High-frequency transposition for determining antibacterial mode of action

Hao Wang, David Claveau, John P. Vaillancourt, Terry Roemer, Timothy Charles Meredith

Research output: Contribution to journalArticle

30 Scopus citations

Abstract

Connecting bacterial growth inhibitors to molecular targets at the whole-cell level is a major impediment to antibacterial development. Herein we report the design of a highly efficient and versatile bacteriophage-based mariner transposon delivery system in Staphylococcus aureus for determining inhibitor mode of action. Using bacteriophage-mediated delivery of concatameric minitransposon cassettes, we generated nonclonal transposant libraries with genome-wide insertion-site coverage in either laboratory or methicillin- resistant strain backgrounds and screened for drug resistance in situ on a single agar plate in one step. A gradient of gene-target expression levels, along with a correspondingly diverse assortment of drug-resistant phenotypes, was achieved by fitting the transposon cassette with a suite of outward-facing promoters. Using a panel of antibiotics, we demonstrate the ability to unveil not only an inhibitor's molecular target but also its route of cellular entry, efflux susceptibility and other off-target resistance mechanisms.

Original languageEnglish (US)
Pages (from-to)720-729
Number of pages10
JournalNature Chemical Biology
Volume7
Issue number10
DOIs
Publication statusPublished - Jan 1 2011

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All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cell Biology

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