Aggregation of human platelets by gingipain-R from Porphyromonas gingivalis cells and membrane vesicles

Pham, Khoa; Feik, Diane; Hammond, Benjamin F.; Rams, Thomas E.; Whitaker, Eugene J.

doi:10.1080/09537100120104863

Cited by 41 publications

(33 citation statements)

References 33 publications

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“…Previous reports have shown that MVs possess various virulent abilities (9,13,25,39,40,43,48), while our results indicate a new aspect of cellular function impairment by P. gingivalis MVs.…”

Section: Discussionsupporting

confidence: 63%

Entry ofPorphyromonas gingivalisOuter Membrane Vesicles into Epithelial Cells Causes Cellular Functional Impairment

2009

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Bacteria have evolved mechanisms for the secretion of virulence factors into host cells; these virulence factors alter host cell biology and enable bacterial colonization (11). Bacterial outer membrane vesicles (MVs), ubiquitously shed from gramnegative bacteria by a mechanism involving cell wall turnover, consist of a subset of outer membrane and soluble periplasmic components (54). This extracellular secretion system likely plays a part in the strategy utilized by bacterial pathogens to modulate host defense and response and impair host cell function. For example, Pseudomonas aeruginosa (3), Helicobacter pylori (19), and Actinobacillus actinomycetemcomitans (8), as well as pathogenic and nonpathogenic Escherichia coli (7, 51), secrete MVs that contain toxins, proteases, adhesins, and lipopolysaccharide. Therefore, it has been proposed that MVs are bacterial "bombs" (30). However, the molecular mechanism of MV entry into host cells is unclear, while it also remains unknown whether MV-associated virulence factors have cytotoxic effects within the invaded cells. A recent study showed that enterotoxigenic E. coli MVs containing heat-labile enterotoxin and other bacterial envelope components were taken up by a human epithelial cell line via cellular lipid rafts, after which intracellular MVs accumulated in nonacidified compartments inaccessible to the extracellular milieu (28). In addition, a very recent study found that Pseudomonas aeruginosa MVs deliver multiple virulence factors, including ␤-lactamase, alkaline phosphatase, hemolytic phospholipase C, and Cif (cystic fibrosis transmembrane conductance regulator inhibitory factor), directly into the host cytoplasm via fusion of MVs with lipid rafts in the host plasma membrane, which has an effect on host cell biology (5). Thus, it is likely MVs function as specifically targeted transport vehicles to mediate entry of bacterial virulence factors into host cells. However, no other related results have been reported.Periodontitis, one of the most common infectious diseases seen in humans (52), is characterized by gingival inflammation, as well as loss of connective tissue and bone from around the roots of the teeth, which leads to eventual tooth exfoliation. Porphyromonas gingivalis is considered to be a bona fide pathogen that causes several forms of severe periodontal disease. The bacterium releases MVs in an extracellular manner; these MVs retain the full components of outer membrane constituents, including lipopolysaccharide, muramic acid, a capsule, fimbriae, and proteases termed gingipains (13,35). Fimbriae reportedly mediate bacterial adherence to and entry into periodontal cells (2), while gingipains, which consist of arginine (Arg-gingipain [Rgp])-and lysine (Lys-gingipain [Kgp])-specific cysteine proteinases, contribute to the destruction of periodontal tissues (24). Gingipains degrade collagen and fibronectin and inhibit interactions between host cells and the extracellular matrix. In addition, they degrade various cytokines, resulting in a disturbance of t...

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Section: Discussionsupporting

confidence: 63%

Entry ofPorphyromonas gingivalisOuter Membrane Vesicles into Epithelial Cells Causes Cellular Functional Impairment

2009

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“…In addition, MVs have been reported to possess various virulent abilities (9,13,21,39,41,47,51). We intend to conduct additional studies to elucidate the cellular functions impaired by invading MVs.…”

Section: Discussionmentioning

confidence: 99%

Porphyromonas gingivalis Outer Membrane Vesicles Enter Human Epithelial Cells via an Endocytic Pathway and Are Sorted to Lysosomal Compartments

et al. 2009

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Porphyromonas gingivalis, a periodontal pathogen, secretes outer membrane vesicles (MVs) that contain major virulence factors, including major fimbriae and proteases termed gingipains, although it is not confirmed whether MVs enter host cells. In this study, we analyzed the mechanisms involved in the interactions of P. gingivalis MVs with human epithelial cells. Our results showed that MVs swiftly adhered to HeLa and immortalized human gingival epithelial cells in a fimbria-dependent manner and then entered via a lipid raft-dependent endocytic pathway. The intracellular MVs were subsequently routed to early endosome antigen 1-associated compartments and then were sorted to lysosomal compartments within 90 min, suggesting that intracellular MVs were ultimately degraded by the cellular digestive machinery. However, P. gingivalis MVs remained there for over 24 h and significantly induced acidified compartment formation after being taken up by the cellular digestive machinery. In addition, MV entry was shown to be mediated by a novel pathway for transmission of bacterial products into host cells, a Rac1-regulated pinocytic pathway that is independent of caveolin, dynamin, and clathrin. Our findings indicate that P. gingivalis MVs efficiently enter host cells via an endocytic pathway and survive within the endocyte organelles for an extended period, which provides better understanding of the role of MVs in the etiology of periodontitis.

