactams, not only in molecular detail but also in their physiological consequences, will be crucial to improving these drugs and any counterresistances. In this work, we investigate the piperacillin paradox, by which this β-lactam selects primarily variants of PBP2b, whereas its most reactive target is PBP2x. These PBPs are both essential monofunctional transpeptidases involved in peptidoglycan assembly. PBP2x participates in septal synthesis, while PBP2b functions in peripheral elongation. The formation of the “lemon”-shaped cells induced by piperacillin treatment is consistent with the inhibition of PBP2x. Following the examination of treated and untreated cells by electron microscopy, the localization of the PBPs by epifluorescence microscopy, and the determination of the inhibition time course of the different PBPs, we propose a model of peptidoglycan assembly that accounts for the piperacillin paradox.

scholarly journals Transcriptional Repressor PtvR Regulates Phenotypic Tolerance to Vancomycin in Streptococcus pneumoniae

2017 ◽  
Vol 199 (14) ◽  
Author(s):  
Xue Liu ◽  
Jing-Wen Li ◽  
Zhixing Feng ◽  
Youfu Luo ◽  
Jan-Willem Veening ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Treatment Failure ◽  
Molecular Mechanisms ◽  
Transcriptional Repressor ◽  
Drug Tolerance ◽  
Microbial Populations ◽  
Luciferase Reporter ◽  
Chronic Infections ◽  
Mobility Shift ◽  
Content Type

ABSTRACT Reversible or phenotypic tolerance to antibiotics within microbial populations has been implicated in treatment failure of chronic infections and development of persister cells. However, the molecular mechanisms regulating phenotypic drug tolerance are largely unknown. In this study, we identified a four-gene operon in Streptococcus pneumoniae that contributes to phenotypic tolerance to vancomycin (ptv). RNA sequencing, quantiative reverse transcriptase PCR, and transcriptional luciferase reporter experiments revealed that transcription of the ptv operon (consisting of ptvR, ptvA, ptvB, and ptvC) is induced by exposure to vancomycin. Further investigation showed that transcription of the ptv operon is repressed by PtvR, a PadR family repressor. Transcriptional induction of the ptv operon by vancomycin was achieved by transcriptional derepression of this locus, which was mediated by PtvR. Importantly, fully derepressing ptvABC by deleting ptvR or overexpressing the ptv operon with an exogenous promoter significantly enhanced vancomycin tolerance. Gene deletion analysis revealed that PtvA, PtvB, and PtvC are all required for the PtvR-regulated phenotypic tolerance to vancomycin. Finally, the results of an electrophoretic mobility shift assay with recombinant PtvR showed that PtvR represses the transcription of the ptv operon by binding to two palindromic sequences within the ptv promoter. Together, the ptv locus represents an inducible system in S. pneumoniae in response to stressful conditions, including those caused by antibiotics. IMPORTANCE Reversible or phenotypic tolerance to antibiotics within microbial populations is associated with treatment failure of bacterial diseases, but the underlying mechanisms regulating phenotypic drug tolerance remain obscure. This study reports our finding of a multigene locus that contributes to inducible tolerance to vancomycin in Streptococcus pneumoniae, an important opportunistic human pathogen. The vancomycin tolerance phenotype depends on the PtvR transcriptional repressor and three predicted membrane-associated proteins encoded by the ptv locus. This represents the first example of a gene locus in S. pneumoniae that is responsible for antibiotic tolerance and has important implications for further understanding bacterial responses and phenotypic tolerance to antibiotic treatment in this and other pathogens.

scholarly journals Inactivation of Transcriptional Regulator FabT Influences Colony Phase Variation of Streptococcus pneumoniae

mBio ◽  
2021 ◽  
Author(s):  
Jinghui Zhang ◽  
Weijie Ye ◽  
Kaifeng Wu ◽  
Shengnan Xiao ◽  
Yuqiang Zheng ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Cell Surface ◽  
Virulence Factors ◽  
Capsular Polysaccharide ◽  
Phase Variation ◽  
Transcriptional Regulator ◽  
Human Pathogen ◽  
Content Type ◽  
Reversible Phase

Streptococcus pneumoniae is a major human pathogen, and its virulence factors and especially the capsular polysaccharide have been extensively studied. In addition to virulence components that are present on its cell surface that directly interact with the host, S. pneumoniae undergoes a spontaneous and reversible phase variation that allows survival in different host environments.

scholarly journals Pivotal Roles for Ribonucleases in Streptococcus pneumoniae Pathogenesis

mBio ◽  
2021 ◽  
Author(s):  
Dhriti Sinha ◽  
Jacob P. Frick ◽  
Kristen Clemons ◽  
Malcolm E. Winkler ◽  
Nicholas R. De Lay
Keyword(s):  
Gene Expression ◽  
Streptococcus Pneumoniae ◽  
Host Cell ◽  
Cell Interactions ◽  
Human Pathogen ◽  
Protein Coding ◽  
Content Type ◽  
Key Players ◽  
Host Cell Interactions ◽  
Host Tissues

Streptococcus pneumoniae is a notorious human pathogen that adapts to conditions in distinct host tissues and responds to host cell interactions by adjusting gene expression. RNases are key players that modulate gene expression by mediating the turnover of regulatory and protein-coding transcripts.

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