Peptide Mapping of Bacterial Fimbrial Epitopes Interacting with Pattern Recognition Receptors

George Hajishengallis,Pukar Ratti,Evlambia Harokopakis

doi:10.1074/jbc.m507326200

Abstract

The fimbriae of the oral pathogen Porphyromonas gingivalis induce Toll-like receptor 2 (TLR2)-dependent macrophage activation upon their recognition by CD14 and the beta(2) integrin CD11b/CD18. To map functional epitopes of fimbriae that interact with these pattern recognition receptors (PRRs), we examined 20 synthetic peptides covering the entire length of the 41-kDa fimbrillin subunit. Using direct or competitive inhibition assays for receptor binding or cell activation, the CD14 binding activity of fimbriae was localized to residues 69-90 and was essential for TLR2-dependent cytokine induction. The CD11b/CD18 binding activity of fimbriae was localized to two neighboring epitopes defined by residues 166-185 and 206-225. Unlike epitope 69-90 that constitutively bound CD14, the CD11b/CD18 binding activity of epitopes 166-185 and 206-225 was inducible by integrin activators. The CD11b/CD18 binding activity played a contributory role to TLR2-dependent induction of tumor necrosis factor-alpha by fimbriae but was involved in specific down-regulation of interleukin-12. Cell activation by a combination of fimbrillin peptides corresponding to the CD14 and CD11b/CD18 binding activities resulted in higher tumor necrosis factor-alpha responses than would be expected from a simply additive effect, attributable to CD14-dependent inside-out signaling leading to enhanced binding interactions with CD11b/CD18. These data suggest that P. gingivalis fimbriae display a modular structure that interacts through discrete epitopes and in a regulated mode with distinct PRRs, which in turn differentially modulate the state of cell activation. Elucidation of pathogen interactions with PRRs at the molecular level may glean insight into host defense mechanisms as well as into microbial strategies that subvert innate immunity.

Highlights

Monocytes/macrophages constitute a major source of TNF-␣ production, and their number increases in periodontal inflammation compared with healthy periodontal tissue [16, 17]
In our efforts to determine the pattern recognition receptors (PRRs) and the mechanisms involved in innate immune recognition of P. gingivalis fimbriae, we have found that CD14 directly binds fimbriae [22] and mediates their ability to stimulate Toll-like receptor 2 (TLR2)-dependent TNF-␣ release [14, 23]
In this study we mapped functional epitopes of the P. gingivalis fimbriae interacting with PRRs involved in monocyte/macrophage activation

Summary

Introduction

Monocytes/macrophages constitute a major source of TNF-␣ production, and their number increases in periodontal inflammation compared with healthy periodontal tissue [16, 17] These cells express multiple PRRs, including CD14 and CD11b/CD18, which play an important accessory role in Toll-like receptor (TLR)-dependent innate immune and inflammatory responses (18 –20). CD14 was found to be indispensable for cell activation by fimbrial peptide or intact protein, whereas fimbriae-CD11b/CD18 interactions are involved in both positive and negative regulation of innate immune responses. It appears, that P. gingivalis fimbriae interact through discrete domains with CD14 and CD11b/CD18, which in turn differentially modulate the activation state of monocytes/macrophages

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Conclusion