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“…This commensal can colonize, invade and multiply within gingival epithelial cells, as well as penetrate into deeper epithelial cell layers, potentially releasing the whole organism and/or virulence factors into the bloodstream (reviewed by Yilmaz, 2008). In addition to its ability to cause disease in the oral cavity, there are data indicating a role in systemic disease, including its ability to invade vascular endothelial cells (Dorn et al, 2000(Dorn et al, , 2002Jandik et al, 2008) and to cause aggregation of platelets (Pham et al, 2002). For many pathogenic bacteria, surface polysaccharides play a key role in immune modulation and evasion (Comstock & Kasper, 2006).…”

Section: Introductionmentioning

confidence: 99%

The nucleoid-associated protein HUβ affects global gene expression in Porphyromonas gingivalis

et al. 2013

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HU is a non-sequence-specific DNA-binding protein and one of the most abundant nucleoidassociated proteins in the bacterial cell. Like Escherichia coli, the genome of Porphyromonas gingivalis is predicted to encode both the HUa (PG1258) and the HUb (PG0121) subunit. We have previously reported that PG0121 encodes a non-specific DNA-binding protein and that PG0121 is co-transcribed with the K-antigen capsule synthesis operon. We also reported that deletion of PG0121 resulted in downregulation of capsule operon expression and produced a P. gingivalis strain that is phenotypically deficient in surface polysaccharide production. Here, we show through complementation experiments in an E. coli MG1655 hupAB double mutant strain that PG0121 encodes a functional HU homologue. Microarray and quantitative RT-PCR analysis were used to further investigate global transcriptional regulation by HUb using comparative expression profiling of the PG0121 (HUb) mutant strain to the parent strain, W83. Our analysis determined that expression of genes encoding proteins involved in a variety of biological functions, including iron acquisition, cell division and translation, as well as a number of predicted nucleoid associated proteins were altered in the PG0121 mutant. Phenotypic and quantitative real-time-PCR (qRT-PCR) analyses determined that under iron-limiting growth conditions, cell division and viability were defective in the PG0121 mutant. Collectively, our studies show that PG0121 does indeed encode a functional HU homologue, and HUb has global regulatory functions in P. gingivalis; it affects not only production of capsular polysaccharides but also expression of genes involved in basic functions, such as cell wall synthesis, cell division and iron uptake. INTRODUCTIONPorphyromonas gingivalis is a Gram-negative obligate anaerobe belonging to the family Bacteroidaceae that persists as a natural member of the human oral microbiota. A shift in the microbial community leading to outgrowth of this anaerobe is directly linked to periodontitis, a chronic inflammatory disease that leads to destruction of the tissues supporting the gums and ultimately, exfoliation of the teeth (Choil et al., 1990;Dzink et al., 1988;Grossi et al., 1994;Lamont & Jenkinson, 2000;Moore et al., 1991). This commensal can colonize, invade and multiply within gingival epithelial cells, as well as penetrate into deeper epithelial cell layers, potentially releasing the whole organism and/or virulence factors into the bloodstream (reviewed by Yilmaz, 2008). In addition to its ability to cause disease in the oral cavity, there are data indicating a role in systemic disease, including its ability to invade vascular endothelial cells (Dorn et al., 2000(Dorn et al., , 2002Jandik et al., 2008) and to cause aggregation of platelets (Pham et al., 2002). For many pathogenic bacteria, surface polysaccharides play a key role in immune modulation et al., 1996). HU typically acts as an accessory protein in virtually all types of nucleoprotein-mediated processes (reviewed by...

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Aggregation of human platelets by gingipain-R from Porphyromonas gingivalis cells and membrane vesicles

Cited by 41 publications

References 33 publications

Entry ofPorphyromonas gingivalisOuter Membrane Vesicles into Epithelial Cells Causes Cellular Functional Impairment

Entry ofPorphyromonas gingivalisOuter Membrane Vesicles into Epithelial Cells Causes Cellular Functional Impairment

Porphyromonas gingivalis Outer Membrane Vesicles Enter Human Epithelial Cells via an Endocytic Pathway and Are Sorted to Lysosomal Compartments

The nucleoid-associated protein HUβ affects global gene expression in Porphyromonas gingivalis

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