Deciphering the role of the OmpC-Mla system in bile salt resistance

The outer membrane (OM) of Gram-negative bacteria presents a formidable barrier against external insults, in part contributing to the survival of Enterobacteriaceae in the mammalian gut. The lipid asymmetry of the OM, where lipopolysaccharides (LPS) form a tight outer layer of low permeability, effectively restricts the passage of toxic substances across the bilayer. In the gut, however, bile salts pose a unique challenge to the bacterial cell due to their ability to form micelles and solubilize membranes; yet, mechanisms to prevent dissolution of the OM by such detergents are not well understood. In this study, we define a distinct role in bile salt resistance for the OmpC-Mla system in Escherichia coli, which is better known for its function in maintaining OM lipid asymmetry. We show that cells lacking a functional OmpC-Mla system are sensitive to bile salts, but only at or above critical micellar concentrations. Furthermore, we observe that these cells still exhibit bile salt sensitivity even when defects in OM lipid asymmetry have been corrected, suggesting that the OmpC-Mla system contributes to bile salt resistance independent of its role in lipid asymmetry. Finally, we demonstrate that MlaD, one of the key lipid-binding components of the system, displays specific binding to bile salts in vitro. Since the OmpC-Mla system maintains OM lipid asymmetry by transporting mislocalized PLs, our findings here support a model where this system also additionally removes bile salts that have intercalated into the OM, to ultimately prevent dissolution and disruption of this important barrier. IMPORTANCEBile salts are important components secreted into the human gut to help solubilize fats from our diet, yet they also possess anti-microbial properties due to their corresponding ability to dissolve bacterial membranes. For Enterobacteriaceae to survive in the gut environment, these bacterial cells must prevent intracellular build-up of bile salts by either restricting entry or by pumping out these molecules. Ultimately, they must resist bile salt-mediated dissolution of their membranes, particularly their outer membranes, which serve as a protective barrier against toxic substances. In this study, we reveal that a known lipid transport system in Escherichia coli has a distinct role in bile salt resistance independent of its role in maintaining outer membrane lipid asymmetry; it does so likely by removing bile salts from the outer membrane, thus preventing dissolution. Our work highlights the possibility of targeting this lipid transport system for the treatment of Enterobacteriaceae infections.