Biomedical subjects

Laurence Van Melderen

Publications and source records attributed to Laurence Van Melderen.

11 recordsLinked to original sources

PubMed2026

Acquisition and erosion of toxin-antitoxin systems in bacterial chromosomes.

Toxin-antitoxin systems (TAs) are widespread in bacterial genomes. Yet, their integration, persistence, and impact in chromosome dynamics remain unclear. Here, we identified 80 type II TAs in the single chromosome of Photorhabdus laumondii TT01, 50 of which were experimentally validated. Comparative analysis across the Photorhabdus genus revealed a highly heterogeneous distribution, with TAs frequently clustering within discrete genomic regions, either alone or associated with cointegrate-forming transposases and integrases. TAs rarely clustered with other putative defense systems and are preferentially associated with different types of recombinases, suggesting distinct pathways of acquisition for the two types of functions. Functional analyses showed that most validated TAs display addictive properties and stabilize plasmids. These addictive TAs are preferentially located in genomic regions characterized by high gene turnover, consistent with recent acquisition events. Despite their plasmid-stabilizing capacity, TAs do not promote long-term conservation of their immediate chromosomal neighborhoods. Instead, we observed frequent TA loss, either through complete deletion or toxin pseudogenization, indicating relaxed selection for their persistence in bacterial lineages. We propose a stepwise model for TA evolution in bacterial chromosomes: initial acquisition mediated by mobile genetic elements, preferential integration into permissive genomic regions, subsequent genetic streamlining of linked loci, and progressive gene loss. The short-lasting linkage between TAs and their genomic neighborhoods is consistent with the view that TA modules can behave as autonomous, selfish genetic elements.

Journal Article↗

PubMed2006

Differences between Belgian and Brazilian group A Streptococcus epidemiologic landscape.

BACKGROUND: Group A Streptococcus (GAS) clinical and molecular epidemiology varies with location and time. These differences are not or are poorly understood. METHODS AND FINDINGS: We prospectively studied the epidemiology of GAS infections among children in outpatient hospital clinics in Brussels (Belgium) and Brasília (Brazil). Clinical questionnaires were filled out and microbiological sampling was performed. GAS isolates were emm-typed according to the Center for Disease Control protocol. emm pattern was predicted for each isolate. 334 GAS isolates were recovered from 706 children. Skin infections were frequent in Brasília (48% of the GAS infections), whereas pharyngitis were predominant (88%) in Brussels. The mean age of children with GAS pharyngitis in Brussels was lower than in Brasília (65/92 months, p<0.001). emm-typing revealed striking differences between Brazilian and Belgian GAS isolates. While 20 distinct emm-types were identified among 200 Belgian isolates, 48 were found among 128 Brazilian isolates. Belgian isolates belong mainly to emm pattern A-C (55%) and E (42.5%) while emm pattern E (51.5%) and D (36%) were predominant in Brasília. In Brasília, emm pattern D isolates were recovered from 18.5% of the pharyngitis, although this emm pattern is supposed to have a skin tropism. By contrast, A-C pattern isolates were infrequently recovered in a region where rheumatic fever is still highly prevalent. CONCLUSIONS: Epidemiologic features of GAS from a pediatric population were very different in an industrialised country and a low incomes region, not only in term of clinical presentation, but also in terms of genetic diversity and distribution of emm patterns. These differences should be taken into account for designing treatment guidelines and vaccine strategies.

Adolescent↗

PubMed2006

Structural basis for nucleic acid and toxin recognition of the bacterial antitoxin CcdA.

Toxin-antitoxin systems are highly abundant in plasmids and bacterial chromosomes. They ensure plasmid maintenance by killing bacteria that have lost the plasmid. Their expression is autoregulated at the level of transcription. Here, we present the solution structure of CcdA, the antitoxin of the ccd system, as a free protein (16.7 kDa) and in complex with its cognate DNA (25.3 kDa). CcdA is composed of two distinct and independent domains: the N-terminal domain, responsible for DNA binding, which establishes a new family of the ribbon-helix-helix fold and the C-terminal region, which is responsible for the interaction with the toxin CcdB. The C-terminal domain is intrinsically unstructured and forms a tight complex with the toxin. We show that CcdA specifically recognizes a 6 bp palindromic DNA sequence within the operator-promoter (OP) region of the ccd operon and binds to DNA by insertion of the positively charged N-terminal beta-sheet into the major groove. The binding of up to three CcdA dimers to a 33mer DNA of its operator-promoter region was studied by NMR spectroscopy, isothermal titration calorimetry and single point mutation. The highly flexible C-terminal region of free CcdA explains its susceptibility to proteolysis by the Lon ATP-dependent protease.

