High-throughput identification and rational design of synergistic small-molecule pairs for combating and bypassing antibiotic resistance
Fig 4
The SOS response is induced by trimethoprim and azidothymidine (AZT).
(A) SOS response measured by a green fluorescent protein (GFP) reporter gene under control of the sulA reporter. Small molecules were added at 50% minimum inhibitory concentration (MIC). Expected sulA induction (green) is either the induction by the second molecule or, if the sulA reporter is repressed by the second molecule, no induction or repression. Trimethoprim does not induce the sulA reporter, so it is considered to not have any contribution to the expected value, and we do not simply sum the induction of trimethoprim + molecule #2. The observed sulA induction (purple) is significantly higher than expected in the trimethoprim and AZT combination but not nonsynergistic combinations, such as trimethoprim + hydroxyurea or trimethoprim + rifampicin. Since the trimethoprim + sulfamethizole combination does not induce sulA, the molecular mechanisms underlying trimethoprim + sulfamethizole synergy likely differ from trimethoprim + AZT synergy. Significance was calculated using a Mann-Whitney test. Error bars represent the standard deviation. In all cases when we observed a significant difference between expected and observed, we also found a significant difference between induction by molecule #2 and induction in the combination. The data for these graphs are in S7 Table. (B) Fluctuation assay measures the mutation rate following small-molecule treatment. Small molecules alone (solid colors) do not significantly increase mutation rate. Trimethoprim combined (striped bars) with synergistic partners AZT or mitomycin C increases mutation rate. P values were calculated using Fisher’s exact test. Error bars represent the 95% confidence interval. The data for these graphs are in S8 Table.