Genetic modules for sensing and detoxification of novel antibacterial compounds

• Applicant: Privatdozent Dr. Sven Halbedel
• Subject Area: Medical Microbiology and Mycology, Hygiene, Molecular Infection Biology ; Metabolism, Biochemistry and Genetics of Microorganisms
• Term: from 2019 to 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 433766034

• Final Report Year: 2025

Final Report Abstract

The bacterium Listeria monocytogenes is the causative agent of listeriosis, a foodborne infectious human disease with high mortality. L. monocytogenes is found in various environmental habitats. From there it often enters the food processing chain and leads to food contamination, human infections and disease outbreaks. In the environment, L. monocytogenes is exposed to antimicrobial compounds produced by competing species. Resistance against such compounds can depend on multi drug resistance (MDR) transporters, which export harmful substances out of the cell. L. monocytogenes has 24 of these transporters, but their natural substrates are mostly unknown. Their identification would be beneficial for the better specification of the pathogen´s natural reservoir, but also for the identification of novel antimicrobial compounds. We here generated different L. monocytogenes reporter strains for compound screens with collections of naturally occurring antimicrobial compounds. These screens yielded tartrolon B as the substrate of the so far uncharacterized MDR transporter TimAB. The genes for this transporter are encoded in the timABR operon together with a transcriptional repressor (timR). We showed that the TimR repressor senses tartrolon B to regulate timABR expression. Tartrolon B is a macrodioloide produced by terrestrial and marine bacteria, suggesting that marine environments could play a role as reservoir of L. monocytogenes in addition to its known terrestrial reservoirs. Mode of action studies showed that tartrolon B exposure caused leakage of potassium ions from L. monocytogenes cells. However, suppressor mutants with high levels of tartrolon B resistance were found that were not related to potassium transport. These mutants carried inactivating mutations in the clpP2 gene, which encodes the main proteolytic component of Clp proteasomes. Various genetic experiments then showed that the transcription factor (and Clp substrate) SpxA1, its protease adaptor YjbH, the two Clp ATPases ClpC and ClpX, the SpxA1-dependent cytochrome oxidase CydAB and several other respiratory chain genes are also essential for tartrolon B resistance. The respiratory chain is important for the generation of a proton gradient across the membrane. For these and other reasons, it seems plausible that tartrolon B could act as a coupled potassium ionophore/protonophore dissipating transmembrane gradients of potassium ions and protons simultaneously. With these results, our work has generated novel insights in the ecological interactions of an important human pathogen and useful tools for antimicrobial compound discovery.

Publications

• Elements in the LftR Repressor Operator Interface Contributing to Regulation of Aurantimycin Resistance in Listeria monocytogenes. Journal of Bacteriology, 203(10).
  Hauf, Samuel; Engelgeh, Tim & Halbedel, Sven
  
• Insights into tartrolon resistance mechanisms in Listeria monocytogenes. Poster at the Annual Meeting of the Association for General and Applied Microbiology. 21-23 February. Online.
  Engelgeh T., Herrmann J., Müller R. & Halbedel S.
  
• Listeria monocytogenes genes supporting growth under standard laboratory cultivation conditions and during macrophage infection. Genome Research, 32(9), 1711-1726.
  Fischer, Martin A.; Engelgeh, Tim; Rothe, Patricia; Fuchs, Stephan; Thürmer, Andrea & Halbedel, Sven
  
• Perception and detoxification of the polyketide tartrolon B by Listeria monocytogenes. Poster at the 21st International Conference on Bacilli & Gram-Positive Bacteria. 14-17 June. Prague, Czechia.
  Engelgeh T., Herrmann J., Müller R. & Halbedel S.
  
• ListiWiki: A database for the foodborne pathogen Listeria monocytogenes. International Journal of Medical Microbiology, 313(6), 151591.
  Elfmann, Christoph; Zhu, Bingyao; Stülke, Jörg & Halbedel, Sven
  
• Perception and detoxification of the polyketide tartrolon B by Listeria monocytogenes. Talk at the BACELL Meeting 2023. 19-20 June. Hohenheim, Germany.
  Engelgeh T., Rothe P., Wamp S., Herrmann J., Müller R. & Halbedel S.
  
• Tartrolon resistance mechanisms in Listeria monocytogenes depend on the TimAB transporter and the ClpP2 proteasome. Poster at the Annual Meeting of the Association for General and Applied Microbiology. 10-23 September. Göttingen, Germany
  Engelgeh T., Herrmann J., Jansen R., Müller R. & Halbedel S.
  
• Tartrolon sensing and detoxification by the Listeria monocytogenes timABR resistance operon. Molecular Microbiology, 120(5), 629-644.
  Engelgeh, Tim; Herrmann, Jennifer; Jansen, Rolf; Müller, Rolf & Halbedel, Sven
  
• ClpP2 proteasomes and SpxA1 determine Listeria monocytogenes tartrolon B hyper-resistance. PLOS Genetics, 21(4), e1011621.
  Engelgeh, Tim; Wamp, Sabrina; Rothe, Patricia; Herrmann, Jennifer; Fischer, Martin A.; Müller, Rolf & Halbedel, Sven