Rationally designed pooled CRISPRi-seq uncovers an inhibitor of bacterial peptidyl-tRNA hydrolase.

Autor:	Rahman ASMZ; Department of Microbiology, University of Manitoba, Winnipeg, MB R3T 2N2, Canada., Syroegin EA; Department of Biological Sciences, University of Illinois at Chicago, Chicago, IL 60607, USA., Novomisky Nechcoff J; Department of Microbiology, University of Manitoba, Winnipeg, MB R3T 2N2, Canada., Devarajan A; Department of Microbiology, University of Manitoba, Winnipeg, MB R3T 2N2, Canada., Polikanov YS; Department of Biological Sciences, University of Illinois at Chicago, Chicago, IL 60607, USA; Center for Biomolecular Sciences, University of Illinois at Chicago, Chicago, IL 60607, USA; Department of Pharmaceutical Sciences, University of Illinois at Chicago, Chicago, IL 60607, USA., Cardona ST; Department of Microbiology, University of Manitoba, Winnipeg, MB R3T 2N2, Canada; Department of Medical Microbiology and Infectious Diseases, University of Manitoba, Winnipeg, MB R3E 0J9, Canada. Electronic address: silvia.cardona@umanitoba.ca.
Jazyk:	angličtina
Zdroj:	Cell reports [Cell Rep] 2024 Nov 26; Vol. 43 (11), pp. 114967. Date of Electronic Publication: 2024 Nov 15.
DOI:	10.1016/j.celrep.2024.114967
Abstrakt:	Bacterial mutant libraries with downregulated antibiotic targets are useful tools for elucidating the mechanisms of action of antibacterial compounds, a pivotal step in antibiotic discovery. However, achieving genomic coverage of antibacterial targets poses a challenge due to the uneven proliferation of knockdown mutants during pooled growth, leading to the unintended loss of important targets. To overcome this issue, we constructed an arrayed essential gene mutant library (EGML) in the antibiotic-resistant bacterium Burkholderia cenocepacia using CRISPR interference (CRISPRi). By modeling depletion levels and adjusting knockdown mutant inocula, we rationally designed and optimized a CRISPR interference-mediated pooled library of essential genes (CIMPLE) approaching coverage of the bacterial essential genome with mutant sensitization. We exposed CIMPLE to an uncharacterized bacterial growth inhibitor structurally different from antibiotics and discovered that it inhibits the essential peptidyl-tRNA hydrolase. Overall, CIMPLE leverages the advantages of arrayed and pooled CRISPRi libraries to uncover unexplored targets for antibiotic action. Competing Interests: Declaration of interests The authors declare no competing financial interests. (Crown Copyright © 2024. Published by Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
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