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An autoinducer-independent RhlR quorum-sensing receptor enables analysis of RhlR regulation.

Author(s): McCready, Amelia R; Paczkowski, Jon E; Cong, Jian-Ping; Bassler, Bonnie L

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Abstract: 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 (C4HSL), and the RhlR:C4HSL 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 C4HSL 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.
Publication Date: 13-Jun-2019
Citation: McCready, Amelia R, Paczkowski, Jon E, Cong, Jian-Ping, Bassler, Bonnie L. (2019). An autoinducer-independent RhlR quorum-sensing receptor enables analysis of RhlR regulation.. PLoS pathogens, 15 (6), e1007820 - ?. doi:10.1371/journal.ppat.1007820
DOI: doi:10.1371/journal.ppat.1007820
ISSN: 1553-7366
EISSN: 1553-7374
Pages: 1 - 23
Language: eng
Type of Material: Journal Article
Journal/Proceeding Title: PLoS pathogens
Version: Final published version. This is an open access article.

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