Interaction of Prevotella intermedia Strain 17 Leucine-Rich Repeat Domain Protein AdpF with Eukaryotic Cells Promotes Bacterial Internalization

Abstract

Prevotella intermedia is an oral bacterium implicated in a variety of oral diseases. Although internalization of this bacterium by nonphagocytic host cells is well established, the molecular players mediating the process are not well known. Here, the properties of a leucine-rich repeat (LRR) domain protein, designated AdpF, are described. This protein contains a leucine-rich region composed of 663 amino acid residues, and molecular modeling shows that it folds into a classical curved solenoid structure. The cell surface localization of recombinant AdpF (rAdpF) was confirmed by electron and confocal microscopy analyses. The recombinant form of this protein bound fibronectin in a dose-dependent manner. Furthermore, the protein was internalized by host cells, with the majority of the process accomplished within 30 min. The internalization of rAdpF was inhibited by nystatin, cytochalasin, latrunculin, nocodazole, and wortmannin, indicating that microtubules, microfilaments, and signal transduction are required for the invasion. It is noteworthy that preincubation of eukaryotic cells with AdpF increased P. intermedia 17 internalization by 5-and 10-fold for HeLa and NIH 3T3 fibroblast cell lines, respectively. The addition of the rAdpF protein was also very effective in inducing bacterial internalization into the oral epithelial cell line HN4, as well as into primary cells, including human oral keratinocytes (HOKs) and human umbilical vein endothelial cells (HUVECs). Finally, cells exposed to P. intermedia 17 internalized the bacteria more readily upon reinfection. Taken together, our data demonstrate that rAdpF plays a role in the internalization of P. intermedia 17 by a variety of host cells.

Simultaneous Detection of Oral Pathogens in Subgingival Plaque and Prostatic Fluid of Men With Periodontal and Prostatic Diseases

“…Aggregatibacter actinomycetemcomitans (Aa) , which is associated with aggressive periodontitis, may also release soluble bacterial components 14 . Oral bacteria can directly invade endothelial cells without the aid of their byproducts 15 . Pg , which is a red‐complex bacteria and one of the main periodontal pathogens, 16 is able to invade the aorta via the lipid raft in human aortic cells 17 and can also move from one cell to another in cardiovascular endothelial and smooth muscle cells 18 .…”

The complete genome sequencing ofPrevotella intermediastrain OMA14 and a subsequent fine-scale, intra-species genomic comparison reveal an unusual amplification of conjugative and mobile transposons and identify a novelPrevotella-lineage-specific repeat

“…intermedia , such as proteolytic enzyme interpain A (II_0221) 12 and the adhesins AdpC (I_1236) 10 and AdpF (I_1057). 11 The gene set encoding the type IX secretion system (T9SS), which was originally identified in Por. gingivalis , 32 , 33 was also found in the core genome.…”

“…The virulence factors of Pre. intermedia , such as adhesins, 10 , 11 proteolytic enzymes 12 and lipopolysaccharides, 13 have been investigated, but its pathogenicity has not yet been fully elucidated. Three Pre.…”

<em>Porphyromonas gingivalis</em> as a Model Organism for Assessing Interaction of Anaerobic Bacteria with Host Cells

“…Thus, for large-scale quantitative data on the invasive property of bacteria a flow cytometry can also be used 40 . This method involves the use of fluorescently labeled bacteria (that can be accomplished as described for bacteria to be used for microscopy) and allows for studying multiple samples at a time that may be attractive for some researchers.…”

“…In summary, described are a few simple protocols that can be used to examine host-pathogen interaction for anaerobic bacteria. These protocols have also been used to assess the effect of bacterial internalins, proteins promoting bacterial invasion 40 , as well as surrogate non-invasive bacteria expressing a protein conferring invasive property on the bacteria 47 .…”