Team
Team Manager: Polard Patrice
Presentation
The pneumococcal competence and transformation team studies all aspects of competence and genetic transformation in the human pathogen Streptococcus pneumoniae (the pneumococcus). Pneumococcal competence is a transient genetic programme induced in response to various types of stresses, including antibiotics, which targets different cellular processes. Competence promotes the mechanism of transformation, well known to direct horizontal gene transfer, as well as providing cells with other properties, including the killing of non-competent siblings, altered tolerance to lethal drugs, cell colonization and virulence.
Genetic transformation involves capture of exogenous DNA, internalization in the form of single strands and integration into the recipient chromosome by homologous recombination, driven by the recombinase RecA. This process is widespread in the bacterial kingdom and promotes acquisition of new genetic traits. In the pneumococcus, transformation can mediate the spread of antibiotics and vaccine escape.
Our team combines advances techniques of molecular genetics and cell biology to study in the pneumococcus both the transformation mechanism and on competence development, regulation and associated properties. The main projects of our team currently focus on:
– The functional dissection of the mechanism of transformation.
– An analysis of genome dynamics during competence.
– The links between competence and antiobiotic tolerance in the pneumococcus.
– The mechanism of action of the RumC1 bacteriocin.
Through these complementary projects, we aim to further our fundamental understanding of the mechanisms involved in competence and transformation, and how these clinically relevant processes are integrated into the lifestyle of this major human pathogen.
Project 1
Transformation is a conserved mechanism of lateral DNA transfer and recombination promoting the acquisition of new genetic traits. In involves the capture of exogenous DNA followed by its uptake in single strand form (ssDNA) and its integration into the chromosome by RecA-directed homologous recombination (HR). While the proteins involved in transformation are now mostly identified, the molecular mechanisms involved in the process remain poorly characterized. This project aims to further our understanding of how the proteins involved in transformation interact to ensure DNA uptake and integration during the process. The objectives of this project are as follows:
– Use in silico structural modelling and functional assays to explore how the ComEC transmembrane pore coordinates and interacts with proteins on both sides of the membrane to ensure the efficient uptake of transforming ssDNA. This is a collaboration with Raphael GUEROIS and Jessica ANDREANI (I2BC, Paris).
– Explore the maturation of the transformation D-loop structure generated during integration into the recipient chromosome. This involves revealing the roles of the recombination proteins directing this terminal recombinational reactions, and how these are coordinated to ensure chromosomal integration of transforming ssDNA. This is a collaboration with Xavier Charpentier (CIRI, Lyon).
Project 2
Competence is a transient physiological state that reprograms cells with new properties. How competence is integrated into the cell-cycle and affects the genome dynamics remains poorly understood. This project aims to unravel how the competence state impacts the genome at multiple levels:
– RecA-directed HR mechanism is central and common to both genome maintenance and transformation. We are interested in understanding how both HR pathways are managed together during competence, and the mechanisms involved in their spatiotemporal coordination.
– One pathway of competence modulation involves a toxin-antitoxin system, with the toxin targeting the replication protein DnaN. We explore the mechanisms controlling this toxin-antitoxin system in collaboration with Patricia BORDES and Pierre GENEVAUX (CBI, Toulouse).
– We recently revealed that specific proteins are phosphorylated during competence, including effectors involved in DNA repair. This axis explores the general effect of competence and protein phosphorylation on genome architecture. This involves a collaboration with Christophe GRANGEASSE (MMSB, Lyon) and Romain KOZUL (Institut Pasteur, Paris).
Project 3
We recently showed that competence promotes altered tolerance to varied antibiotics. A transient division delay mediated by the competence protein ComM is key for this phenotype, providing populations with time to overcome the stress using stress-specific mechanisms, and thus increasing survival. This finding revealed competence as a general stress response, and this project aims to further our understanding of the links between competence and antibiotic tolerance by:
– Using a reference laboratory strain to explore the underlying mechanisms involved in the competence-mediated tolerance to specific antibiotic stresses.
– Exploring the metabolic shift that occurs in competent cells and relating this to the competence-mediated tolerance phenotype. This is a collaboration with Stéphanie HEUX (TBI) and two linked Genotoul platforms (Get and MetaToul).
– Uncovering the conservation of competence-mediated tolerance in clinical isolates of pneumococci, including lineages multi-resistant to antibiotics and other streptococcal pathogenic species, many of them developing competence by a different regulation mechanism. This is a collaboration with S. LO (University of Bath) and P. HOLS (Catholic university of Louvain).
Project 4
RumC1 is a bacteriocin encoded by Ruminococcus gnavus from the human gut microbiota, It exhibits bactericidal activity against a large spectrum of bacteria, including the pneumococcus and clinical pathogenic strains multi-resistant to antibiotics. RumC1 is exceptionally resistant to human complement, high temperature and extreme pHs, which makes it a promising candidate to design a novel antibiotic. This challenging perspective requires the characterization of its killing mechanism and of its specific target site. We engaged the molecular study of the action mode of RumC1 in S. pneumoniae, by combining the genetic, biochemical and cell biology approaches that we developed for characterizing the unknown roles of proteins involved in competence or transformation. We conduct this molecular characterization of the RumC1 killing mechanism of action in collaboration with V. DUARTE (LBCM, Grenoble) and C. MORLOT (IBS, Grenoble).
– De Lemos D, Soulet AL, Morales V, Bergé MJ, Polard P, Johnston CHG. Competence induction of homologous recombination genes protects pneumococcal cells from genotoxic stress. mBio. 2025 Jan, 8;16(1).
– Structural insights into the mechanism of DNA branch migration during homologous recombination in bacteria. Rosa LT, Vernhes E, Soulet AL, Polard P, Fronzes R. EMBO Journal, 2024 43 :6180-6198.
– Johnston CHG, Prudhomme M, Soulet AL, Boyeldieu A, De Lemos D, Campo N, Polard P. Pneumococcal competence is a populational health sensor driving multilevel heterogeneity in response to antibiotics. Nat Commun. 2024 Jul 10;15(1):5625.
– Maziero M, Lane D, Polard P, Bergé M. Fever-like termperature bursts promote competence development via an HtrA-dependent pathway in Streptococcus pneumoniae. PLoS Genet 2023 Sep 12;19(9).
– Assembly mechanism and cryoEM structure of RecA recombination nucleofilaments from Streptococcus pneumoniae. Hertzog M, Perry T, Dupaigne P, Serres S, Morales V, Soulet AL, Bell J, Margeat E, Kowalczykowski S, Le Cam E, Fronzes R, Polard P. Nucleic Acid Research, 2023 Apr 11;51(6):2800-2817.
– The RecA-directed recombination pathway of natural transformation initiates at chromosomal replication forks in Streptococcus pneumoniae. Johnston CHG, Hope R, Soulet AL, Dewailly M, De Lemos D, Polard P. PNAS. 2023 Feb 21;120(8).
Affiliation
