Evaluation of CpxRA as a Therapeutic Target for Uropathogenic Escherichia coli Infections

Abstract

CpxRA is an envelope stress response system found in all members of the family <i>Enterobacteriaceae</i>; CpxA has kinase activity for CpxR and phosphatase activity for phospho-CpxR, a transcription factor. CpxR also accepts phosphate groups from acetyl phosphate, a glucose metabolite. Activation of CpxR increases the transcription of genes encoding membrane repair and downregulates virulence determinants. We hypothesized that activation of CpxR could serve as an antimicrobial/antivirulence strategy and discovered compounds that activate CpxR in <i>Escherichia coli</i> by inhibiting CpxA phosphatase activity. As a prelude to testing such compounds <i>in vivo</i>, here we constructed <i>cpxA</i> (in the presence of glucose, CpxR is activated because of a lack of CpxA phosphatase) and <i>cpxR</i> (system absent) deletion mutants of uropathogenic <i>E. coli</i> (UPEC) CFT073. By RNA sequencing, few transcriptional differences were noted between the <i>cpxR</i> mutant and its parent, but in the <i>cpxA</i> mutant, several UPEC virulence determinants were downregulated, including the <i>fim</i> and <i>pap</i> operons, and it exhibited reduced mannose-sensitive hemagglutination of guinea pig red blood cells <i>in vitro</i> In competition experiments with mice, both mutants were less fit than the parent in the urine, bladder, and kidney; these fitness defects were complemented in <i>trans</i> Unexpectedly, in single-strain challenges, only the <i>cpxA</i> mutant was attenuated for virulence in the kidney but not in the bladder or urine. For the <i>cpxA</i> mutant, this may be due to the preferential use of amino acids over glucose as a carbon source in the bladder and urine by UPEC. These studies suggest that CpxA phosphatase inhibitors may have some utility for treating complex urinary tract infections.