Upregulation of PBP1B and LpoB in cysB Mutants Confers Mecillinam (Amdinocillin) Resistance in Escherichia coli

Mecillinam (amdinocillin) is a β-lactam antibiotic that inhibits the essential penicillin-binding protein 2 (PBP2). In clinical isolates of Escherichia coli from urinary tract infections, inactivation of the cysB gene (which encodes the main regulator of cysteine biosynthesis, CysB) is the major cause of resistance. How a nonfunctional CysB protein confers resistance is unknown, however, and in this study we wanted to examine the mechanism of resistance. Results show that cysB mutations cause a gene regulatory response that changes the expression of ∼450 genes. Among the proteins that show increased levels are the PBP1B, LpoB, and FtsZ proteins, which are known to be involved in peptidoglycan biosynthesis. Artificial overexpression of either PBP1B or LpoB in a wild-type E. coli strain conferred mecillinam resistance; conversely, inactivation of either the mrcB gene (which encodes PBP1B) or the lpoB gene (which encodes the PBP1B activator LpoB) made cysB mutants susceptible. These results show that expression of the proteins PBP1B and LpoB is both necessary and sufficient to confer mecillinam resistance. The addition of reducing agents to a cysB mutant converted it to full susceptibility, with associated downregulation of PBP1B, LpoB, and FtsZ. We propose a model in which cysB mutants confer mecillinam resistance by inducing a response that causes upregulation of the PBP1B and LpoB proteins. The higher levels of these two proteins can then rescue cells with mecillinam-inhibited PBP2. Our results also show how resistance can be modulated by external conditions such as reducing agents.