Acidification of Cytoplasmic pH in Escherichia coli Provides a Strategy to Cope with Stress and Facilitates Development of Antibiotic Resistance

Abstract

Awareness of the problem of antimicrobial resistance (AMR) has escalated, and drug-resistant infections are named among the most urgent issues facing clinicians today. Bacteria can acquire resistance to antibiotics by a variety of mechanisms that, at times, involve changes in their metabolic status, thus altering diverse biochemical reactions, many of them pH-dependent. In this work, we found that modulation of the cytoplasmic pH (pHi) of Escherichia coli provides a thus far unexplored strategy to support resistance. We show here that the acidification of the cytoplasmic pH is a previously unrecognized consequence of the activation of the marRAB operon. The acidification itself contributes to the full implementation of the resistance phenotype. We measured the pHi of two resistant strains, developed in our laboratory, that carry mutations in marR that activate the marRAB operon. The pHi of both strains is lower than that of the wild type strain. Inactivation of the marRAB response in both strains weakens resistance, and pHi increases back to wild type levels. Likewise, we showed that exposure of wild type cells to weak acids that caused acidification of the cytoplasm induced a resistant phenotype, independent of the marRAB response. We speculate that the decrease of the cytoplasmic pH brought about by activation of the marRAB response provides a signaling mechanism that modifies metabolic pathways and serves to cope with stress and to lower metabolic costs. SIGNIFICANCEThe decreasing effectiveness of antibiotics in treating common infections has quickened in recent years, and resistance has spread worldwide. There is an urgent need to understand the mechanisms underlying acquisition and maintenance of resistance, and here we identify a novel element in the chain of events leading to a full-fledged clinically relevant state. The Escherichia coli multiple antibiotic resistance (mar) regulon is induced by a variety of signals and modulates the activity of dozens of target genes involved in resistance to antibiotics. We report here a thus far unidentified result of this activation: acidification of the cytoplasmic pH. Manipulation of the cytoplasmic pH with weak acids and bases, independently of the mar response, shows that the acidification significantly increases resistance.