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In E. coli, transcriptional activation is often mediated by the C-terminal domain of RpoA, the α subunit of RNA polymerase. Random mutations that prevent activation of the arabinose PBAD promoter are clustered in the RpoA C-terminal domain (α-CTD). We have isolated functional suppressors of rpoA α-CTD mutations that map to araC, the main transcriptional regulator of ara genes, or to the PBAD promoter. No mutation was found in the DNA regulatory region between araC and PBAD. Most suppressors that improve PBAD transcription are localized to the N-terminal domain of AraC. One class of araC mutations generates substitutions in the core of the N-terminal domain, suggesting that they affect its conformation. Other suppressors localize to the flexible N-terminal arm of AraC. Some, but not all, suppressors confer an arabinose constitutive phenotype. Suppression by both classes of araC mutations requires the α-CTD to stimulate expression from PBAD. Surprisingly, in rpoA+ strains lacking Crp, the cAMP receptor protein, these araC mutations largely restore arabinose gene expression and can essentially bypass Crp activation. Thus, the N-terminal domain of AraC exhibits at least three distinct activities: dimerization, arabinose binding, and transcriptional activation. Finally, one mutation maps to the AraC C-terminal domain and can synergize with AraC mutations in the N-terminal domain.

The acquisition of multidrug resistance (MDR) determinants jeopardizes treatment of bacterial infections with antibiotics. The tripartite efflux pump AcrAB-NodT confers adaptive MDR in the polarized α-proteobacterium Caulobacter crescentus via transcriptional induction by first-generation quinolone antibiotics. We discovered that overexpression of AcrAB-NodT by mutation or exogenous inducers confers resistance to cephalosporin and penicillin (β-lactam) antibiotics. Combining 2-step mutagenesis-sequencing (Mut-Seq) and cephalosporin-resistant point mutants, we dissected how TipR uses a common operator of the divergent tipR and acrAB-nodT promoter for adaptive and/or potentiated AcrAB-NodT-directed efflux. Chemical screening identified diverse compounds that interfere with DNA binding by TipR or induce its dependent proteolytic turnover. We found that long-term induction of AcrAB-NodT deforms the envelope and that homeostatic control by TipR includes co-induction of the DnaJ-like co-chaperone DjlA, boosting pump assembly and/or capacity in anticipation of envelope stress. Thus, the adaptive MDR regulatory circuitry reconciles drug efflux with co-chaperone function for trans-envelope assemblies and maintenance.