Dataset Information

Synthesis of Sphingolipids Impacts Survival of Porphyromonas gingivalis and the Presentation of Surface Polysaccharides.

ABSTRACT: Bacteria alter the biophysical properties of their membrane lipids in response to environmental cues, such as shifts in pH or temperature. In essence, lipid composition determines membrane structure, which in turn influences many basic functions, such as transport, secretion, and signaling. Like other members of the phylum Bacteroidetes, the oral anaerobe Porphyromonas gingivalis possesses the ability to synthesize a variety of novel membrane lipids, including species of dihydroceramides that are distinct, yet similar in structure to sphingolipids produced by the human host. The role of dihydroceramides in the physiology and pathogenic potential of the human microbiota is only beginning to be explored; yet there is increasing data indicating that these lipids play a role in human diseases, such as periodontitis and multiple sclerosis. Here, we report on the identification of a gene (PG1780) in the chromosome of P. gingivalis strain W83 encoding a putative serine palmitoyltransferase, the enzyme that catalyzes the first step in sphingolipid biosynthesis. While we were able to detect dihydroceramides in whole lipid extracts of P. gingivalis cells as well as crude preparations of outer membrane vesicles, sphingolipids were absent in the PG1780 mutant strain. Moreover, we show that the synthesis of sphingolipids plays an essential role in the long-term survival of the organism as well as its resistance to oxidative stress. Further, a PG1780 mutant displayed much lower activity of cell-associated arginine and lysine gingipains, yet slightly higher activity in the corresponding culture supernates, which we hypothesize is due to altered membrane properties and anchoring of these proteases to the cell surface. In addition, we determined that sphingolipid production is critical to the presentation of surface polysaccharides, with the mutant strain displaying less K-antigen capsule and more anionic polysaccharide (APS). Overall, we have discovered that, in addition to their role in pathogenicity, the synthesis of sphingolipids is critical to the cellular homeostasis and persistence of this important dental pathogen.

SUBMITTER: Moye ZD

PROVIDER: S-EPMC5126122 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Synthesis of Sphingolipids Impacts Survival of <i>Porphyromonas gingivalis</i> and the Presentation of Surface Polysaccharides.

Moye Zachary D ZD Valiuskyte Kornelija K Dewhirst Floyd E FE Nichols Frank C FC Davey Mary E ME

Frontiers in microbiology 20161129

Bacteria alter the biophysical properties of their membrane lipids in response to environmental cues, such as shifts in pH or temperature. In essence, lipid composition determines membrane structure, which in turn influences many basic functions, such as transport, secretion, and signaling. Like other members of the phylum Bacteroidetes, the oral anaerobe <i>Porphyromonas gingivalis</i> possesses the ability to synthesize a variety of novel membrane lipids, including species of dihydroceramides ...[more]

PMID: 27965646

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Project description:Studies are showing that the stress hormone cortisol can reach high levels in the gingival sulcus and induce shifts in the metatranscriptome of the oral microbiome. Interestingly, it has also been shown that cortisol can influence expression levels of Type IX Secretion System (T9SS) genes involved in gliding motility in bacteria belonging to the phylum Bacteroidota. The objective of this study was to determine if cortisol impacts gene expression and surface translocation of Porphyromonas gingivalis strain W50. To conduct these experiments, P. gingivalis was stabbed to the bottom of soft agar plates containing varying cortisol concentrations (0 μM, 0.13 μM, 1.3 μM, and 13 μM), and surface translocation on the subsurface was observed after 48 h of incubation. The results show that when grown with certain nutrients, i.e., in rich medium with the addition of sheep blood, lactate, or pyruvate, cortisol promotes migration of P. gingivalis in a concentration-dependent manner. To begin to examine the underlying mechanisms, quantitative PCR was used to evaluate differential expression of genes when P. gingivalis was exposed to cortisol. In particular, we focused on differential expression of T9SS-associated genes, including mfa5, since it was previously shown that Mfa5 is required for cell movement and cell-to-cell interactions. The data show that mfa5 is significantly up-regulated in the presence of cortisol. Moreover, an mfa5 deletion mutant showed less surface translocation compared to the wild-type P. gingivalis in the presence of cortisol, and the defects of the mfa5 deletion mutant were restored by complementation. Overall, cortisol can stimulate P. gingivalis surface translocation and this coincides with higher expression levels of T9SS-associated genes, which are known to be essential to gliding motility. Our findings support a high possibility that the stress hormone cortisol from the host can promote surface translocation and potentially virulence of P. gingivalis.

Project description:The asaccharolytic anaerobe Porphyromonas gingivalis metabolizes proteins it encounters in the periodontal pocket, including host-derived glycoproteins such as mucins and immunoglobulins. Often, these proteins are protected by a diverse array of carbohydrates tethered to the polypeptide chain via glycolytic bonds, and P. gingivalis produces enzymes capable of liberating these carbohydrates, exposing the proteinaceous core. In this study, we investigated the effect of individual monosaccharides, including galactose, l-fucose, mannose, and glucose, on the growth and physiology of P. gingivalis Of the carbohydrates tested, only galactose noticeably altered the density of the bacterial culture, and we observed that cultures grown with galactose reached significantly higher densities during stationary phase. Importantly, electron micrographs and plating of P. gingivalis in stationary phase demonstrated that the presence of galactose did not increase cell numbers; instead, the higher densities resulted from the expansion of individual cells which contained large intracellular granules. Initial attempts to characterize these granules revealed only a subtle increase in soluble carbohydrates, suggesting they are likely not composed of stored carbohydrate. Also, an analysis of major surface polysaccharides via an enzyme-linked immunosorbent assay (ELISA) did not reveal significant differences between cells grown with or without galactose. Finally, an initial investigation of the transcriptional changes elicited by galactose in late exponential phase suggested that genes important for cell shape and for the general stress response may play roles in this phenomenon. Overall, galactose, a monosaccharide commonly present on the surfaces of host proteins, substantially alters the physiology of P. gingivalis via the production of large, currently undefined, intracellular granules.IMPORTANCE Environmental perturbations are central to the ability of pathobionts, such as Porphyromonas gingivalis, to promote the development of diseased sites. In the case of periodontal disease, increased local pH, a shift to anaerobic surroundings, and the accumulation of Gram-negative anaerobes at the expense of Gram-positive cocci are known ecological fluctuations prominently associated with progression toward disease. Importantly, in contrast, the alterations to subgingival food webs in disease sites remain poorly characterized. We hypothesized that given the dramatic shift in community structure during disease, it is possible that free carbohydrates, which would typically be readily metabolized by Gram-positive cocci after cleavage from glycoproteins, may increase in concentration locally and thereby affect the physiological state of the subgingival microbiota. In this study, we explored the impact of free monosaccharides on P. gingivalis to gain deeper insight into the effect of dysbiotic conditions on the growth and physiology of this periodontal pathogen.

Dataset Information

Synthesis of Sphingolipids Impacts Survival of Porphyromonas gingivalis and the Presentation of Surface Polysaccharides.

Publications

Synthesis of Sphingolipids Impacts Survival of <i>Porphyromonas gingivalis</i> and the Presentation of Surface Polysaccharides.

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