Interaction of Prevotella intermedia strain 17 leucine-rich repeat domain protein AdpF with eukaryotic cells promotes bacterial internalization.
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.
VCU Philips Institute for Oral Health Research, Virginia Commonwealth University, Richmond, Virginia, USA.
Dipanwita
D
Sengupta
Dae-Joong
DJ
Kang
Cecilia
C
Anaya-Bergman
Tiana
T
Wyant
Arnab K
AK
Ghosh
Hiroshi
H
Miyazaki
Janina P
JP
Lewis
D000704:Analysis of Variance; D001426:Bacterial Proteins / Q000502:physiology*; D005057:Eukaryotic Cells / Q000382:microbiology*; D005347:Fibroblasts / Q000382:microbiology; D005353:Fibronectins / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D006367:HeLa Cells / Q000382:microbiology; D006801:Humans; D000090263:Leucine-Rich Repeat Proteins; D018713:Prevotella intermedia / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology*; D011506:Proteins / Q000502:physiology*
D001426:Bacterial Proteins; D005353:Fibronectins; D000090263:Leucine-Rich Repeat Proteins; D011506:Proteins