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Establishment of a high-throughput setup for screening small molecules that modulate c-di-GMP signaling in pseudomonas aeruginosa

Establishment of a high-throughput setup for screening small molecules that modulate c-di-GMP signaling in pseudomonas aeruginosa
Establishment of a high-throughput setup for screening small molecules that modulate c-di-GMP signaling in pseudomonas aeruginosa

Bacterial resistance to traditional antibiotics has driven research attempts to identify new drug targets in recently discovered regulatory pathways. Regulatory systems that utilize intracellular cyclic di-GMP (c-di-GMP) as a second messenger are one such class of target. c-di-GMP is a signaling molecule found in almost all bacteria that acts to regulate an extensive range of processes including antibiotic resistance, biofilm formation and virulence. The understanding of how c-di-GMP signaling controls aspects of antibiotic resistant biofilm development has suggested approaches whereby alteration of the cellular concentrations of the nucleotide or disruption of these signaling pathways may lead to reduced biofilm formation or increased susceptibility of the biofilms to antibiotics. We describe a simple high-throughput bioreporter protocol, based on green fluorescent protein (GFP), whose expression is under the control of the c-di-GMP responsive promoter cdrA, to rapidly screen for small molecules with the potential to modulate c-di-GMP cellular levels in Pseudomonas aeruginosa (P. aeruginosa). This simple protocol can screen upwards of 3,500 compounds within 48 hours and has the ability to be adapted to multiple microorganisms.

Antibiotic resistance, Biofilm formation, Cyclic di-GMP signaling, Green fluorescent protein reporter, High-throughput screen, Issue 112, Microbiology, Pseudomonas aeruginosa
1940-087X
Rugjee, Kushal N.
38d15aab-f64f-4a4f-9a44-f4489a191b33
An, Shi Qi
0e05f480-cec1-4c0e-bc1d-359d30ea9a6e
Ryan, Robert P.
cd9f1e35-9ffe-456f-a64e-798b1f520298
Rugjee, Kushal N.
38d15aab-f64f-4a4f-9a44-f4489a191b33
An, Shi Qi
0e05f480-cec1-4c0e-bc1d-359d30ea9a6e
Ryan, Robert P.
cd9f1e35-9ffe-456f-a64e-798b1f520298

Rugjee, Kushal N., An, Shi Qi and Ryan, Robert P. (2016) Establishment of a high-throughput setup for screening small molecules that modulate c-di-GMP signaling in pseudomonas aeruginosa. Journal of Visualized Experiments, 2016 (112), [e54115]. (doi:10.3791/54115).

Record type: Article

Abstract

Bacterial resistance to traditional antibiotics has driven research attempts to identify new drug targets in recently discovered regulatory pathways. Regulatory systems that utilize intracellular cyclic di-GMP (c-di-GMP) as a second messenger are one such class of target. c-di-GMP is a signaling molecule found in almost all bacteria that acts to regulate an extensive range of processes including antibiotic resistance, biofilm formation and virulence. The understanding of how c-di-GMP signaling controls aspects of antibiotic resistant biofilm development has suggested approaches whereby alteration of the cellular concentrations of the nucleotide or disruption of these signaling pathways may lead to reduced biofilm formation or increased susceptibility of the biofilms to antibiotics. We describe a simple high-throughput bioreporter protocol, based on green fluorescent protein (GFP), whose expression is under the control of the c-di-GMP responsive promoter cdrA, to rapidly screen for small molecules with the potential to modulate c-di-GMP cellular levels in Pseudomonas aeruginosa (P. aeruginosa). This simple protocol can screen upwards of 3,500 compounds within 48 hours and has the ability to be adapted to multiple microorganisms.

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e-pub ahead of print date: 30 June 2016
Published date: 30 June 2016
Keywords: Antibiotic resistance, Biofilm formation, Cyclic di-GMP signaling, Green fluorescent protein reporter, High-throughput screen, Issue 112, Microbiology, Pseudomonas aeruginosa

Identifiers

Local EPrints ID: 425824
URI: http://eprints.soton.ac.uk/id/eprint/425824
ISSN: 1940-087X
PURE UUID: e9294eba-de42-4a25-ba05-fb6d6dd73121

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Date deposited: 05 Nov 2018 17:30
Last modified: 05 Jun 2024 17:29

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Contributors

Author: Kushal N. Rugjee
Author: Shi Qi An
Author: Robert P. Ryan

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