Modulating Microbial Resistance: Transcription Factor Regulation of Antibiotic Resistance in Escherichia coli

Abstract Introduction/Objective Antibiotic resistance is a global health emergency, with resistance detected to all antibiotics currently in clinical use and only a few novel drugs in the development pipeline. Antibiotics target key biological processes in bacteria, such as protein synthesis, cell wall synthesis, DNA replication and protein translation. Cells tightly control their internal states in different environments by regulating their transcriptional program through the activity of transcription factors (TFs) that control the expression of multiple downstream effector genes. We hypothesized that efficacy of specific antibiotics can be adjusted by altering the transcriptional states of bacteria prior to treatment, using TF knockout (KO) and over- expression (O-E) strains. Methods/Case Report Aim: Calculate the IC50 of 9 TF KO strains and 9 TF O-E strains across across a panel of representative antibiotics. A previous barcode screen study had identified TF KOs which were associated with antibiotic resistance/susceptibility. A serial antibiotic dilution was prepared in M9 media. E. coli TF KO and O-E strains were cultured for 24 hours in M9 media+Antibiotic (37°C, shaking). Each strain/antibiotic combination contained a technical replicate. Growth was measured by OD, allowing for calculation of growth curves, drug response curves, IC50, and EC50. Strain-specific IC50 and EC50 were compared to wild-type (WT) IC50 and EC50 as a measure of drug resistance/susceptibility. Results (if a Case Study enter NA) Experiments demonstrated consistent, TF-specific patterns of drug susceptibility. Compared to WT, TF KO strains demonstrated altered antibiotic IC50s (drug susceptibility). Compared to WT, TF O-E strains also demonstrated altered antibiotic IC50s (drug susceptibility). Conclusion Hypothesis Weakly Supported – TF states affect IC50s (Abx resistance). The relationships outlined in initial screen were consistently represented in experimental trials. Strain resistance/susceptibility is directly related to TF system and drug mechanism of action. For example, drugs that inhibited ribosomal function demonstrated greater effectiveness against KO/O-E strains that relied more heavily on protein synthesis pathways. Drug resistance is not binary but is shaped by a number of internal cellular factors, including gene regulation by TFs.