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An autoinducer-independent RhlR quorum-sensing receptor enables analysis of RhlR regulation
Quorum sensing is a chemical communication process that bacteria use to coordinate group behaviors. Pseudomonas aeruginosa, an opportunistic pathogen, employs multiple quorum-sensing systems to control behaviors including virulence factor production and biofilm formation. One P. aeruginosa quorum-se...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2019
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6564026/ https://www.ncbi.nlm.nih.gov/pubmed/31194839 http://dx.doi.org/10.1371/journal.ppat.1007820 |
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author | McCready, Amelia R. Paczkowski, Jon E. Cong, Jian-Ping Bassler, Bonnie L. |
author_facet | McCready, Amelia R. Paczkowski, Jon E. Cong, Jian-Ping Bassler, Bonnie L. |
author_sort | McCready, Amelia R. |
collection | PubMed |
description | Quorum sensing is a chemical communication process that bacteria use to coordinate group behaviors. Pseudomonas aeruginosa, an opportunistic pathogen, employs multiple quorum-sensing systems to control behaviors including virulence factor production and biofilm formation. One P. aeruginosa quorum-sensing receptor, called RhlR, binds the cognate autoinducer N-butryl-homoserine lactone (C(4)HSL), and the RhlR:C(4)HSL complex activates transcription of target quorum-sensing genes. Here, we use a genetic screen to identify RhlR mutants that function independently of the autoinducer. The RhlR Y64F W68F V133F triple mutant, which we call RhlR*, exhibits ligand-independent activity in vitro and in vivo. RhlR* can drive wildtype biofilm formation and infection in a nematode animal model. The ability of RhlR* to properly regulate quorum-sensing-controlled genes in vivo depends on the quorum-sensing regulator RsaL keeping RhlR* activity in check. RhlR is known to function together with PqsE to control production of the virulence factor called pyocyanin. Likewise, RhlR* requires PqsE for pyocyanin production in planktonic cultures, however, PqsE is dispensable for RhlR*-driven pyocyanin production on surfaces. Finally, wildtype RhlR protein is not sufficiently stabilized by C(4)HSL to allow purification. However, wildtype RhlR can be stabilized by the synthetic ligand mBTL (meta-bromo-thiolactone) and RhlR* is stable without a ligand. These features enabled purification of the RhlR:mBTL complex and of RhlR* for in vitro examination of their biochemical activities. To our knowledge, this work reports the first RhlR protein purification. |
format | Online Article Text |
id | pubmed-6564026 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-65640262019-06-20 An autoinducer-independent RhlR quorum-sensing receptor enables analysis of RhlR regulation McCready, Amelia R. Paczkowski, Jon E. Cong, Jian-Ping Bassler, Bonnie L. PLoS Pathog Research Article Quorum sensing is a chemical communication process that bacteria use to coordinate group behaviors. Pseudomonas aeruginosa, an opportunistic pathogen, employs multiple quorum-sensing systems to control behaviors including virulence factor production and biofilm formation. One P. aeruginosa quorum-sensing receptor, called RhlR, binds the cognate autoinducer N-butryl-homoserine lactone (C(4)HSL), and the RhlR:C(4)HSL complex activates transcription of target quorum-sensing genes. Here, we use a genetic screen to identify RhlR mutants that function independently of the autoinducer. The RhlR Y64F W68F V133F triple mutant, which we call RhlR*, exhibits ligand-independent activity in vitro and in vivo. RhlR* can drive wildtype biofilm formation and infection in a nematode animal model. The ability of RhlR* to properly regulate quorum-sensing-controlled genes in vivo depends on the quorum-sensing regulator RsaL keeping RhlR* activity in check. RhlR is known to function together with PqsE to control production of the virulence factor called pyocyanin. Likewise, RhlR* requires PqsE for pyocyanin production in planktonic cultures, however, PqsE is dispensable for RhlR*-driven pyocyanin production on surfaces. Finally, wildtype RhlR protein is not sufficiently stabilized by C(4)HSL to allow purification. However, wildtype RhlR can be stabilized by the synthetic ligand mBTL (meta-bromo-thiolactone) and RhlR* is stable without a ligand. These features enabled purification of the RhlR:mBTL complex and of RhlR* for in vitro examination of their biochemical activities. To our knowledge, this work reports the first RhlR protein purification. Public Library of Science 2019-06-13 /pmc/articles/PMC6564026/ /pubmed/31194839 http://dx.doi.org/10.1371/journal.ppat.1007820 Text en © 2019 McCready et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article McCready, Amelia R. Paczkowski, Jon E. Cong, Jian-Ping Bassler, Bonnie L. An autoinducer-independent RhlR quorum-sensing receptor enables analysis of RhlR regulation |
title | An autoinducer-independent RhlR quorum-sensing receptor enables analysis of RhlR regulation |
title_full | An autoinducer-independent RhlR quorum-sensing receptor enables analysis of RhlR regulation |
title_fullStr | An autoinducer-independent RhlR quorum-sensing receptor enables analysis of RhlR regulation |
title_full_unstemmed | An autoinducer-independent RhlR quorum-sensing receptor enables analysis of RhlR regulation |
title_short | An autoinducer-independent RhlR quorum-sensing receptor enables analysis of RhlR regulation |
title_sort | autoinducer-independent rhlr quorum-sensing receptor enables analysis of rhlr regulation |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6564026/ https://www.ncbi.nlm.nih.gov/pubmed/31194839 http://dx.doi.org/10.1371/journal.ppat.1007820 |
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