The Rcs-Phosphorelay Pathway Is Not Essential for Intrinsic Antibiotic Resistance to β-lactam Antibiotics in Escherichia coli

Rcs phosphorelay pathway in Escherichia coli is a response system that senses outer membrane stress conditions and regulating capsule and biofilm formation which can result in antibiotic resistance. The Rcs phosphorelay system consists of the outer membrane sensor protein RcsF, a sensor kinase RcsC, a response
regulator RcsB, and a non-coding RNA rprA. RcsF signals downstream of the Rcs pathway to regulate expression of the Rcs regulon, including rprA, which induces the activity of the general stress response regulator RpoS. The role of RcsB and RcsF in regulating intrinsic antibiotic resistance remains unclear. Here we address this question by testing the minimum inhibitory antibiotic concentrations to four β-lactam antibiotics in E. coli strains DH311 and BW25113 which bear chromosomal deletions of rcsB and rcsF, respectively. We hypothesized that the disruption of the Rcs pathway would decrease RpoS expression, which in turn increases the antibiotic susceptibility in the mutant strains compared to wild type E. coli. We also predicted a correlation between increased rprA expression levels and antibiotic resistance. E. coli strains designed with an rprA::lacZ fusion were tested via β-galactosidase assay to measure rprA expression after treatment with β-lactam antibiotics. We found that the wild type and the mutant strains of RcsB displayed the identical levels of intrinsic antibiotic resistance against cefsulodin, ampicillin, phosphomycin, and penicillin. Wild type E. coli strain DH300 showed 5 times higher rprA expression compared to an rcsB deletion mutant when treated with antibiotics and a 1.5 times higher level of rprA expression compared to the untreated sample. These data suggest that RcsB directly contributes to rprA expression
but deletion of RcsB or RcsF does not result in a detectable difference in the intrinsic antibiotic susceptibility of E. coli to β-lactams. Our data provides information on further understanding the contributions of the Rcs pathway to the regulation of E. coli stress response regulator RpoS.