PubMed · 2497962
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R C Judd. 1989. Protein I: structure, function, and genetics.. https://doi.org/10.1128/cmr.2.suppl.s41
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In Proteobacteria, the outer membrane protein TamA and the inner membrane-anchored protein TamB form the Translocation and Assembly Module (TAM) complex, which facilitates the transport of autotransporters, virulence factors, and likely lipids across the two membranes. In Bacteroidota, TamA is replaced by TamL, a TamA-like lipoprotein with a lipid modification at its N-terminus that likely anchors it to the outer membrane. This structural difference suggests that TamL may have a distinct function compared to TamA. However, the role of TAM in bacterial phyla other than Proteobacteria remains unexplored. Our study aimed to elucidate the function of TamL in Flavobacterium johnsoniae, an environmental Bacteroidota. Unlike its homologs in Proteobacteria, we found that TamL and TamB are essential in F. johnsoniae. Through genetic, phenotypic, proteomic, and lipidomic analyses, we show that TamL depletion severely compromises outer membrane integrity, as evidenced by reduced cell viability, altered cell shape, increased susceptibility to membrane-disrupting agents, and elevated levels of outer membrane lipoproteins. Notably, we did not observe an overall decrease in the levels of β-barrel outer membrane proteins, nor substantial alterations in outer membrane lipid composition. By pull-down assays, we found TamL co-purifying with TamB in F. johnsoniae, suggesting an interaction. Furthermore, we found that while TamL and TamB monocistronic genes are conserved among Bacteroidota, only some species encode multiple TamL, TamB and TamA proteins. To our knowledge, this study is the first to provide functional insights into a TAM subunit beyond Proteobacteria.
The human pathogen Neisseria gonorrhoeae produces an array of diseases ranging from urethritis to disseminated gonococcal infections. Early events in the establishment of infection involve interactions between N. gonorrhoeae and the mucosal epithelium, which leads to the local release of inflammatory mediators. Because of this, it is important to identify the bacterial virulence factors and host cell components that contribute to inflammation. Using a series of column chromatography steps, we purified a lipoprotein from N. gonorrhoeae strain F62 called Lip. This outer membrane antigen expresses a conserved epitope known as H.8, which is common to all pathogenic Neisseria species. We found the purified preparation of Lip to be a potent inflammatory mediator capable of inducing the release of the chemokine interleukin (IL)-8 and the cytokine IL-6 by immortalized human endocervical epithelial cells and the production of IL-8 and the activation of the transcription factor NF-kappaB by human embryonic kidney 293 (HEK) cells transfected with toll-like receptor (TLR) 2. Upon removal of Lip by immunoprecipitation, the ability of the H.8/Lip preparation to stimulate NF-kappaB activation was abolished. In addition to TLR2, the activation of NF-kappaB by H.8/Lip in HEK cells was enhanced upon coexpression of TLR1 but not TLR6. These observations provide evidence that Lip is capable of inducing the release of inflammatory mediators from epithelial cells in a TLR2-dependent manner.
OBJECTIVES: Multidrug efflux pumps are thought to be involved in mediating multidrug resistance in Pseudomonas aeruginosa. Here we aim to characterize hitherto uncharacterized multidrug efflux pumps from P. aeruginosa. MATERIALS AND METHODS: We isolated a mutant, YM442, which showed elevated resistance to several antimicrobial agents from P. aeruginosa YM44 lacking four major multidrug efflux pumps, MexAB, MexCD-OprJ, MexEF-OprN and MexXY. We cloned genes responsible for the resistance from chromosomal DNA of YM442 using YM44 as host. RESULTS: We designated the genes mexVW. Introduction of a recombinant plasmid pTAJ2 carrying the mexVW into YM44 cells conferred resistance to fluoroquinolones, tetracycline, chloramphenicol, erythromycin, ethidium bromide and acriflavine. Elevated ethidium bromide extrusion was observed with cells of YM442 and of YM44/pTAJ2. An outer membrane protein OprM was able to cooperate with MexVW. Elevated expression of the mexV gene was observed with YM442 compared with YM44. CONCLUSIONS: MexV (membrane fusion protein)-MexW (RND-type membrane protein)-OprM is a tripartite multidrug efflux pump. It is suggested that other outer membrane component(s) could cooperate with MexVW.