β-Lactam Resistance Response Triggered by Inactivation of a Nonessential Penicillin-Binding Protein

Abstract

It has long been recognized that the modification of penicillin-binding proteins (PBPs) to reduce their affinity for β-lactams is an important mechanism (target modification) by which Gram-positive cocci acquire antibiotic resistance. Among Gram-negative rods (GNR), however, this mechanism has been considered unusual, and restricted to clinically irrelevant laboratory mutants for most species. Using as a model Pseudomonas aeruginosa, high up on the list of pathogens causing life-threatening infections in hospitalized patients worldwide, we show that PBPs may also play a major role in β-lactam resistance in GNR, but through a totally distinct mechanism. Through a detailed genetic investigation, including whole-genome analysis approaches, we demonstrate that high-level (clinical) β-lactam resistance in vitro, in vivo, and in the clinical setting is driven by the inactivation of the dacB-encoded nonessential PBP4, which behaves as a trap target for β-lactams. The inactivation of this PBP is shown to determine a highly efficient and complex β-lactam resistance response, triggering overproduction of the chromosomal β-lactamase AmpC and the specific activation of the CreBC (BlrAB) two-component regulator, which in turn plays a major role in resistance. These findings are a major step forward in our understanding of β-lactam resistance biology, and, more importantly, they open up new perspectives on potential antibiotic targets for the treatment of infectious diseases. Decades after their discovery, β-lactams remain key components of our antimicrobial armamentarium for the treatment of infectious diseases. Nevertheless, resistance to these antibiotics is increasing alarmingly. There are two major bacterial strategies to develop resistance to β-lactam antibiotics: the production of enzymes that inactivate them (β-lactamases), or the modification of their targets in the cell wall (the essential penicillin-binding proteins, PBPs). Using the pathogen Pseudomonas aeruginosa as a model microorganism, we show that high-level (clinical) β-lactam resistance in vitro and in vivo frequently occurs through a previously unrecognized, totally distinct resistance pathway, driven by the mutational inactivation of a nonessential PBP (PBP4) that behaves as a trap target for β-lactams. We show that mutation of this PBP determines a highly efficient and complex β-lactam resistance response, triggering overproduction of the chromosomal β-lactamase AmpC and the specific activation of a two-component regulator, which in turn plays a key role in resistance. These findings are a major step forward in our understanding of β-lactam resistance biology, and, more importantly, they open up new perspectives on potential antibiotic targets for the treatment of infectious diseases.