A tunable anthranilate-inducible gene expression system for Pseudomonas species
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APPLIED GENETICS AND MOLECULAR BIOTECHNOLOGY
A tunable anthranilate-inducible gene expression system for Pseudomonas species Lena Hoffmann 1 & Michael-Frederick Sugue 1 & Thomas Brüser 1 Received: 5 October 2020 / Revised: 18 November 2020 / Accepted: 23 November 2020 # The Author(s) 2020
Abstract Pseudomonads are among the most common bacteria in soils, limnic ecosystems, and human, animal, or plant host environments, including intensively studied species such as Pseudomonas aeruginosa, P. putida, or P. fluorescens. Various gene expression systems are established for some species, but there is still a need for a simple system that is suitable for a wide range of pseudomonads and that can be used for physiological applications, i.e., with a tuning capacity at lower expression levels. Here, we report the establishment of the anthranilate-dependent PantA promoter for tunable gene expression in pseudomonads. During studies on P. fluorescens, we constructed an anthranilate-inducible AntR/PantA-based expression system, named pUCP20-ANT, and used GFP as reporter to analyze gene expression. This system was compared with the rhamnose-inducible RhaSR/PrhaB-based expression system in an otherwise identical vector background. While the rhamnose-inducible system did not respond to lower inducer concentrations and always reached high levels over time when induced, expression levels of the pUCP20-ANT system could be adjusted to a range of distinct lower or higher levels by variation of anthranilate concentrations in the medium. Importantly, the anthranilate-inducible expression system worked also in strains of P. aeruginosa and P. putida and therefore will be most likely useful for physiological and biotechnological purposes in a wide range of pseudomonads. Key points • We established an anthranilate-inducible gene expression system for pseudomonads. • This system permits tuning of gene expression in a wide range of pseudomonads. • It will be very useful for physiological and biotechnological applications. Keywords Anthranilate . Recombinant expression . Gene regulation . Pseudomonas fluorescens . Pseudomonas aeruginosa
Introduction Pseudomonads play important roles in many ecosystems, and they often occur in mutualistic or pathogenic life styles associated with plants, animals, or humans (Peix et al. 2018). Prominent model organisms are, among others, the plant growth-promoting P. fluorescens, the human opportunistic pathogen P. aeruginosa, and P. putida, which is used for bioremediation and biotechnology (Diggle and Whiteley 2020; Kim and Anderson 2018; Poblete-Castro et al. 2012). Pseudomonads are frequently isolated and often there is the
* Thomas Brüser [email protected] 1
Institute of Microbiology, Leibniz University Hannover, Herrenhäuser Str. 2, 30419 Hannover, Germany
wish to produce functional proteins in these strains to study complementation, interaction, localization, transport, or other physiological aspects related to these proteins. For that purpose, there is a need for a regulated, tightly contr
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