Genetic evidence for functional diversification of gram-negative intermembrane phospholipid transporters

Abstract

The gram-negative outer membrane protects against environmental stress and antibiotics. Phospholipid transport to this membrane is under intense investigation and, in Escherichia coli, YhdP, TamB, and YdbH have recently been shown to mediate this transport. However, strains without the main transporters, YhdP and TamB, have different phenotypes, demonstrating distinct functions. We investigated these functions using the synthetic cold sensitivity of a deletion strain of yhdP and fadR, a fatty acid metabolism regulator. Cold sensitivity is suppressed by ΔtamB forcing transport to YdbH and demonstrating TamB dysfunction. Increased cardiolipin and fatty acid saturation are necessary for cold sensitivity and lowering either suppresses the sensitivity. Our data and simulations support a model where YhdP transports more saturated phospholipids and TamB transports more unsaturated phospholipids, allowing independent regulation of inner and outer membrane saturation by altering the YhdP:TamB activity ratio and providing a mechanism for maintaining outer membrane homeostasis under changing conditions.

The translocation assembly module (TAM) catalyzes the assembly of bacterial outer membrane proteinsin vitro

“…A second possibility is that partially folded OMPs “piggyback” on phospholipids (or vice-versa) and both types of molecules reach the OM together. Finally, in light of recent evidence that TamB and YhdP primarily transport distinct classes of lipids and that the phospholipid composition of the OM is regulated by changing the relative activity of each protein 102 , it is also possible that TAM transports OMPs when it is not needed for lipid transport.…”

Genetic interaction mapping reveals functional relationships between peptidoglycan endopeptidases and carboxypeptidases

“…Among the conditionally essential genes, this screen revealed PG synthesis and recycling factors such as the main PBP, pbp1A, its accessory factor csiV [13], and several PG recycling pathway components, namely the muropeptide ligase mpl, the muropeptide fragment permease ampG [14] and the lytic transglycosylase mltB (Fig 1B). Another synthetic lethal hit was yhdP, which likely fulfils phospholipid transport functions to the OM, with a preference for saturated fatty acids [15]. ShyA L109K overexpression causes spheroplast formation, and spheroplast survival has been shown to depend on YhdP [16]; this hit may thus suggest that saturated fatty acids contribute to spheroplast stability.…”