ED Sciences de la Vie et de la Santé
Mechanisms of ribosome inhibition by context-dependent antibiotics targeting the decoding center of the bacterial ribosome
by Arunima BHATTACHARYA (Acides nucléiques : Régulations Naturelles et Artificielles)
The defense will take place at 14h00 - Amphithéâtre Université de Bordeaux, 2 rue Dr Hoffmann Martinot, Bâtiment BBS, 33000 Bordeaux
in front of the jury composed of
- Axel INNIS - Directeur de recherche - Université de Bordeaux - Directeur de these
- Célia PLISSON-CHASTANG - Directrice de recherche - Université de Toulouse - Rapporteur
- Reynald GILLET - Professeur - Université de Rennes - Rapporteur
- Eric ENNIFAR - Directeur de recherche - Université de Strasbourg - Examinateur
- Grégory BOEL - Directeur de recherche - Université Paris Cité - Examinateur
Antibiotics are essential in modern medicine to combat bacteria pathogenic to humans, but resistance to existing antibiotics is a serious threat to patient lives and the sustainability of healthcare infrastructure. While innovation is necessary to introduce novel treatments, it is also important to revisit existing drugs in detail to understand their complete mechanisms of action and explore ways to improve them to combat resistant pathogens. Antibiotics targeting the ribosome, the main player in the bacterial protein synthesis machinery, account for the majority of clinical drugs in use. Research over the last decade has revealed that contrary to the long-held view that ribosome-targeting drugs inhibit the synthesis of all proteins with the same efficiency, many antibiotics are selective and inhibit translation in a sequence-specific manner. Probing into the mechanism of action of such context-dependent antibiotics is important since this can lead to novel methods of counteracting drug-resistant pathogens in a species-specific manner. Ribosome-targeting drugs can inhibit translation by binding to several functionally important sites on the ribosome, and the decoding center, where the genetic code encoded in the mRNA is translated into proteins, is a critical drug target, since perturbations in the decoding function can lead to mistranslation and production of aberrant proteins, ultimately compromising cell viability. While several decoding center-targeting antibiotics exist in clinical use, their context-dependent mode of action has remained underexplored. In this work, I study the sequence-dependent mechanism of action of two classes of antibiotics targeting the decoding center of the bacterial ribosome – odilorhabdins, a relatively new class of peptide antibiotics discovered in the last decade, and tigecycline, a third-generation tetracycline in regular clinical use. To identify the sequence contexts where these drugs induce ribosomal stalling in addition to their impact on decoding fidelity, I used inverse toeprinting coupled to high-throughput sequencing (iTP-Seq), an in vitro profiling technique that can locate ribosomes stalled on mRNA with codon resolution. Further, to investigate the mechanism of action of odilorhabdins, I used cryo-electron microscopy and single-particle analysis and studied multiple conformational states of drug-bound ribosomes stalled on a representative arrest motif. Overall, this thesis reveals the sequence-dependent mode of inhibition of two antibiotics targeting the decoding center of the ribosome, and elaborates the mechanism of action of a recently discovered class of antibiotics at the structural level, contributing to the foundational knowledge needed for structure-guided development of next-generation ribosome-targeting antibiotics.