# A New Quorum-Sensing Autoinducer Acts with the RhlR Receptor to Control Virulence and Biofilms in Pseudomonas Aeruginosa

> **NIH NIH R00** · UNIVERSITY OF CHICAGO · 2021 · $248,945

## Abstract

PROJECT SUMMARY
Hospital-acquired secondary infections are an escalating problem of global significance. Indeed, multi-drug
resistant Pseudomonas aeruginosa is the leading cause of hospital-acquired infections in the USA and P.
aeruginosa is now a priority pathogen on the CDC ESKAPE pathogen list. P. aeruginosa infection is a particular
problem in cystic fibrosis, microbial keratitis, in third-degree burn units, and in cancer sufferers and HIV patients.
P. aeruginosa virulence and biofilm development depend on the bacterial cell-to-cell communication process
called quorum sensing. The known P. aeruginosa quorum-sensing circuit possesses two canonical LuxI/R type
signaling pathways: LasI/R and RhlI/R, that, together, control an estimated 10% of the genes in the genome.
The known circuit functions as follows: LasI produces and LasR responds to the autoinducer 3OC12-homoserine
lactone. The LasR:3OC12-homoserine lactone complex activates transcription of many genes including rhlR,
encoding a second quorum-sensing receptor. RhlR binds to the autoinducer C4-homoserine lactone, the product
of RhlI. RhlR:C4-homoserine lactone also directs a large regulon of genes including those encoding virulence
factors such as pyocyanin, elastases, rhamnolipids and genes required for biofilm formation. Typically, mutations
in quorum-sensing luxI-type and luxR-type genes (i.e., lasI-lasR and rhlI-rhlR) confer identical phenotypes
because each component of the pair needs the other to function. However, using biofilm analyses, transcriptional
reporter assays, RNA-seq studies, and animal infection assays, I discovered that RhlR directs both RhlI-
dependent and RhlI-independent regulons. Importantly, I showed that ΔrhlI mutant cell-free culture fluids, i.e.,
that lack C4-homoserine lactone, contain an activity that stimulates RhlR-dependent gene expression. I
hypothesize that RhlR responds to an alternative ligand, in addition to the traditional C4-homoserine lactone
autoinducer. Supporting this notion, I showed that the enzyme PqsE is required for alternative ligand synthesis.
Finally, I demonstrated that while the RhlR-RhlI system is dispensable, the RhlR-PqsE system is the crucial
quorum-sensing system required for biofilm formation and for virulence in two animal models of infection. Here,
I propose to 1) determine the chemical structure of the alternative ligand and define how it interacts with RhlR;
2) characterize the PqsE active site, discover small molecule inhibitors of PqsE, and identify and characterize
additional factors involved in alternative ligand synthesis; 3) map the alternative ligand and/or C4-homoserine
lactone-dependent RhlR regulon(s) required for biofilm development at the single cell level and at the level of
community. The proposed research will contribute significant insights about the chemical lexicon used by a
clinically important bacterium in biofilms and in disease. Moreover, the proposed research will provide a
mechanistic understanding of ...

## Key facts

- **NIH application ID:** 10247826
- **Project number:** 5R00GM129424-04
- **Recipient organization:** UNIVERSITY OF CHICAGO
- **Principal Investigator:** Sampriti Mukherjee
- **Activity code:** R00 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $248,945
- **Award type:** 5
- **Project period:** 2018-09-01 → 2023-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10247826

## Citation

> US National Institutes of Health, RePORTER application 10247826, A New Quorum-Sensing Autoinducer Acts with the RhlR Receptor to Control Virulence and Biofilms in Pseudomonas Aeruginosa (5R00GM129424-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10247826. Licensed CC0.

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