Identification of MupP as a New Peptidoglycan Recycling Factor and Antibiotic Resistance Determinant in Pseudomonas aeruginosa

Peptidoglycan (PG) is an essential cross-linked polymer that surrounds most bacterial cells to prevent osmotic rupture of the cytoplasmic membrane. Its synthesis relies on penicillin-binding proteins, the targets of beta-lactam antibiotics. Many Gram-negative bacteria, including the opportunistic pathogen Pseudomonas aeruginosa, are resistant to beta-lactams because of a chromosomally encoded beta-lactamase called AmpC. In P. aeruginosa, expression of the ampC gene is tightly regulated and its induction is linked to cell wall stress. We reasoned that a reporter gene fusion to the ampC promoter would allow us to identify mutants defective in maintaining cell wall homeostasis and thereby uncover new factors involved in the process. A library of transposon-mutagenized P. aeruginosa was therefore screened for mutants with elevated ampC promoter activity. As an indication that the screen was working as expected, mutants with transposons disrupting the dacB gene were isolated. Defects in DacB have previously been implicated in ampC induction and clinical resistance to beta-lactam antibiotics. The screen also uncovered murU and PA3172 mutants that, upon further characterization, displayed nearly identical drug resistance and sensitivity profiles. We present genetic evidence that PA3172, renamed mupP, encodes the missing phosphatase predicted to function in the MurU PG recycling pathway that is widely distributed among Gram-negative bacteria.IMPORTANCE The cell wall biogenesis pathway is the target of many of our best antibiotics, including penicillin and related beta-lactam drugs. Resistance to these therapies is on the rise, particularly among Gram-negative species like Pseudomonas aeruginosa, a problematic opportunistic pathogen. To better understand how these organisms resist cell wall-targeting antibiotics, we screened for P. aeruginosa mutants defective in maintaining cell wall homeostasis. The screen identified a new factor, called MupP, involved in the recycling of cell wall turnover products. Characterization of MupP and other components of the pathway revealed that cell wall recycling plays important roles in both the resistance and the sensitivity of P. aeruginosa to cell wall-targeting antibiotics.