3-O-deacylation of lipid A by PagL, a PhoP/PhoQ-regulated deacylase of Salmonella typhimurium, modulates signaling through Toll-like receptor 4

Toll-like receptor 4 (TLR4)-mediated responses, which are induced by the lipid A portion of lipopolysaccharide, are important for host defense against Salmonellae infection. A variety of different data indicate that the acylation state of lipid A can alter TLR4-mediated responses. The S. typhimurium virulence gene product PhoP/PhoQ signals the presence of host microenvironments to regulate the expression of a lipid A 3-O-deacylase, PagL, and a lipid A palmitoyltransferase, PagP. We now demonstrate that 3-O-deacylation and palmitoylation of lipid A decreases its ability to induce TLR4-mediated signaling. Deacylated lipid A, deacylated and palmitoylated lipid A, palmitoylated lipid A, and unmodified lipid A species were purified from Escherichia coli heterologously expressing PagL and/or PagP. The purified lipid A preparations showed spectra of a single lipid A species on mass spectrometry and gave a single band on thin layer chromatography. The activity of purified lipid A species was examined using human and mouse cell lines that express recombinant human TLR4. Compared with unmodified lipid A, the modified lipid A species are 30-100-fold less active in the ability to induce NF-kappaB-dependent reporter activation. These results suggest that the lipid A modifications reduce TLR4-signaling as part of Salmonellae adaptation to host environments.