| Fluorescence-based reporter for gauging cyclic di-GMP levels in Pseudomonas aeruginosa. | |
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MedLine Citation:
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PMID: 22582064 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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The increased tolerance toward the host immune system and antibiotics displayed by biofilm-forming Pseudomonas aeruginosa and other bacteria in chronic infections such as cystic fibrosis bronchopneumonia is of major concern. Targeting of biofilm formation is believed to be a key aspect in the development of novel antipathogenic drugs that can augment the effect of classic antibiotics by decreasing antimicrobial tolerance. The second messenger cyclic di-GMP is a positive regulator of biofilm formation, and cyclic di-GMP signaling is now regarded as a potential target for the development of antipathogenic compounds. Here we describe the development of fluorescent monitors that can gauge the cellular level of cyclic di-GMP in P. aeruginosa. We have created cyclic di-GMP level reporters by transcriptionally fusing the cyclic di-GMP-responsive cdrA promoter to genes encoding green fluorescent protein. We show that the reporter constructs give a fluorescent readout of the intracellular level of cyclic di-GMP in P. aeruginosa strains with different levels of cyclic di-GMP. Furthermore, we show that the reporters are able to detect increased turnover of cyclic di-GMP mediated by treatment of P. aeruginosa with the phosphodiesterase inducer nitric oxide. Considering that biofilm formation is a necessity for the subsequent development of a chronic infection and therefore a pathogenicity trait, the reporters display a significant potential for use in the identification of novel antipathogenic compounds targeting cyclic di-GMP signaling, as well as for use in research aiming at understanding the biofilm biology of P. aeruginosa. |
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Authors:
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Morten T Rybtke; Bradley R Borlee; Keiji Murakami; Yasuhiko Irie; Morten Hentzer; Thomas E Nielsen; Michael Givskov; Matthew R Parsek; Tim Tolker-Nielsen |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2012-05-11 |
Journal Detail:
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Title: Applied and environmental microbiology Volume: 78 ISSN: 1098-5336 ISO Abbreviation: Appl. Environ. Microbiol. Publication Date: 2012 Aug |
Date Detail:
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Created Date: 2012-07-19 Completed Date: 2012-12-10 Revised Date: 2013-05-20 |
Medline Journal Info:
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Nlm Unique ID: 7605801 Medline TA: Appl Environ Microbiol Country: United States |
Other Details:
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Languages: eng Pagination: 5060-9 Citation Subset: IM |
Affiliation:
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Department of International Health, Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Adhesins, Bacterial
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genetics Biofilms* Cyclic GMP / analysis* Fluorescence* Genes, Reporter / genetics* Green Fluorescent Proteins / genetics Plasmids / genetics Promoter Regions, Genetic / genetics Pseudomonas aeruginosa / chemistry* |
| Grant Support | |
ID/Acronym/Agency:
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R01 AI077628/AI/NIAID NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Adhesins, Bacterial; 0/CdrA protein, Pseudomonas aeruginosa; 147336-22-9/Green Fluorescent Proteins; 7665-99-8/Cyclic GMP |
| Comments/Corrections | |
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