Reporter-Guided Transposon Mutant Selection for Activation of Silent Gene Clusters in Burkholderia thailandensis.

TitleReporter-Guided Transposon Mutant Selection for Activation of Silent Gene Clusters in Burkholderia thailandensis.
Publication TypeJournal Article
Year of Publication2020
AuthorsMao, D, Yoshimura, A, Wang, R, Seyedsayamdost, MR
JournalChembiochem
Volume21
Issue13
Pagination1826-1831
Date Published2020 Jul 01
ISSN1439-7633
Abstract

<p>Most natural product biosynthetic gene clusters that can be observed bioinformatically are silent. This insight has prompted the development of several methodologies for inducing their expression. One of the more recent methods, termed reporter-guided mutant selection (RGMS), entails creation of a library of mutants that is then screened for the desired phenotype via reporter gene expression. Herein, we apply a similar approach to Burkholderia thailandensis and, using transposon mutagenesis, mutagenize three strains, each carrying a fluorescent reporter in the malleilactone (mal), capistruin (cap), or an unidentified ribosomal peptide (tomm) gene cluster. We show that even a small library of <500 mutants can be used to induce expression of each cluster. We also explore the mechanism of activation and find that inhibition of pyrimidine biosynthesis is linked to the induction of the mal cluster. Both a transposon insertion into pyrF as well as small-molecule-mediated inhibition of PyrF trigger malleilactone biosynthesis. Our results pave the way toward the broad application of RGMS and related approaches to Burkholderia spp.</p>

DOI10.1002/cbic.201900748
Alternate JournalChembiochem
PubMed ID31984619
PubMed Central IDPMC7493819
Grant ListDP2 AI124786 / AI / NIAID NIH HHS / United States
DP2-AI-124786 / / Foundation for the National Institutes of Health /
Postodoctoral Fellowship / / Uehara Memorial Foundation /