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Porphyromonas gingivalis fimbriae induce a 68-kilodalton phosphorylated protein in macrophages
Y Murakami1, S Hanazawa, A Watanabe
1Department of Oral Microbiology, Meikai University School of Dentistry, Saitama, Japan.
Abstract:
The present study was performed to examine whether Porphyromonas gingivalis fimbriae induce specifically a protein kinase-mediated phosphorylated protein that is involved in the mechanism of signal transduction. The fimbriae induced a 68-kDa phosphorylated protein (pp68) in a dose-dependent manner in mouse peritoneal macrophages. A marked appearance of pp68 was observed 20 min after the initiation of fimbrial treatment. The fimbria-induced pp68 was inhibited dramatically by staurosporine, a potent inhibitor of protein kinase C. pp68 induction was also inhibited by H-7, a potent inhibitor of several types of protein kinase. However, the induction was not inhibited by HA-1004 and H-8, relatively high-affinity inhibitors of protein kinase A. Phorbol myristate acetate and 1-oleoyl-2-acetyl-sn-glycerol, activators of protein kinase C, were able to induce pp68 in mouse peritoneal macrophages. This protein was localized in the cytosolic fraction of fimbria-treated macrophages. pp68 also was induced in fimbria-treated human monocyte-like cells. Finally, we observed that gene expression of the fimbria-induced neutrophil chemoattractant KC was inhibited markedly by staurosporine.
Insights
Porphyromonas gingivalis fimbriae trigger a specific protein kinase pathway, leading to the induction of a 68-kDa phosphorylated protein (pp68) involved in cellular signaling. This finding sheds light on bacterial mechanisms in immune cell responses.
Area of Science:
- Immunology
- Molecular Biology
- Cellular Signaling
Background:
- Porphyromonas gingivalis is a key pathogen in periodontitis.
- Bacterial fimbriae are crucial virulence factors involved in host-pathogen interactions.
- Signal transduction pathways are essential for cellular responses to external stimuli.
Purpose of the Study:
- To investigate if Porphyromonas gingivalis fimbriae induce a specific protein kinase-mediated phosphorylation event.
- To identify the role of this phosphorylated protein in cellular signal transduction mechanisms.
Main Methods:
- Treatment of mouse peritoneal macrophages and human monocyte-like cells with Porphyromonas gingivalis fimbriae.
- Dose-dependent and time-course analysis of protein phosphorylation.
- Inhibition studies using specific protein kinase inhibitors (staurosporine, H-7, HA-1004, H-8).
- Activation studies using protein kinase C activators (phorbol myristate acetate, 1-oleoyl-2-acetyl-sn-glycerol).
- Subcellular localization of the phosphorylated protein.
- Analysis of neutrophil chemoattractant KC gene expression.
Main Results:
- Porphyromonas gingivalis fimbriae induced a 68-kDa phosphorylated protein (pp68) in a dose- and time-dependent manner.
- pp68 induction was significantly inhibited by staurosporine (protein kinase C inhibitor) and H-7, but not by HA-1004 or H-8 (protein kinase A inhibitors).
- Activators of protein kinase C also induced pp68.
- pp68 was localized in the cytosolic fraction.
- Fimbriae-induced expression of neutrophil chemoattractant KC was inhibited by staurosporine.
Conclusions:
- Porphyromonas gingivalis fimbriae activate a protein kinase C-mediated signaling pathway.
- This pathway results in the phosphorylation of a 68-kDa protein (pp68) crucial for signal transduction.
- The findings provide insights into the molecular mechanisms by which P. gingivalis fimbriae modulate host cell responses, including neutrophil recruitment.