The anti-sigma factor MucA of Pseudomonas aeruginosa: Dramatic differences of a mucA22 vs. a ΔmucA mutant in anaerobic acidified nitrite sensitivity of planktonic and biofilm bacteria in vitro and during chronic murine lung infection

Mucoid mucA22 Pseudomonas aeruginosa (PA) is an opportunistic lung pathogen of cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD) patients that is highly sensitive to acidified nitrite (A-NO(2)(-)). In this study, we first screened PA mutant strains for sensitivity or resistance to 20 mM A-NO(2)(-) under anaerobic conditions that represent the chronic stages of the aforementioned diseases. Mutants found to be sensitive to A-NO(2)(-) included PA0964 (pmpR, PQS biosynthesis), PA4455 (probable ABC transporter permease), katA (major catalase, KatA) and rhlR (quorum sensing regulator). In contrast, mutants lacking PA0450 (a putative phosphate transporter) and PA1505 (moaA2) were A-NO(2)(-) resistant. However, we were puzzled when we discovered that mucA22 mutant bacteria, a frequently isolated mucA allele in CF and to a lesser extent COPD, were more sensitive to A-NO(2)(-) than a truncated ΔmucA deletion (Δ157–194) mutant in planktonic and biofilm culture, as well as during a chronic murine lung infection. Subsequent transcriptional profiling of anaerobic, A-NO(2)(-)-treated bacteria revealed restoration of near wild-type transcript levels of protective NO(2)(-) and nitric oxide (NO) reductase (nirS and norCB, respectively) in the ΔmucA mutant in contrast to extremely low levels in the A-NO(2)(-)-sensitive mucA22 mutant. Proteins that were S-nitrosylated by NO derived from A-NO(2)(-) reduction in the sensitive mucA22 strain were those involved in anaerobic respiration (NirQ, NirS), pyruvate fermentation (UspK), global gene regulation (Vfr), the TCA cycle (succinate dehydrogenase, SdhB) and several double mutants were even more sensitive to A-NO(2)(-). Bioinformatic-based data point to future studies designed to elucidate potential cellular binding partners for MucA and MucA22. Given that A-NO(2)(-) is a potentially viable treatment strategy to combat PA and other infections, this study offers novel developments as to how clinicians might better treat problematic PA infections in COPD and CF airway diseases.