Proteolysis of human monocyte CD14 by cysteine proteinases (gingipains) from Porphyromonas gingivalis leading to lipopolysaccharide hyporesponsiveness.
Cysteine proteinases (gingipains) elaborated from Porphyromonas gingivalis exhibit enzymatic activities against a broad range of host proteins and are considered key virulence factors in the onset and development of adult periodontitis and host defense evasion. In this study, we examined the ability of arginine-specific gingipains (high molecular mass Arg-specific gingipain (HRGP) and Arg-specific gingipain 2) and lysine-specific gingipain (KGP) to cleave monocyte CD14, the main receptor for bacterial cell surface components such as LPS. Binding of anti-CD14 mAb MY4 to human monocytes was almost completely abolished by 0.3 microM HRGP and KGP treatments for 15 min, and 1 microM RGP2 for 30 min. In contrast, the expressions of Toll-like receptor 4, and CD18, CD54, CD59, and HLA-A, -B, -C on monocytes were slightly increased and decreased, respectively, by 0. 3 microM HRGP and KGP. This down-regulation resulted from direct proteolysis, because 1) gingipains eliminated MY4 binding even to fixed monocytes, and 2) CD14 fragments were detected in the extracellular medium by immunoblot analysis. Human rCD14 was degraded by all three gingipains, which confirmed that CD14 was a substrate for gingipains. TNF-alpha production by monocytes after HRGP and KGP treatments was decreased at 1 ng/ml, but not at 20 microg/ml LPS, indicating that gingipains inhibited a CD14-dependent cell activation. These results suggest that gingipains preferentially cleave monocyte CD14, resulting in attenuation of the cellular recognition of bacteria, and as a consequence sustain chronic inflammation.
Sugawara
S
S
Department of Microbiology and Immunology, Tohoku University School of Dentistry, Sendai, Japan. sugawars@mail.cc.tohoku.ac.jp
Nemoto
E
E
Tada
H
H
Miyake
K
K
Imamura
T
T
Takada
H
H
D018829:Adhesins, Bacterial; D000328:Adult; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D001666:Binding Sites, Antibody / Q000187:drug effects; D001667:Binding, Competitive / Q000276:immunology; D001769:Blood / Q000276:immunology; D003546:Cysteine Endopeptidases / Q000378:metabolism* / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004151:Dipeptides / Q000494:pharmacology; D005404:Fixatives / Q000494:pharmacology; D005557:Formaldehyde / Q000494:pharmacology; D000080867:Gingipain Cysteine Endopeptidases; D006388:Hemagglutinins / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D006868:Hydrolysis; D007108:Immune Tolerance / Q000187:drug effects; D015151:Immunoblotting; D007527:Isoenzymes / Q000378:metabolism / Q000494:pharmacology; D007659:Ketones / Q000494:pharmacology; D018950:Lipopolysaccharide Receptors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000276:immunology*; D009000:Monocytes / Q000201:enzymology* / Q000276:immunology* / Q000378:metabolism; D011108:Polymers / Q000494:pharmacology; D016966:Porphyromonas gingivalis / Q000201:enzymology*; D011994:Recombinant Proteins / Q000378:metabolism; D012995:Solubility; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors / Q000096:biosynthesis
D018829:Adhesins, Bacterial; D000590:Amino Acid Chloromethyl Ketones; D015853:Cysteine Proteinase Inhibitors; D004151:Dipeptides; D005404:Fixatives; D000080867:Gingipain Cysteine Endopeptidases; D006388:Hemagglutinins; D007527:Isoenzymes; D007659:Ketones; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D011108:Polymers; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; C091190:benzyloxycarbonyl-phenylalanyl-lysyl-2,4,6-trimethylbenzoyloxymethyl ketone; D005557:Formaldehyde; D003546:Cysteine Endopeptidases; C040308:phenylalanyl-prolyl-arginine-chloromethyl ketone; C003043:paraform