Biology subjects

Fernandes, S. E.

Publications and source records attributed to Fernandes, S. E..

3 recordsLinked to original sources

bioRxiv2026

Proteolytic feedback of quorum sensing promotes infection by Pseudomonas aeruginosa

Quorum sensing (QS) enables bacteria to coordinate collective behaviors in response to population density. However, the mechanisms by which QS circuits are reset or attenuated remain poorly understood. In Pseudomonas aeruginosa, the LuxR-type transcription factor RhlR controls virulence gene expression at high cell density, requiring both its cognate acyl-homoserine lactone signal and the accessory protein PqsE for full activity. Here, we show that RhlR abundance is actively controlled by ATP-dependent proteolysis through a self-limiting negative feedback loop. We identify an N-terminal degron within RhlR that is protected from protease recognition by PqsE binding and is required for degradation by both the Lon and the ClpP1-ClpP2 protease complex. Removal of this degron stabilizes RhlR, advancing the timing of QS-regulated gene expression, enhancing biofilm formation, imposes a competitive fitness cost in vitro, and reduced virulence in an acute mouse lung infection. Mechanistically, RhlR directly activates transcription of clpP2, encoding a subunit of the protease complex required for its own degradation, establishing a self-limiting negative feedback loop. Disruption of RhlR DNA binding stabilizes the receptor through dual effects on reduced clpP2 expression and increased PqsE availability via the loss of RhlR-dependent repression of pqsE. Together, these findings reveal proteolytic turnover as an underappreciated regulatory layer of QS-activation and establish a mechanism for how receptor turnover may facilitate the transition away from high-cell density states.

microbiology↗

bioRxiv2024

A chronic Pseudomonas aeruginosa mouse lung infection modeling the pathophysiology and inflammation of human cystic fibrosis

Investigation of chronic cystic fibrosis (CF) lung infections has been limited by a lack of murine models that reproduce obstructive lung pathology, chronicity of bacterial infections, and complex inflammation in human CF lung pathology. Three different approaches have been used separately to address these limitations, including using transgenic Scnn1b-Tg mice overexpressing a lung epithelial sodium channel to mimic the mucus-rich and hyperinflammatory CF lung environment, using synthetic CF sputum medium (SCFM) in an acute infection to induce bacterial phenotypes consistent with human CF, or using agar beads to promote chronic infections. Here, we combine these three models to establish a chronic Pseudomonas aeruginosa lung infection model using SCFM agar beads and Scnn1b-Tg mice (SCFM-Tg-mice) to recapitulate nutrients, mucus, and inflammation characteristic of the human CF lung environment. Like people with CF, SCFM-Tg-mice failed to clear bacterial infections. Lung function measurements showed that infected SCFM-Tg-mice had decreased inspiratory capacity and compliance, elevated airway resistance, and significantly reduced FVC and FEV0.1. Using spectral flow cytometry and multiplex cytokine arrays we show that, like people with CF, SCFM-Tg-mice developed inflammation characterized by eosinophil infiltration and Th2 lymphocytic cytokine responses. Chronically infected SCFM-Tg-mice developed an exacerbated mix of innate and Th1, Th2, and Th17-mediated inflammation, causing higher lung cellular damage, and elevated numbers of unusual Siglec F+ neutrophils. Thus, SCFM-Tg-mice represents a powerful tool to investigate bacterial pathogenesis and potential treatments for chronic CF lung infections and reveal a potential role for Siglec F+ neutrophils in CF inflammation. ImportanceHost-pathogen interaction studies of Pseudomonas aeruginosa cystic fibrosis (CF) lung infections have been hampered by limitations of mouse infection models. Here we combine synthetic CF sputum medium (SCFM) agar beads and Scnn1b-Tg transgenic mice to model the mucus obstructed airways and complex inflammatory characteristic of the human cystic fibrosis lung environment. In this model, which we name SCFM-Tg-mice, we use SCFM to cause changes in bacterial gene expression consistent with sputum collected from people with CF and the Scnn1b-Tg mice produce excessive airway mucus like people with CF. We show that SCMF-Tg-mice infected with P. aeruginosa have defects in lung function and increased inflammation that is consistent with human CF lung infections. This model can be adapted for other bacterial species and can be used to test hypotheses about bacterial pathogenesis and potential treatments in a CF human-like system.

microbiology↗

bioRxiv2020

Aging associated altered response to intracellular bacterial infections and its implication on the host

The effects of senescence and aging on geriatric diseases has been well explored but how these influence infections in the elderly have been scarcely addressed. Here, we show that several innate immune responses are elevated in senescent cells and old mice, allowing them to promptly respond to bacterial infections. We have identified higher levels of iNOS as a crucial host response and show that p38 MAPK in senescent cells acts as a negative regulator of iNOS transcription. In old mice, however the ability to impede bacterial proliferation does not correlate with increased survival as elevated immune responses persist unabated eventually affecting the host. The use of anti-inflammatory drugs that could consequently be recommended also decreases iNOS disarming the host of a critical innate immune response. Overall, our study highlights that infection associated mortality in the elderly is not merely an outcome of pathogen load but is also influenced by the hosts ability to resolve inflammation induced damage. Summary statementUsing cellular models and old mice we demonstrate the effect of aging on host response to bacterial infections. Aged systems mount a more effective anti-bacterial innate immune response but its persistence results in mortality of the host.

microbiology↗