From drug export to envelope fitness: characterizing the regulatory landscape of selective Escherichia coli ABC efflux pumps
TL;DR
The data show that ABC pumps are tightly integrated into a regulatory network governing envelope stress, osmotic adaptation, and biofilm formation—not just drug export.
Abstract
Background Antimicrobial resistance remains a top global health threat. While ATP-binding cassette (ABC) efflux pumps contribute to multidrug resistance in Gram-negative bacteria, their transcriptional regulation is far less understood than that of the RND and MFS families. Methods We used DNA pull-down followed by LC–MS/MS to identify proteins binding to the promoters of five E. coli ABC efflux pump genes (msbA, macB, ybhF, yddA, and yadG). Candidate regulators were analyzed using bioinformatics, domain scanning, and molecular docking. Results Binding patterns differed sharply across the five genes. MsbA, macB, and ybhF pulled down complex interaction networks, while yddA and yadG bound far fewer regulators. CpxR was among the most highly enriched regulatory proteins, recovered at the msbA, macB, and ybhF promoters with fold changes of 30.57, 57.17, and 17.39, respectively. OmpR and ArcA exhibited significant enrichment at the ybhF promoter (FC = 64.77 and 238.28, respectively). Both regulators are implicated in osmotic and redox signaling, suggesting a potential link to ybhF transcriptional control under such conditions. LrhA was prominently enriched at the yddA promoter (FC = 103.21). This observation raises the possibility of a connection between LrhA and biofilm-related regulatory pathways, which remains to be validated experimentally. Structurally, the ybhFSR operon is organized as a split-type system: ybhF encodes the nucleotide-binding domains (NBDs), while ybhS and ybhR encode the transmembrane domains (TMDs). The system likely assembles with YbhG and TolC into a tripartite complex. By contrast, based on structural prediction, yddA and yddB may form a minimal two-subunit ABC transporter, marked by atypical Walker A (GYSGAGKTT) and ABC signature (LSSGE) motifs. Conclusion Our data show that ABC pumps are tightly integrated into a regulatory network governing envelope stress, osmotic adaptation, and biofilm formation—not just drug export. These findings identify candidate regulators whose putative promoter binding requires experimental validation.
