TY - JOUR

T1 - A tunable anthranilate-inducible gene expression system for Pseudomonas species

AU - Hoffmann, Lena

AU - Sugue, Michael Frederick

AU - Brüser, Thomas

N1 - Funding Information: We thank Katrin Gunka for manifold suggestions and excellent technical assistance, and Jochen Meens and Nina Janze of the Institute of Microbiology, University of Veterinary Medicine Hannover (TiHo) for the support with flow cytometry. We are grateful to Michael Ringel for the support in early phases of this project. We thank Joyce E. Loper (Oregon State University) for kindly providing the strain P. fluorescens A506, and we also thank Susanne H?ussler (Institute of Molecular Bacteriology, TWINCORE, Hannover & Helmholtz Centre for Infection Research, Braunschweig) for donation of the vector pUCP20 and P. aeruginosa PAO1.

PY - 2021/1

Y1 - 2021/1

N2 - 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.

AB - 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.

KW - Anthranilate

KW - Gene regulation

KW - Pseudomonas aeruginosa

KW - Pseudomonas fluorescens

KW - Recombinant expression

KW - Gene expression

KW - Pseudomonas fluorescens

KW - Pseudomonas aeruginosa

KW - Recombinant expression

UR - http://www.scopus.com/inward/record.url?scp=85097172309&partnerID=8YFLogxK

U2 - 10.1007/s00253-020-11034-8

DO - 10.1007/s00253-020-11034-8

M3 - Article

C2 - 33270152

AN - SCOPUS:85097172309

VL - 105

SP - 247

EP - 258

JO - Applied Microbiology and Biotechnology

JF - Applied Microbiology and Biotechnology

SN - 0175-7598

ER -