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Bacillus subtilis remains translationally active after CRISPRi-mediated replication initiation arrest.
Muñoz-Gutierrez, Vanessa; Cornejo, Fabián A; Schmidt, Katja; Frese, Christian K; Halte, Manuel; Erhardt, Marc; Elsholz, Alexander K W; Turgay, Kürsad; Charpentier, Emmanuelle.
Afiliación
  • Muñoz-Gutierrez V; Max Planck Unit for the Science of Pathogens, Berlin, Germany.
  • Cornejo FA; Institute of Microbiology, Leibniz Universität Hannover, Hannover, Germany.
  • Schmidt K; Max Planck Unit for the Science of Pathogens, Berlin, Germany.
  • Frese CK; Max Planck Unit for the Science of Pathogens, Berlin, Germany.
  • Halte M; Max Planck Unit for the Science of Pathogens, Berlin, Germany.
  • Erhardt M; Humboldt-Universität zu Berlin, Institute of Biology - Molecular Microbiology, Berlin, Germany.
  • Elsholz AKW; Max Planck Unit for the Science of Pathogens, Berlin, Germany.
  • Turgay K; Humboldt-Universität zu Berlin, Institute of Biology - Molecular Microbiology, Berlin, Germany.
  • Charpentier E; Max Planck Unit for the Science of Pathogens, Berlin, Germany.
mSystems ; 9(4): e0022124, 2024 Apr 16.
Article en En | MEDLINE | ID: mdl-38546227
ABSTRACT
Initiation of bacterial DNA replication takes place at the origin of replication (oriC), a region characterized by the presence of multiple DnaA boxes that serve as the binding sites for the master initiator protein DnaA. This process is tightly controlled by modulation of the availability or activity of DnaA and oriC during development or stress conditions. Here, we aimed to uncover the physiological and molecular consequences of stopping replication in the model bacterium Bacillus subtilis. We successfully arrested replication in B. subtilis by employing a clustered regularly interspaced short palindromic repeats interference (CRISPRi) approach to specifically target the key DnaA boxes 6 and 7, preventing DnaA binding to oriC. In this way, other functions of DnaA, such as a transcriptional regulator, were not significantly affected. When replication initiation was halted by this specific artificial and early blockage, we observed that non-replicating cells continued translation and cell growth, and the initial replication arrest did not induce global stress conditions such as the SOS response.IMPORTANCEAlthough bacteria constantly replicate under laboratory conditions, natural environments expose them to various stresses such as lack of nutrients, high salinity, and pH changes, which can trigger non-replicating states. These states can enable bacteria to (i) become tolerant to antibiotics (persisters), (ii) remain inactive in specific niches for an extended period (dormancy), and (iii) adjust to hostile environments. Non-replicating states have also been studied because of the possibility of repurposing energy for the production of additional metabolites or proteins. Using clustered regularly interspaced short palindromic repeats interference (CRISPRi) targeting bacterial replication initiation sequences, we were able to successfully control replication initiation in Bacillus subtilis. This precise approach makes it possible to study non-replicating phenotypes, contributing to a better understanding of bacterial adaptive strategies.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Bacillus subtilis / Proteínas de Unión al ADN Idioma: En Revista: MSystems Año: 2024 Tipo del documento: Article País de afiliación: Alemania

Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Bacillus subtilis / Proteínas de Unión al ADN Idioma: En Revista: MSystems Año: 2024 Tipo del documento: Article País de afiliación: Alemania
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