Amino Acid Sequence↗

PubMed2006

Biological roles of the Lon ATP-dependent protease.

The Lon ATP-dependent protease plays a major role in protein quality control. An increasing number of regulatory proteins, however, are being identified as Lon substrates, thus indicating that in addition to its housekeeping function, Lon plays an important role in regulating many biological processes in bacteria. This review presents and discusses the involvement of Lon in different aspects of bacterial physiology, including cell differentiation, sporulation, pathogenicity and survival under starvation conditions.

Bacillus subtilis↗

PubMed2006

Expression of the F plasmid ccd toxin-antitoxin system in Escherichia coli cells under nutritional stress.

The ccd system of the F plasmid encodes CcdB, a protein toxic to DNA-gyrase, and CcdA, its antitoxin. The function attributed to this system is to contribute to plasmid stability by killing bacteria that lose the plasmid during cell division. However, the function of ccd in resting bacteria is not clear. Results presented show that ccd transcription increases as bacteria enter stationary phase and that the amount of the Ccd proteins is higher in bacteria under nutritional stress than in growing bacteria. Moreover, an increase in the frequency of Lac+ "adaptive" mutations was observed in stationary-phase bacteria that over-express the Ccd proteins.

Bacterial Proteins↗

PubMed2006

Pharyngitis in low-resources settings: a pragmatic clinical approach to reduce unnecessary antibiotic use.

OBJECTIVE: Existing scoring systems for the diagnosis of group A streptococcus pharyngitis are insensitive or inapplicable in low-resources settings. Bacterial cultures and rapid tests can allow for antibiotic prescription abstention in high-income regions. These techniques are not feasible in many low-resources settings, and antibiotics often are prescribed for any pharyngitis episode. However, judicious antibiotics prescription in the community also is of concern in low-income countries. The objective of this study was to develop a clinical decision rule that allows for the reduction of empirical antibiotic therapy for children with pharyngitis in low-resources settings by identifying non-group A streptococcus pharyngitis. PATIENTS AND METHODS: We prospectively included children with pharyngitis in 3 public hospitals of Brazil during 9 months in 2004. We filled out clinical questionnaires and performed throat swabs. Bilateral chi2 (2-tailed test) and multivariate analysis were used to determine score categories. The outcome measures were sensitivity, specificity, positive likelihood ratio, and posttest probability of non-group A streptococcus infection with the clinical approach as compared with throat culture. RESULTS: A total of 163 of the 220 children had non-group A streptococcus pharyngitis (negative culture). We established a 3-questions decision rule (age and viral and bacterial signs) with 3 possible answers. The use of this score would prevent 41% to 55% of unnecessary antimicrobial prescriptions. The specificity of the score for non-group A streptococcus pharyngitis was >84%. CONCLUSION: Such a clinical decision rule could be helpful to reduce significantly unnecessary antibiotic prescriptions for pharyngitis in children in low-resources settings.

Anti-Bacterial Agents↗

PubMed2005

Molecular basis of gyrase poisoning by the addiction toxin CcdB.

Gyrase is an ubiquitous bacterial enzyme that is responsible for disentangling DNA during DNA replication and transcription. It is the target of the toxin CcdB, a paradigm for plasmid addiction systems and related bacterial toxin-antitoxin systems. The crystal structure of CcdB and the dimerization domain of the A subunit of gyrase (GyrA14) dictates an open conformation for the catalytic domain of gyrase when CcdB is bound. The action of CcdB is one of a wedge that stabilizes a dead-end covalent gyrase:DNA adduct. Although CcdB and GyrA14 form a globally symmetric complex where the two 2-fold axes of both dimers align, the complex is asymmetric in its details. At the centre of the interaction site, the Trp99 pair of CcdB stacks with the Arg462 pair of GyrA14, explaining why the Arg462Cys mutation in the A subunit of gyrase confers resistance to CcdB. Overexpression of GyrA14 protects Escherichia coli cells against CcdB, mimicking the action of the antidote CcdA.

Bacterial Proteins↗

PubMed2004

Crystallization of CcdB in complex with a GyrA fragment.

