PERSONAL DE APOYO
FERNÁNDEZ Jorge GermÁn
congresos y reuniones científicas
Título:
IDENTIFICATION OF ESSENTIAL FACTORS INVOLVED IN THE RELATIONSHIP BE- TWEEN THE SIGMA FACTOR ALGT AND ITS ANTI-SIGMA MUCA IN THE ACIDIFIED NI- TRITE SENSITIVITY DURING ANAEROBIC GROWTH IN PSEUDOMONAS AERUGINOSA
Autor/es:
HEDEMANN, GABRIELA; LÓPEZ, VERÓNICA; FERNÁNDEZ, JORGE GERMÁN; TRIBELLI, PAULA; LÓPEZ, NANCY; MOYANO, ALEJANDRO; SMANIA, ANDREA
Reunión:
Congreso; XVII Congreso Argentino de Microbiología General; 2022
Institución organizadora:
Sociedad Argentina de Microbiología General
Resumen:
Pseudomonas aeruginosa is an opportunistic pathogen that chronically infects the airways of cystic fibrosis (CF) patients. Mucoid, mucA mutant, phenotype marks the onset of chronic infection and constitutes a sign of poor prognosis. The mucA gene is an anti-σ fac- tor that negatively regulates alginate production by sequestration of AlgT, an alternative σ factor responsible for the transcription of the alginate biosynthetic operon. The most frequent mutation responsible for mucoid conversion is a deletion of a G residue within a homopolymeric track of five Gs (G5426), also known as mucA22 allele, causing the truncation of MucA C-terminal periplasmic domain. Mutations in mucA gene can also affect major determinants for bacterial persistence such as quorum-sensing (QS) signals, flagel- lum biosynthesis or survival under anaerobic and osmotic stress conditions. Evidence has shown that mucoid mucA22 is highly sensitive to acidified nitrite (A-NO2-) perishing during anaerobic exposure to 15Mm nitrite at pH 6.5. Previously, we confirmed mucA22 sensitivity to nitrite and demonstrated that this phenotype was restored to almost wild-type levels upon algT deletion, indicating that it might be consequence of the σ factor deregulation. To better understand the relationship between algT-mucA22 and NO2- sensitivity during anaerobic growth, we performed a proteomic analysis using algTmucA (WT) strain and ΔalgTmucA22 and mucA22 mutants that were treated and non-treated with A-NO2- under anaerobic conditions. For protein extracts, we performed a subcellular fractiona- tion from cytoplasm, periplasm and membrane. Quantitative liquid chromatography - tandem mass spectrometry (LC-MS/MS) analysis was performed using a Q-Exactive Orbitrap Mass Spectrometers. Protein identification was achieved using Proteome Discoverer Software version 2.2 (Thermo Scientific). As expected, when bacteria were NO2- treated, proteins involved in processes such as cell elongation, DNA biosynthesis, and peptidoglycan biosynthesis were downregulated. Instead, proteins from biofilm formation, stress response and membrane vesicle formation were upregulated. Importantly, whereas the mucA22 strain showed a decreased expression of NirS and NirF proteins, essential factors for nitrite reduction to nitric oxide, in both, treated and non-treated condition, the ex- pression of these factors in the ΔalgTmucA22 mutant was significantly increased, which may explain the bases of A-NO2- sensitivity and anaerobic growth impairment upon AlgT deregulation. A-NO2-sensitivity complementation in mucA22 strain by nirS and nirF ex- pression are currently carrying out. Our work shed light on the complex regulatory pathways connecting mucoid conversion and anaerobic growth, providing potential targets for future therapeutic strategies to control chronic P. aeruginosa.