Inactivation of Membrane Tumor Necrosis Factor Alpha by Gingipains fromPorphyromonas gingivalis

Abstract

Gingipains are cysteine proteinases produced by Porphyromonas gingivalis, a major causative bacterium of adult periodontitis. They consist of arginine-specific (HRgpA and RgpB) and lysine-specific (Kgp) proteinases. Gingipains strongly affect the host defense system by degrading some cytokines, components of the complement system, and several immune cell receptors. In an in vitro model, gingipains were shown to degrade soluble tumor necrosis factor alpha (TNF-␣). However, since membrane TNF-␣ shows strong biological activity, especially in local inflammatory lesions, it was worth investigating whether gingipains might also destroy membrane TNF-␣ and limit its biological activities. To avoid a possible influence of gingipains on ADAM17, the secretase of TNF-␣, the majority of experiments were performed using ADAM17 ؊/؊ fibroblasts stably transfected with cDNA of human pro-TNF-␣ (ADAM17 ؊/؊ TNF ؉ ). Arginine-specific gingipains (Rgp's) strongly diminished the level of TNF-␣ on the cell surface as measured by flow cytometry, and this process was not accompanied by an increased concentration of soluble TNF-␣ in the culture medium. Degradation of membrane TNF-␣ by Rgp's correlated with a strong decrease in TNF-␣-mediated biological activities of ADAM17 ؊/؊ TNF ؉ cells. First, the activation state of transcription factor NF-B was suppressed; second, the cells were no longer able to induce apoptosis in HL-60 cells. Kgp was also able to cleave membrane TNF-␣, but its effect was much weaker than that of Rgp's. Gingipains also limited the binding of native TNF-␣ to the target cells. Thus, gingipains are able not only to cleave soluble TNF-␣ but also to destroy the membrane form of the cytokine, which may additionally dysregulate the cytokine network.

Gingipain-dependent interactions with the host are important for survival of Porphyromonas gingivalis

“…This suggests that BCAEC are in some way able to protect Ncadherin from cleavage by Kgp activity at later time points in order to counteract the apoptotic signaling and/or stimulate survival signaling. A similar phenomenon was observed in the gingipain-mediated cleavage of TNF-α in fibroblasts, in which HRgpA was shown to have the most activity toward TNF-α, but Kgp-induced cleavage of TNF-α was greatly increased if TNF-α re-expression was prevented, suggesting that the loss of TNF-α through Kgp cleavage could be compensated for by de novo synthesis (128), allowing cells to offset the Kgp-induced effects. Moreover, we have observed that a high concentration of Kgp can cause BCAEC detachment and apoptosis (our unpublished observations) suggesting that there is a threshold level of gingipain activity below which a cell can remain viable, but at gingipain activity above this level the cell cannot overcome gingipain-induced effects and becomes apoptotic.…”

The Native 67-Kilodalton Minor Fimbria of Porphyromonas gingivalis Is a Novel Glycoprotein with DC-SIGN-Targeting Motifs

“…P. gingivalis utilizes a myriad of virulence factors that contribute to chronic periodontitis. Among these are a polysaccharide capsule, fimbriae, proteases for opsonins C3 and IgG, gingipains (21,30,43,52), bacterial lipopolysaccharides (LPS) (22,44), and toxins and hemagglutinins (10,25).…”

Tumor necrosis factor-α converting enzyme (TACE) increases RANKL expression in osteoblasts and serves as a potential biomarker of periodontitis

“…Periodontitis is one of the most common inflammatory diseases and is characterized by the presence of (mainly gram-negative) anaerobes, accumulation of immune cells, formation of periodontal pockets, and loss of tooth attachment (1,2). Recent studies have investigated the roles of cytokines and secreted proteinases in periodontitis (3).…”