Plasmid addiction systems consist of a plasmid-encoded toxin-antidote pair that serves to stabilize low-copy-number plasmids in bacterial populations. CcdB, the toxin from the ccd system on the Escherichia coli F plasmid, acts as a gyrase poison. A 14 kDa fragment of gyrase, GyrA14, was found to bind to the toxin CcdB with an affinity of 1.75 x 10(-8) M. Crystals of the (GyrA14)(2) dimer in its free state belong to space group P4(3)2(1)2, with unit-cell parameters a = 86.4, c = 89.4 angstroms, and diffract to 2.4 angstroms. Crystals of the (GyrA14)(2)-(CcdB)(2) complex belong to space group P2(1)2(1)2(1), with a = 52.1, b = 83.3, c = 110.9 angstroms, and diffract to 2.8 angstroms resolution.

Bacterial Proteins↗

PubMed2004

Overproduction of the Lon protease triggers inhibition of translation in Escherichia coli: involvement of the yefM-yoeB toxin-antitoxin system.

In Escherichia coli, the Lon ATP-dependent protease is responsible for degradation of several regulatory proteins and for the elimination of abnormal proteins. Previous studies have shown that the overproduction of Lon is lethal. Here, we showed that Lon overproduction specifically inhibits translation through at least two different pathways. We have identified one of the pathways as being the chromosomal yefM-yoeB toxin-antitoxin system. The existence of a second pathway is demonstrated by the observation that the deletion of the yefM-yoeB system did not completely suppress lethality and translation inhibition. We also showed that the YoeB toxin induces cleavage of translated mRNAs and that Lon overproduction specifically activates YoeB-dependent mRNAs cleavage. Indeed, none of the other identified chromosomal toxin-antitoxin systems (relBE, mazEF, chpB and dinJ-yafQ) was involved in Lon-dependent lethality, translation inhibition and mRNA cleavage even though the RelB and MazE antitoxins are known to be Lon substrates. Based on our results and other studies, translation inhibition appears to be the key element that triggers chromosomal toxin-antitoxin systems. We propose that under Lon overproduction conditions, translation inhibition is mediated by Lon degradation of a component of the YoeB-independent pathway, in turn activating the YoeB toxin by preventing synthesis of its unstable YefM antidote.

ATP-Dependent Proteases↗

PubMed2002

Molecular interactions of the CcdB poison with its bacterial target, the DNA gyrase.

The ccd poison/antidote system of the F plasmid encodes CcdB, a toxin targeting the essential DNA gyrase of E. coli, and CcdA, the unstable antidote that interacts with CcdB to neutralise its toxicity. Gyrase belongs to the topoisomerase II class of enzymes and is a well-validated target for efficient therapeutic drugs, i. e. the quinolones. CcdB acts on gyrase in a similar way as quinolones do, both compounds induce double-strand breaks in DNA. Interestingly, the CcdB-binding domain of gyrase is different than that of quinolones. Therefore, novel classes of therapeutic drugs could be derived from the analysis of the interaction between CcdB and gyrase.

Bacterial Proteins↗

PubMed2002

The highly conserved TldD and TldE proteins of Escherichia coli are involved in microcin B17 processing and in CcdA degradation.

Microcin B17 (MccB17) is a peptide antibiotic produced by Escherichia coli strains carrying the pMccB17 plasmid. MccB17 is synthesized as a precursor containing an amino-terminal leader peptide that is cleaved during maturation. Maturation requires the product of the chromosomal tldE (pmbA) gene. Mature microcin is exported across the cytoplasmic membrane by a dedicated ABC transporter. In sensitive cells, MccB17 targets the essential topoisomerase II DNA gyrase. Independently, tldE as well as tldD mutants were isolated as being resistant to CcdB, another natural poison of gyrase encoded by the ccd poison-antidote system of plasmid F. This led to the idea that TldD and TldE could regulate gyrase function. We present in vivo evidence supporting the hypothesis that TldD and TldE have proteolytic activity. We show that in bacterial mutants devoid of either TldD or TldE activity, the MccB17 precursor accumulates and is not exported. Similarly, in the ccd system, we found that TldD and TldE are involved in CcdA and CcdA41 antidote degradation rather than being involved in the CcdB resistance mechanism. Interestingly, sequence database comparisons revealed that these two proteins have homologues in eubacteria and archaebacteria, suggesting a broader physiological role.

Antidotes↗